Role of sRNA molecules in the rye–leaf rust interaction

Role of sRNA molecules in the rye–leaf rust interaction | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Role of sRNA molecules in the rye–leaf rust interaction Marek Daniel Koter, Mateusz Matuszkiewicz, Monika Rakoczy-Trojanowska This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-3875675/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Leaf rust (LR) caused by Puccinia recondita f. sp. secalis ( Prs ) is one of the most damaging diseases of rye. However, the genetic basis of the rye response to LR remains relatively unknown. Specifically, the involvement of small RNAs (sRNAs) in the rye– Prs interaction has not been characterized. In this study, the changes in various sRNAs in response to LR were revealed in three rye lines (D33, D39, and L318) that vary regarding their immune responses to LR and two Prs isolates that cause compatible (CP) or incompatible (ICP) interactions with the host. The sRNAs were analyzed using miRDeep2 and ShortStack algorithms, with true miRNAs detected on the basis of strict miRNA biosynthesis-related parameters. The differential expression of all sRNAs and miRNAs was analyzed and potential targets (i.e., mRNA) were detected. Additionally, the targets in the CP and ICP host–pathogen interactions were compared. Moreover, different isoforms of miRNAs originating from the same miRNA gene (isomiRs) were manually identified and the diversity in their potential targets were examined. The isomiRs can broaden the range of targets related to plant defense responses to pathogens. For example, UGUGUUCUCAGGUCGCCCCCG targets the transcripts of 11 endoglucanase genes and two superoxide dismutase genes, which encode proteins with plant defense-related functions. A shorter sequence (UCAGGUCGCCCCCGCUGGAG) was revealed to target 16 mRNAs encoding transcription factors, including those from the MADS box family. Finally, miRNA-like RNAs (milRNAs) from Prs and their predicted targets in rye were identified. To examine the effects of the experimental design, the differences in the number of milRNAs between CP and ICP interactions as well as the milRNA expression dynamics at two time-points were analyzed. Compared with the ICP interactions, the CP interactions resulted in more milRNAs, which affected many more genes, including some related to biotic stress responses. Molecular Genetics sRNA leaf rust isomiR milRNA ck-RNAi Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 1. Introduction Despite being considered a cereal that is highly resistant to various environmental stresses, rye ( Secale cereale L.) is susceptible to leaf rust (LR), which can lead to yield losses of up to 40%. Leaf rust is caused by an airborne pathogen, namely the obligate biotrophic basidiomycete P. f. sp. secalis ( Prs ) [ 1 ]. The genetic basis of the interaction between rye and the LR pathogen has been studied for over 20 years. A total of 17 resistance genes ( Pr1–5 , Pr-d–f , Pr-i–l , Pr-n , Pr-p , Pr-r , and Pr-t ) located on chromosomes 1R, 2R, 4R, 6R, and 7R have been identified in studies that applied a Mendelian genetics approach [ 2 , 3 ]. The recent sequencing of the rye genome allowed for increasingly precise analyses at the molecular level, resulting in the identification of the genes responsive to LR [ 4 ]. Additionally, a recent genome-wide association study mapped five LR resistance-associated quantitative trait loci on chromosome arms 1RS (short arm of chromosome 1R), 1RL (long arm of chromosome 1R), 2RL, 5RL, and 7RS [ 5 , 6 ]. The most important resistance-associated marker on chromosome arm 1RS was physically co-localized with molecular markers delimiting Pr3 , which is a previously characterized gene. The second region on 7RS comprised numerous nucleotide-binding leucine-rich repeat (NLR)-encoding genes, one of which, provisionally denoted Pr6 , was similar (at the protein level) to the wheat LR response-related Lr1 gene on chromosome 5DL. Krępski et al. (2023) recently showed that several rye orthologs of wheat Lr genes ( ScLr1_3 , ScLr1_4 , ScLr1_8 , and ScRga2_6 ) may be critical for the rye response to LR [ 7 ]. The expression of many plant genes is regulated by small RNAs (sRNAs), which are non-coding sequences consisting of 20–32 nucleotides (nt). The sRNAs have been divided into a few main classes according to the differences in their biogenesis and function. One of the main sRNA classes includes short interfering RNAs (siRNAs) and microRNAs (miRNAs). These molecules can bind to specific proteins in the RNA-induced silencing complex, which then recognizes and targets complementary mRNA sequences, leading to translational inhibition or transcript degradation [ 8 ]. The results of an analysis of miRNAs classified according to new biosynthesis-based criteria have been published [ 9 ]. The large-scale miRNome sequencing of different plant and animal species has revealed distinct miRNA sequences originating from a single miRNA locus. These isoforms, which are more frequently referred to as isomiRs, represent variants with heterogenous ends, sequences, and lengths [ 10 – 12 ]. IsomiRs are variants of canonical miRNAs that differ in their length or sequence. They can be classified into two groups: templated isomiRs, which are encoded in their respective genes and result from alternative DCL cleavage, and non-templated isomiRs, which are modified by enzymes after transcription and may have nucleotide additions, deletions, or substitutions. Moreover, isomiRs originating from the same gene may be active in different plant tissues or developmental stages [ 13 ]. Studies on miRNA expression revealed fold-change dynamics, where the addition or loss of two or three nucleotides may result in completely different expression profiles [ 14 ]. Relatively little is known about the role of isomiRs in biotic stress responses. Recent studies have confirmed the distinct differential expression of individual isomiRs from a single sequence during the complex response of tomato to the potato cyst nematode [ 15 ]. Small RNAs also exist in fungi. They were first detected in the filamentous fungus Neurospora crassa in a study in which the transformation with homologous sequences led to the transient inactivation of endogenous transcripts [ 16 ]. Earlier research on tomato infected with Botrytis cinerea identified fungal miRNA-like RNAs (milRNAs) targeting the host transcripts of many genes related to pathogen resistance [ 17 ]. Since then, milRNAs have been identified in many fungal species, including plant parasites [ 18 , 19 , 20 ]. During an infection, the sRNA from the host plant can enter the cells of the pathogen and silence the pathogen genes and vice versa [ 19 , 21 , 22 ]. Small RNAs are a component of the fungal effectome [ 23 ]. Several researchers have explored the interactions between the sRNAs of different Puccinia fungal species and defense-related plant genes. For example, Mueth et al. (2015) identified Puccinia striiformis RNAs and predicted the host target genes [ 22 ]. Of the 22 target genes homologous to known plant stress resistance genes, seven encode nucleotide-binding site leucine-rich repeat (NBS-LRR) proteins and one encodes an ATP-binding cassette (ABC) transporter C family member. Wang et al. (2017) subsequently identified a novel sRNA (Pst-milR1) produced by P. striiformis f. sp. tritici infecting wheat [ 19 ]. Because the transcript of the pathogenesis-related gene Pr2 is targeted by Pst-milR1, the authors concluded that Pst-milR1 functions as an effector that suppresses the wheat immune response to stripe rust. Studies conducted over the last 10 years have shown that miRNAs induced by pathogens can be effective regulators of plant resistance ( R ) genes encoding diverse proteins, including the following: NBS-LRR proteins; pathogenesis-related proteins; proteins associated with amino acid metabolism, replication, transport, and chelation of reactive oxygen species; cysteine-rich receptor-like protein kinases; ABC transporters; and other proteins related to plant immune responses [ 17 , 21 , 24 – 28 ]. Many R genes expressed specifically in response to rusts have recently been identified in wheat and rye [ 29 , 30 ]. In terms of rust fungi, miRNAs regulate the defense responses of wheat infected with Puccinia graminis f. sp. tritici , which causes stem rust [ 24 ]. The authors suggested that different miRNAs might independently regulate the response of infected plants. In another study, Kumar et al. (2014) analyzed wheat inoculated with Puccinia triticina (causing wheat LR) and detected 52 miRNAs, of which 22 were differentially expressed between the susceptible and resistant near-isogenic lines (NILs) [ 21 ]. Additionally, there were more upregulated miRNAs in the susceptible NILs than in the resistant NILs. The authors identified 1,306 potential target genes, including genes encoding universal stress-responsive proteins (e.g., Pr1) or ABC transporters. However, there were no NLR-encoding genes targeted by miRNAs. In a later study, Kumar et al. (2017) identified 1,056 mature miRNAs (including 559 novel miRNAs) [ 31 ]. Following the treatment with the pathogen, there were more miRNAs in the resistant plants than in the susceptible plants. However, the miRNA counts increased in the susceptible line, but decreased in the resistant line. The degradome analysis identified 701 predicted target genes associated with defense responses, signal transduction, development, metabolism, and transcriptional regulation. Interestingly, Feng et al. (2016) detected 1,316 miRNA candidates activated by P. striiformis f. sp. tritici , which is responsible for yellow rust [ 32 ]. For the upregulated miRNAs, the authors detected ata-miR1432-5p-targeted genes encoding proteins from the calcium ion-binding family and PC-3p-7484 (wheat specific miRNA)-targeted genes encoding various proteins (e.g., ubiquillin, mitochondrial-processing peptidase, tubby-like F-box protein, and other hypothetical proteins) [ 32 ]. Nair et al. (2022) identified multiple miRNAs associated with Sr36 and Lr45 , which confer resistance to stem rust and LR, respectively [ 28 ]. The mining for miRNAs revealed 26 known miRNAs and seven novel miRNAs in cultivars infected with stem rust as well as 22 known miRNAs and four novel miRNAs in LR-infected samples. The expression of various miRNAs originating from different subgenomes may reflect a potential relationship between the mechanisms underlying the defense responses to stem rust and LR mediated by specific R genes. In plants, isomiRs are prospective host factors responsive to pathogen regulators. Specifically, they target genes involved in developmental processes, such as leucine catabolism, and genes encoding proteins that protect against stress conditions (e.g., GST). One of the isomiRs (isomiR523-4) detected in an earlier study reportedly targets Puccinia mRNAs from the SunT superfamily, which is associated with defense responses [ 25 ]. There is considerable information regarding the effects of rust infections on sRNAs during plant–pathogen interactions, but there has been relatively little related research on rye. 2. Materials and Methods 2.1. Plant materials, growth conditions, and treatment with the rust pathogen This study was conducted using the following three rye inbred lines: L318, which was bred by the Department of Plant Genetics, Breeding and Biotechnology, Warsaw University of Life Sciences (Warsaw, Poland), and D33 and D39, which were developed by Danko Plant Breeding, Ltd. (Poland). The inbred lines were selected according to the previously reported benzoxazinoid (BX) contents of field-grown plants after a natural vernalization period [ 33 ] and differential seedling stage responses to an artificial infection with a partially compatible Prs isolate [ 34 ]. Four Prs isolates (1/1.6, 83/2/2.2_5x, 81/r/5_5x, and 88/o/1_5x) that differed in terms of their interactions with host plants were used to inoculate lines D33, D39, and L318 (Table 1 ). Table 1 Interactions between rye inbred lines and Prs isolates Prs isolate Rye line 1/1.6 83/2/2.2_5x 81/r/5_5x 88/o/1_5x D33 incompatible compatible D39 incompatible compatible L318 incompatible compatible Rye seeds were sown on moistened tissue paper in Petri dishes, which were incubated in a darkened room for 2 days at 22°C. The germinated seeds were transferred to plastic pots (12 cm diameter) filled with sterilized peat substrate (10 germinated seeds per pot). The seedlings were cultivated for 10 days in a growth chamber at 22°C with a 16-h light (60 µmol m − 2 s − 1 )/8-h dark cycle. At 12 days after sowing, the plants were inoculated with a Prs spore suspension or the control solution. The plants were then covered with black boxes to maintain darkness and 100% humidity and incubated for 24 h at 18°C. They were subsequently transferred to a phytotron with a 16 h (22°C)/8 h (18°C) cycle. For the treatment, spores from a single-spore Prs isolate were suspended in Novec™ 7100 engineered fluid (3M, Maplewood, USA) at a density of 1 mg/ml. The control (mock-inoculated) plants were inoculated with Novec™ 7100 engineered fluid lacking Prs spores. The treatments were performed using leaf glass diffusers (Carl Roth, Karlsruhe, Germany). For each pot, leaves were collected from five plants at 20 hpt and from the other five plants at 36 hpt. All leaf samples were collected in tubes, frozen immediately in liquid nitrogen, and stored at − 80°C. The leaves were weighed and then 100 mg leaf material was ground in a mortar for the RNA extraction using the mirVana™ miRNA Isolation Kit (ThermoFisher, Waltham, USA). The RNA condition and concentration were determined using the Qubit 4 fluorometer (ThermoFisher) prior to the next-generation sequencing (NGS) analysis, which was completed by Genomed S.A. (Warsaw, Poland) as previously described [ 33 ]. 2.2. Bioinformatics analyses Sequencing data in FASTQ files were checked for quality using FastQC (version 0.11.8), while sequencing adapters were trimmed using Cutadapt (version 3.0) [ 35 , 36 ]. The reads were mapped to the rye genome with Bowtie (version 1.3) [ 37 ]. In the next step, miRDeep2 (version 0.1.3) was used to detect known miRNAs (i.e., deposited in miRbase) and novel miRNAs [ 38 ]. The quantitative analysis was performed using the DESeq2 package (version 1.32.0) from the Bioconductor repository [ 39 ]. The ShortStack software (version 3.5.8) was used to separate miRNAs from among the sRNAs according to the latest guidelines [ 9 , 40 ]. The sRNAs were quantitatively analyzed using the edgeR package (version 3.34.1) from the Bioconductor repository [ 41 ]. The isomiRs were detected manually using miRNA sequences aligned by Bowtie (version 1.3). The Puccinia -specific milRNAs were selected as follows. Sequencing reads from the mock-inoculated and Prs -infected rye leaves were mapped to the Prs genome as well as to the rye Lo7 genome. The Venn algorithm was used to select Prs -specific sRNA sequences. The detected miRNAs were compared with those in the miRBase and PmiREN 2.0 databases [ 42 , 43 ]. The psRNATarget web server was used to identify potential sRNA targets; the default parameters were applied, with the exception of maximum expectation, which was set to 3 [ 44 , 45 ]. For the functional enrichment analysis, the standard parameters of g:Profiler were used [ 46 , 47 ]. 3. Results 3.1. Identification of sRNAs in rye A comprehensive sRNA analysis pipeline (Fig. 1 ) was developed to identify sRNAs in rye. The Illumina sRNA sequencing generated 642,040,931 reads, of which 12.01% (77,123,956) were unique reads 18–35 nt long (Fig. 2 ). The most common read length was 32 nt, but the most diverse sRNAs consisted of 24 nt. The fact the reads consisting of 32 nt were the most abundant was likely due to the tRFs/tsRNAs, which are regulatory RNAs originating from tRNA segments. The sequences comprising 21–24 nt, which were mostly miRNAs and other sRNAs, were also over-represented. The rye sRNAs were recently surveyed in the Lo7 sequencing project, during which in-house scripts were used for identifying and annotating 1,005 different sRNA sequences [ 4 ]. In the current study, two algorithms in the miRDeep2 and ShortStack programs detected 65 and 462 sRNAs, respectively. Next, the sRNAs were compared with the Lo7 sRNA sequences and with all other sRNAs in the miRBase and PmiREN databases. A Venn diagram was constructed to separate experiment- and database-specific sRNA sequences (Fig. 3 ). In total, 43 and 390 novel sRNA sequences were detected using miRDeep2 and ShortStack, respectively. 3.2. Detecting differentially expressed sRNAs at different time-points during the rye–rust interaction This study initially focused on the dynamic expression of sRNAs in three rye lines infected with LR. Two algorithms detected 316 sequences, with miRDeep2 and ShortStack detecting 14 and 302 sequences, respectively. The subsequent analysis focused on the differentially expressed sRNAs and whether they decrease or increase the expression of genes important for plant defenses against fungal pathogens. The following threshold was applied for the expression analysis: |log 2 (fold change)| >1 (Fig. 4 ). Next, the compatible (CP) and incompatible (ICP) host–pathogen interactions were analyzed and compared to detect the sRNAs influencing gene expression during these two interactions. For the CP interaction, there were 225 sRNAs and seven miRNAs, whereas for the ICP interaction, there were only 57 sRNAs and one miRNA. Notably, 12 sRNAs were common to both interactions. The sRNA expression dynamics varied among the time-points, especially at 20 and 36 h post-treatment (hpt). During this period, the number of differentially expressed sRNAs increased by 84 (from 104 to 188), while the number of differentially expressed miRNAs decreased from five to two. Additionally, similar to the previous findings, there were several sequences that were common between the two time-points, including two sRNAs and two miRNAs (Fig. 5 A and B). 3.3. Host isomiRs responsive to the pathogen Differentially expressed miRNAs and their highly expressed cognate sequences (isomiRs) were manually selected (Fig. 7 A). They differed regarding their sequences as well as in terms of the number of specific target sequences (i.e., 10, 10, 19, and 72; Fig. 7 B). The differences in the isomiR sequences influence target recognition, thereby increasing the range of targeted transcripts important for plant defenses against pathogens. 3.4. Predicted targets for the rye sRNAs The psRNATarget tool and the rye Lo7 transcriptome were used for predicting sRNA targets. The resulting hits were enriched among the highly expressed |log 2 (fold change)| > 2and strongly expressed (baseMean 50) transcripts (Table 1 ). Multiple targets with known potential roles in host defenses against rust infections were detected (Table S1), including targets encoding NBS-LRR disease resistance proteins, receptor kinases, enzymes associated with sugar transport and metabolism, and ABC transporters. More specifically, some of the targets encoded a glutamate receptor, which mediate electrical signaling in response to fungal infections or inhibit fungal growth [ 48 ]. To highlight the functional diversity among the predicted targets of the rye sRNAs, we performed a gene functional enrichment analysis using g:profiler [ 46 ] (Supplementary Table S1) Table 2. Selected sRNAs and their highly expressed targets or targets expressed in all three rye lines (L318, D33, and D39) 3.5. milRNA dynamics and their predicted targets in rye A total of 53 LR-specific sequences were detected. The CP and ICP rust–host interactions were due to different Psr isolates. Additionally, the control rye lines were free of Psr . Thus, milRNA dynamics were examined at 20 and 36 h posttreatment (hpt). There were more differentially expressed milRNAs in the CP interactions than in the ICP interactions. Line D39 had the most differentially expressed milRNAs (20), followed by lines L318 (five) and D33 (three) (Fig. 9 A). For the ICP interactions, there was only one specific sequence in lines L318 and D33. There were many stably expressed milRNAs in all lines and interactions, but there were more in the CP interactions (106, 118, and 133 in D39, L318, and D33, respectively) than in the ICP interactions (77, 58, and 90 in D39, L318, and D33, respectively) (Fig. 9 B). Not all active milRNAs have to be differentially expressed and have important regulatory effects on host gene expression. There were more potential milRNA targets in the CP interactions than in the ICP interactions. The potential targets for all and differentially expressed milRNAs were compared (Fig. 9 C and D). The full list of targets is provided in supplementary Table S2 and S3. 4. Discussion In this study, sRNAs, miRNAs, and milRNAs were detected and analyzed in terms of their expression dynamics in rye– Prs interactions, with a particular focus on their potential targets and regulatory effects on the host transcriptome. There are reports describing the mobilization of sRNAs due to LR infections of wheat [ 21 , 25 , 28 ]. Unfortunately, similar studies have not been conducted for rye, even though it is the closest relative of wheat. 4.1. Comparison of the effectiveness of different algorithms for identifying sRNAs The recently published genomes and transcriptomes of two rye varieties, Chinese Weining and European Lo7 [ 4 , 49 ], significantly accelerated the molecular research on rye. The study conducted by Rabanus-Wallace et al. (2021) explored many aspects of rye transcriptomics, including sRNAs. The authors used their own in-house script to detect sRNAs, which were imprecisely referred to as miRNAs, and identified 13,238 sequences belonging to 90 families. The process underlying miRNA biogenesis is relatively well known and was used to accurately identify miRNAs [ 9 ]. A software package called ShortStack was developed on the basis of a published article [ 40 ]. However, one of the most popular tools for detecting sRNAs is the miRDeep2 software package [ 38 ]. Since its release, it has been used in multiple studies on sRNAs in different organisms. In the current study, the rye sRNAome was analyzed using both ShortStack and miRDeep2, which involve different algorithms, and the resulting sequences were compared with rye transcriptome sequencing data and the sequences deposited in the largest sRNA databases (miRBase and PmiREN). Using this approach, 433 novel sRNA sequences were identified in rye. In the subsequent sRNA transcriptomic analysis, milRNAs synthesized only in response to LR were examined. Because of the lack of an available whole Prs transcriptome, the sRNAome was filtered and screened for milRNAs. This strategy involved dual genome mapping. More specifically, the sequencing reads generated for the LR-infected and mock-inoculated rye leaves were preliminarily mapped to both the Prs and rye genomes. In addition, a Venn diagram was constructed to separate the resulting sRNA sequences. ShortStack was used to identify milRNAs. Using both algorithms to detect sRNAs enabled us to identify more differentially expressed sRNAs than if only one algorithm was used and to analyze their target interactions. Eight miRNAs and 293 sRNAs were detected using ShortStack, whereas 14 sRNAs were detected using miRDeep2. Clearly distinguishing between sRNA and miRNA sequences is conducive to accurately predicting their functions. 4.2. Different interactions between plant gene and s/miRNA transcripts The thorough investigation of the interplay between sRNAs and the targeted mRNAs in plants indicated whole sRNA sequences interact with compatible parts of mRNAs. There are many reports describing the interaction between one sRNA and many targets and vice versa. A single target sequence may have many so-called miRNA-responsive elements (MREs) and interact with many different sRNAs. This phenomenon involving complex multidimensional interactions between different molecules, which is common in diverse biochemical pathways, enhances the regulation of gene expression and the coordination of host responses to pathogen attacks. This interplay among sRNAs, long non-coding RNAs (lncRNAs), and mRNAs results in diverse sRNA–target dynamics (upregulated or downregulated) [ 50 ]. The lncRNAs interact with miRNAs either as target mimics that form competing endogenous RNAs (ceRNAs) or as target lncRNAs that are spliced by miRNAs. Additionally, miRNAs can bind to transcribed pseudogenes and lncRNAs through MREs, which compete with these miRNAs for mRNA-binding sites. These RNAs function as molecular sponges or decoys and suppress the targeting of mRNAs by miRNAs, which explains why they are referred to as ceRNAs. To be able to serve as a ceRNA, the lncRNA must contain an MRE that is partially complementary to the miRNA sequence [ 51 ]. The effect of the interplay between miRNAs and lncRNAs has been investigated in many host–pathogen interactions, including the interaction between wheat and the pathogens responsible for rusts, including LR [ 52 ], yellow rust [ 53 ], and stripe rust [ 54 ]. In the present study, the following distinct interactions between the sRNAs and their targets were detected in the same comparison (i.e., same time-point): (i) sRNAs and their targets were upregulated (i.e., +/+); (ii) sRNAs and their targets were downregulated (i.e., −/−); (iii) sRNAs and their targets had the opposite changes (i.e., +/− or −/+). In all models, many of the targets were for genes with confirmed roles in the immune response to rust fungi, including genes encoding CYP450 [ 34 ], beta-glucanase [ 55 , 56 ], glycosyltransferase [ 57 ], acid invertase [ 58 ], and kaurene synthase [ 59 ] in the +/+ model; chlorophyll a / b -binding protein [ 60 ], beta-glucosidase [ 61 ], fatty acid hydroxylase [ 62 ], and SAUR auxin-responsive protein [ 63 ] in the −/− model; and glutamate receptor in oat [ 64 ] and ammonium transporter [ 65 ] in the +/− model. No immune response-specific genes were identified in the −/+ model. However, the biological significance of these different types of interactions is unclear and may reflect the considerable complexity of the systems regulating various types of non-coding RNA molecules [ 66 ]. The interactions between differentially expressed sRNAs and their targets described in the previous paragraph were for the same time-point, but their interactions can be more dynamic. For example, high or low sRNA levels at one time-point can influence the expression of their targets at a later time-point. Thus, the sRNAs and their targets were examined at different time-points. The downregulated sRNAs at 20 hpt affected the upregulated transcripts of important biotic stress-responsive genes at 36 hpt. Earlier research indicated some of these genes encode receptor-like kinases, proteins involved in detecting pathogens, and ABC transporters belonging to a large family with known roles in the resistance to multiple fungal pathogens in cereals [ 67 ]. In the reverse configuration, upregulated sRNAs at 20 hpt affected downregulated targets at 36 hpt where there were fewer target transcripts, one of which encodes a longifolia protein that regulates xyloglucan endotransglycosylases/hydrolases (XTHs). The XTHs and xyloglucan endotransglycosylases (XETs) are enzymes that repair hemicellulose. Decreases in the expression of the corresponding genes lead to decreases in XET activity and xyloglucan endotransglucosylation, ultimately altering the xyloglucan structure [ 68 ]. These changes might promote fruit softening and wall disassembly, facilitating fungal colonizations and infections [ 69 ]. Another enzyme encoded by a transcript in the reverse configuration group was identified as a strictosidine synthase family member that regulates the secondary metabolism associated with plant responses to pathogens. 4.3. Identification of s/miRNAs in CP and ICP interactions A new era in plant–pathogen interaction-based research was initiated after Flor determined the flax response to a flax rust ( Melampsora lini ) infection and described his gene-for-gene theory in his seminal paper [ 70 ]. The gene-for-gene theory explains the genetic basis of the CP and ICP interactions between plants and pathogens. The theory states that a plant is resistant to a pathogen if it has a resistance ( R ) gene that matches an avirulence ( Avr ) gene from the pathogen. For wheat, which is closely related to rye, more than 200 R genes conferring resistance to rusts (including LR) have been identified (Catalogue of Gene Symbols for Wheat; https://wheat.pw.usda.gov/GG3/wgc ). The expression of many of these genes may be regulated by sRNAs. The changes induced by two Prs isolates associated with CP and ICP interactions were compared. There were more differentially expressed sRNAs and miRNAs in the CP interactions (225 and seven, respectively) than in the ICP interactions (seven and one, respectively), suggesting the response to Prs is more complex during CP interactions than during ICP interactions. The greater number of regulatory sRNAs in CP interactions than in ICP interactions may indicate an increase in the number of targeted transcripts and a more dynamic and complex regulation of the host response during CP interactions. These findings are inconsistent with the results of an earlier study on the potato response to two Phytophthora infestans isolates, in which there were more differentially expressed transcripts in the ICP interaction than in the CP interaction. The reason for this discrepancy might be related to the differences in the selected time-points between the earlier study [ 71 ] and the current study. 4.4. Host genotype-specific targets The three rye lines included in the present study (L318, D33, and D39) differ regarding their response to LR under field and phytotron conditions [ 33 , 34 ]. Lines L318 and D39 are respectively the most susceptible and resistant to LR. In addition, 73, 107, and 55 sRNAs targeting 182, 544, and 116 potential mRNAs were detected in lines L318, D33, and D39, respectively. All but one of the sRNAs were specific to a particular rye line. Interestingly, the most susceptible line (L318) had the most upregulated and downregulated sRNA sequences (62 and 12, respectively). In contrast, in the most resistant line (D39), there were 32 and 23 upregulated and downregulated sRNAs, respectively (i.e., relatively even number of altered sRNAs). This may be related to the number of targets detected in L318 and D39 (i.e. 182 and 116, respectively). Compared with the resistant line, the susceptible line may require the activation of more genes following an LR infection. 4.5. IsomiRs: the influence of small sequence differences on target selection The identified isomiRs in rye infected with a fungal pathogen included 20 and 21 nt sequences with 14 nt in common. The isomiRs that differed by only one nucleotide were observed to target different mRNAs in the host. The longer sequence (UGUGUUCUCAGGUCGCCCCCG) targeted the transcripts of 11 endoglucanase genes and two superoxide dismutase genes, which are well known for their roles in plant defenses. Endoglucanases are required for the formation of cellulose microfibrils, but recent research indicated fungi can hijack a ubiquitous plant apoplastic endoglucanase, leading to the release of a reactive oxygen species-scavenging β-glucan decasaccharide that subverts immune responses [ 72 ]. The shorter sequence (UCAGGUCGCCCCCGCUGGAG) targeted the transcripts of 16 transcription factor genes (e.g., MADS box gene). Some of the proteins encoded by the targets contribute to plant stress responses. SPIRAL1 plays a vital role in stress-induced microtubule disassembly, which is required for cell wall rearrangements and anisotropic cell growth. The AP2-like ethylene-responsive transcription factor is a member of a protein group that functions at a hub in hormone and stress signaling pathways [ 73 ]. Chloroplastic chlorophyll a / b -binding proteins decrease oxidative stress, while CsAtPR5 belongs to a pathogen-responsive protein family. 4.6. Role of cross-kingdom RNA interference in CP and ICP interactions Cross-kingdom RNA interference (ck-RNAi) is an sRNA-mediated gene silencing mechanism affecting hosts and pathogens [ 74 ]. Fungi and oomycetes lack typical miRNAs, but similar sRNA sequences were identified in the fungus N. crassa and designated as milRNAs. This new class of small non-coding RNAs was subsequently detected in diverse fungi and oomycetes, including plant pathogenic species. In fact, the first example of a non-protein effector influencing the response of tomato to gray mold caused by B. cinerea was a fungal milRNA that silenced the transcripts of host defense response-related genes [ 17 ]. Since then, similar examples of pathogen factors silencing host genes have been reported for various agriculturally important crops, including wheat, barley, rice, and tomato [ 20 , 75 , 76 ]. The host targets encoded several proteins involved in responses to pathogen attacks, including pathogenesis-related proteins with NB-ARC domains. The transcripts targeted by milRNAs encoded lipid metabolism-related enzymes crucial for plant responses to biotic stress. For example, fatty acyl reductase catalyzes the NADPH-dependent reduction of fatty acyl-CoA or acyl-ACP substrates to primary fatty alcohols, which are intermediate metabolites or metabolic end products that contribute to the formation of plant extracellular protective lipid barriers (e.g., cuticular wax, sporopollenin, suberin, and taproot wax) [ 77 ]. Notably, one of the targets was a transcript encoding patatin. Both patatin and patatin-like proteins (PLPs) are involved in plant responses to pathogens, especially fungi. A recent study showed that in Nicotiana attenuata , NaPLP expression increases substantially following an infection by Alternaria alternata [ 78 ]. Another targeted transcript, which encodes a methyl esterase, was downregulated by the upregulated milRNAs. This finding is in accordance with the results of an earlier study in which methyl esterase 1 (PvMES1) promoted the salicylic acid pathway and enhanced the resistance of common bean to Fusarium wilt [ 79 ]. The basic helix-loop-helix (bHLH) DNA-binding proteins are involved in the biosynthesis of diterpenoid phytoalexins, which might be induced in blast-infected rice[ 80 ]. Interestingly, milRNA can reportedly target transcripts encoding histone 3, which is post-transcriptionally modified in response to different biotic stresses [ 81 ]. 4.7. Utility of sRNAs for protecting crops and promoting sustainable agriculture The global application of pesticides on crops reached 2.66 tons in 2020. Their harmful effects on the environment are well known [ 82 ]. The research community has proposed many environmentally friendly alternatives to pesticides, one of which involves host-induced gene silencing (HIGS). In this technique, a plant is transformed with a target gene (partial or complete sequence) originating from a pathogen. The expression of the transgene triggers RNA interference, thereby effectively silencing the expression of an essential pathogen gene. Recent research confirmed that HIGS is effective for controlling hemibiotrophic or necrotrophic fungal pathogens (e.g., Verticillium dahliae and B. cinerea ) as well as the obligate biotroph P. triticina and Fusarium species. A simpler technique that does not require the production of transgenic plants is spray-induced gene silencing (SIGS), which involves spraying host plants with the RNA of pathogen genes. After the SIGS technique was first developed in 2016, it was used to protect certain crops from pathogens [ 83 ], including canola infected with B. cinerea and Sclerotinia sclerotiorum , barley infected with Fusarium species, and wheat infected with Rhizoctonia cerealis . The RNAs used to silence genes were sequences targeting transcripts related to thioredoxin reductase and ergosterol biosynthesis. Interestingly, the earlier study also provided evidence that SIGS may have the same effects as a semi-systemic fungicide. When plants were sprayed with CYP3-dsRNAs, they were protected from the disease even in parts that were not sprayed (e.g., distal leaf parts). Moreover, CYP3-dsRNAs were detected in the sprayed and unsprayed leaf parts. The subsequent analysis of dsRNA localization using green fluorescent dye ATTO 488-labeled dsRNAs indicated that the dsRNAs at the leaf surface were taken up by plant cells and then moved to different parts of the plant, including the stomata, xylem cells, companion cells, phloem parenchyma cells, mesophyll cells, and trichomes [ 84 ]. Accordingly, these dsRNAs are eco-friendly fungicides that are safe for the environment and humans. There has been research conducted to address some of the issues that may need to be addressed, including the stability and efficient delivery of sRNAs to pathogens. To increase RNA stability, researchers docked an antiviral dsRNA to double hydroxide clay nanosheets, which resulted in enhanced protection against phytopathogenic viruses [ 85 ]. In addition, the use of artificial vesicles or liposomes to protect RNAs may improve SIGS techniques for protecting plants and developing antifungal compounds, especially for fungi that can efficiently take up extracellular vesicles (EVs) [ 84 ]. In conclusion, we detected rye and Prs sRNAs that target vital transcripts for host–pathogen interactions, including transcripts encoding glucanases, proteins with NBS-LRR domains, peroxidases, and different transcription factors. Furthermore, newly detected Prs milRNAs targeting host resistance-related transcripts (e.g., those encoding NBS-LRR proteins, peroxidases, ABC transporters, and pathogenesis-related proteins) may be silenced or eliminated using Short Tandem Target Mimic (STTMs) as part of a SIGS-based strategy to prevent rust infections. This approach may protect rye against fungal infections (at least LR) and is fully compliant with the current EU regulations that favor this type of agro-technical solution over the genetic modification of crop plants. Declarations CRediT authorship contribution statement Marek Daniel Koter: Data analysis, Writing. Mateusz Matuszkiewicz : Data analysis. Monika Rakoczy-Trojanowska : Conceptualization, Writing - review & editing, Funding acquisition. Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Acknowledgments This work was financially supported by the Polish National Science Centre (grant number 2018/31/B/NZ9/00439). We thank Edanz (www.edanz.com/ac) for editing a draft of this manuscript. References K. Peksa, B. Bankina, Characterization of Puccinia recondita, the causal agent of brown rust: A review, Research for Rural Development 2 (2019) 70–76. O.V. Solodukhina, V.D. Kobylyanskij, Problems of winter rye breeding for resistance to leaf and stem rusts, Plant Breed Seed Sci. 48(2/2) (2003) 87–96. S.R. Roux, B. Hackauf, B. Ruge-Wehling, A. Linz, P. Wehling, Exploitation and comprehensive characterization of leaf rust resistance in rye, Vortr. Pflanzenzüchtung 71 (2007) 144–150. M.T. Rabanus-Wallace, B. Hackauf, M. Mascher, T. Lux, T. Wicker, H. Gundlach, M. Baez, A. Houben, K.F.X. Mayer, L. Guo, J. Poland, C.J. Pozniak, S. Walkowiak, J. Melonek, C.R. Praz, M. Schreiber, H. Budak, M. Heuberger, B. Steuernagel, B. Wulff, A. Börner, B. Byrns, J. Čížková, D.B. Fowler, A. Fritz, A. Himmelbach, G. Kaithakottil, J. Keilwagen, B. Keller, D. Konkin, J. Larsen, Q. Li, B. Myśków, S. Padmarasu, N. Rawat, U. Sesiz, S. Biyiklioglu-Kaya, A. Sharpe, H. Šimková, I. Small, D. Swarbreck, H. Toegelová, N. Tsvetkova, A.V. Voylokov, J. Vrána, E. Bauer, H. Bolibok-Bragoszewska, J. Doležel, A. Hall, J. Jia, V. Korzun, A. Laroche, X.F. Ma, F. Ordon, H. Özkan, M. Rakoczy-Trojanowska, U. Scholz, A.H. Schulman, D. Siekmann, S. Stojałowski, V.K. Tiwari, M. Spannagl, N. Stein, Chromosome-scale genome assembly provides insights into rye biology, evolution and agronomic potential, Nat Genet 53(4) (2021) 564–573. N.M. Vendelbo, K. Mahmood, P. Sarup, M.S. Hovmøller, A.F. Justesen, P.S. Kristensen, J. Orabi, A. Jahoor, Discovery of a Novel Leaf Rust (Puccinia recondita) Resistance Gene in Rye (Secale cereale L.) Using Association Genomics, Cells 11(1) (2021) 64. N.M. Vendelbo, K. Mahmood, B. Steuernagel, B.B.H. Wulff, P. Sarup, M.S. Hovmøller, A.F. Justesen, P.S. Kristensen, J. Orabi, A. Jahoor, Discovery of Resistance Genes in Rye by Targeted Long-Read Sequencing and Association Genetics, Cells 11(8) (2022). T. Krępski, M. Olechowski, I. Samborska-Skutnik, M. Święcicka, A. Grądzielewska, M. Rakoczy-Trojanowska, Identification and characteristics of wheat Lr orthologs in three rye inbred lines, PLoS One 18(7) (2023) e0288520. P. Guleria, M. Mahajan, J. Bhardwaj, S.K. Yadav, Plant Small RNAs: Biogenesis, Mode of Action and Their Roles in Abiotic Stresses, Genomics, Proteomics & Bioinformatics 9(6) (2011) 183–199. M.J. Axtell, B.C. Meyers, Revisiting Criteria for Plant MicroRNA Annotation in the Era of Big Data, Plant Cell 30(2) (2018) 272–284. H. Budak, M. Kantar, R. Bulut, B.A. Akpinar, Stress responsive miRNAs and isomiRs in cereals, Plant Sci 235 (2015) 1–13. E.M. Fard, S. Moradi, N.N. Salekdeh, B. Bakhshi, M.R. Ghaffari, M. Zeinalabedini, G.H. Salekdeh, Plant isomiRs: origins, biogenesis, and biological functions, Genomics 112(5) (2020) 3382–3395. C.T. Neilsen, G.J. Goodall, C.P. Bracken, IsomiRs–the overlooked repertoire in the dynamic microRNAome, Trends Genet 28(11) (2012) 544–9. C. Lelandais-Brière, L. Naya, E. Sallet, F. Calenge, F. Frugier, C. Hartmann, J. Gouzy, M. Crespi, Genome-wide Medicago truncatula small RNA analysis revealed novel microRNAs and isoforms differentially regulated in roots and nodules, Plant Cell 21(9) (2009) 2780–96. M.D. Koter, M. Święcicka, M. Matuszkiewicz, A. Pacak, N. Derebecka, M. Filipecki, The miRNAome dynamics during developmental and metabolic reprogramming of tomato root infected with potato cyst nematode, Plant Sci 268 (2018) 18–29. M. Święcicka, W. Skowron, P. Cieszyński, J. Dąbrowska-Bronk, M. Matuszkiewicz, M. Filipecki, M.D. Koter, The suppression of tomato defence response genes upon potato cyst nematode infection indicates a key regulatory role of miRNAs, Plant Physiol Biochem 113 (2017) 51–55. N. Romano, G. Macino, Quelling: transient inactivation of gene expression in Neurospora crassa by transformation with homologous sequences, Mol Microbiol 6(22) (1992) 3343–53. A. Weiberg, M. Wang, M. Bellinger, H. Jin, Small RNAs: a new paradigm in plant-microbe interactions, Annu Rev Phytopathol 52 (2014) 495–516. B. Wang, Y. Sun, N. Song, M. Zhao, R. Liu, H. Feng, X. Wang, Z. Kang, Puccinia striiformis f. sp. tritici microRNA-like RNA 1 (Pst-milR1), an important pathogenicity factor of Pst, impairs wheat resistance to Pst by suppressing the wheat pathogenesis-related 2 gene, New Phytologist 215(1) (2017) 338–350. B. Wang, Y. Sun, N. Song, M. Zhao, R. Liu, H. Feng, X. Wang, Z. Kang, Puccinia striiformis f. sp. tritici microRNA-like RNA 1 (Pst-milR1), an important pathogenicity factor of Pst, impairs wheat resistance to Pst by suppressing the wheat pathogenesis-related 2 gene, New Phytol 215(1) (2017) 338–350. M. Xu, G. Li, Y. Guo, Y. Gao, L. Zhu, Z. Liu, R. Tian, C. Gao, P. Han, N. Wang, F. Guo, J. Bao, C. Jia, H. Feng, L. Huang, A fungal microRNA-like RNA subverts host immunity and facilitates pathogen infection by silencing two host receptor‐like kinase genes, New Phytologist 233(6) (2022) 2503–2519. D. Kumar, D. Singh, P. Kanodia, K.V. Prabhu, M. Kumar, K. Mukhopadhyay, Discovery of Novel Leaf Rust Responsive microRNAs in Wheat and Prediction of Their Target Genes, J Nucleic Acids 2014 (2014) 570176. N.A. Mueth, S.R. Ramachandran, S.H. Hulbert, Small RNAs from the wheat stripe rust fungus ( Puccinia striiformis f.sp. tritici ), BMC Genomics 16(1) (2015) 718. R.S. Poethig, Small RNAs and developmental timing in plants, Curr Opin Genet Dev 19(4) (2009) 374–8. O.P. Gupta, V. Permar, V. Koundal, U.D. Singh, S. Praveen, MicroRNA regulated defense responses in Triticum aestivum L. during Puccinia graminis f.sp. tritici infection, Mol Biol Rep 39(2) (2012) 817–24. S. Dutta, M. Kumar, K. Mukhopadhyay, Identification and Validation of Leaf Rust Responsive Wheat isomiRs and their Target Genes in both Wheat and Puccinia triticina, (2019). S. Dutta, D. Kumar, S. Jha, K.V. Prabhu, M. Kumar, K. Mukhopadhyay, Identification and molecular characterization of a trans-acting small interfering RNA producing locus regulating leaf rust responsive gene expression in wheat ( Triticum aestivum L.), Planta 246(5) (2017) 939–957. Ö. Bilir, D. Göl, Y. Hong, J.M. McDowell, M. Tör, Small RNA-based plant protection against diseases, Front Plant Sci 13 (2022) 951097. M.M. Nair, S.H.K. Kumar, S. Jyothsna, K.T. Sundaram, C. Manjunatha, M. Sivasamy, M. Alagu, Stem and leaf rust-induced miRNAome in bread wheat near-isogenic lines and their comparative analysis, Appl Microbiol Biotechnol 106(24) (2022) 8211–8232. P. Gruner, A. Witzke, K. Flath, J. Eifler, B. Schmiedchen, M. Schmidt, A. Gordillo, D. Siekmann, F.J. Fromme, S. Koch, H.P. Piepho, T. Miedaner, Studying Stem Rust and Leaf Rust Resistances of Self-Fertile Rye Breeding Populations, Int J Mol Sci 23(22) (2022). K. Kumar, I. Jan, G. Saripalli, P.K. Sharma, R.R. Mir, H.S. Balyan, P.K. Gupta, An Update on Resistance Genes and Their Use in the Development of Leaf Rust Resistant Cultivars in Wheat, Front Genet 13 (2022) 816057. D. Kumar, S. Dutta, D. Singh, K.V. Prabhu, M. Kumar, K. Mukhopadhyay, Uncovering leaf rust responsive miRNAs in wheat ( Triticum aestivum L.) using high-throughput sequencing and prediction of their targets through degradome analysis, Planta 245(1) (2017) 161–182. H. Feng, T. Wang, C. Feng, Q. Zhang, X. Zhang, L. Huang, X. Wang, Z. Kang, Identification of microRNAs and their corresponding targets involved in the susceptibility interaction of wheat response to Puccinia striiformis f. sp. tritici , Physiol Plant 157(1) (2016) 95–107. M. Rakoczy-Trojanowska, P. Krajewski, J. Bocianowski, M. Schollenberger, W. Wakuliński, P. Milczarski, P. Masojć, M. Targońska-Karasek, Z. Banaszak, K. Banaszak, W. Brukwiński, W. Orczyk, A. Kilian, Identification of Single Nucleotide Polymorphisms Associated with Brown Rust Resistance, α-Amylase Activity and Pre-harvest Sprouting in Rye (Plant Mol Biol Report 35(3) (2017) 366–378. M. Święcicka, M. Dmochowska-Boguta, W. Orczyk, A. Grądzielewska, A. Stochmal, M. Kowalczyk, L. Bolibok, M. Rakoczy-Trojanowska, Changes in benzoxazinoid contents and the expression of the associated genes in rye ( Secale cereale L.) due to brown rust and the inoculation procedure, PLoS One 15(5) (2020) e0233807. M. Martin, Cutadapt removes adapter sequences from high-throughput sequencing reads, EMBnet.journal 17(1) (2011) 10. S. Andrews, FASTQC. A quality control tool for high throughput sequence data, 2010. B. Langmead, C. Trapnell, M. Pop, S.L. Salzberg, Ultrafast and memory-efficient alignment of short DNA sequences to the human genome, Genome Biol 10(3) (2009) R25. S.D. Mackowiak, Identification of novel and known miRNAs in deep-sequencing data with miRDeep2, Curr Protoc Bioinformatics Chap. 12 (2011) Unit 12.10. M.I. Love, W. Huber, S. Anders, Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2, Genome Biol 15(12) (2014) 550. M.J. Axtell, ShortStack: comprehensive annotation and quantification of small RNA genes, RNA 19(6) (2013) 740–51. M.D. Robinson, D.J. McCarthy, G.K. Smyth, edgeR: a Bioconductor package for differential expression analysis of digital gene expression data, Bioinformatics 26(1) (2010) 139–40. A. Kozomara, M. Birgaoanu, S. Griffiths-Jones, miRBase: from microRNA sequences to function, Nucleic Acids Res 47(D1) (2019) D155-D162. Z. Guo, Z. Kuang, Y. Wang, Y. Zhao, Y. Tao, C. Cheng, J. Yang, X. Lu, C. Hao, T. Wang, X. Cao, J. Wei, L. Li, X. Yang, PmiREN: a comprehensive encyclopedia of plant miRNAs, Nucleic Acids Res 48(D1) (2020) D1114-D1121. X. Dai, Z. Zhuang, P.X. Zhao, psRNATarget: a plant small RNA target analysis server (2017 release), Nucleic Acids Research 46(W1) (2018) W49-W54. L. Moyo, S.V. Ramesh, M. Kappagantu, N. Mitter, V. Sathuvalli, H.R. Pappu, The effects of potato virus Y-derived virus small interfering RNAs of three biologically distinct strains on potato ( Solanum tuberosum ) transcriptome, Virology Journal 14(1) (2017). U. Raudvere, L. Kolberg, I. Kuzmin, T. Arak, P. Adler, H. Peterson, J. Vilo, g:Profiler: a web server for functional enrichment analysis and conversions of gene lists (2019 update), Nucleic Acids Res 47(W1) (2019) W191-W198. L. Kolberg, U. Raudvere, I. Kuzmin, J. Vilo, H. Peterson, gprofiler2 -- an R package for gene list functional enrichment analysis and namespace conversion toolset g:Profiler, F1000Res 9 (2020). S. Feng, C. Pan, S. Ding, Q. Ma, C. Hu, P. Wang, K. Shi, The Glutamate Receptor Plays a Role in Defense against Botrytis cinerea through Electrical Signaling in Tomato, Applied Sciences 11(23) (2021) 11217. G. Li, L. Wang, J. Yang, H. He, H. Jin, X. Li, T. Ren, Z. Ren, F. Li, X. Han, X. Zhao, L. Dong, Y. Li, Z. Song, Z. Yan, N. Zheng, C. Shi, Z. Wang, S. Yang, Z. Xiong, M. Zhang, G. Sun, X. Zheng, M. Gou, C. Ji, J. Du, H. Zheng, J. Doležel, X.W. Deng, N. Stein, Q. Yang, K. Zhang, D. Wang, A high-quality genome assembly highlights rye genomic characteristics and agronomically important genes, Nat Genet 53(4) (2021) 574–584. X. Yang, L. Zhang, Y. Yang, M. Schmid, Y. Wang, miRNA Mediated Regulation and Interaction between Plants and Pathogens, Int J Mol Sci 22(6) (2021). A. Biswas, B. Sen, S. Bandyopadhyay, C. Mal, Co-regulatory functions of miRNA and lncRNA in adapting biotic and abiotic stress in economically important dicot plants, Plant Gene 26 (2021). N. Jain, S. Rani, C. Sharma, N. Sinha, A. Singh, J.B. Sharma, P. Prasad, G. Saripalli, P.K. Sharma, H.S. Balyan, P.K. Gupta, K.V. Prabhu, Large-scale stage-specific regulation of gene expression during host-pathogen interactions in CSP44 bread wheat carrying APR gene Lr48, Funct Plant Biol 47(3) (2020) 203–225. S. Das, D. Singh, H.S. Meena, S.K. Jha, J. Kumari, V. Chinnusamy, L. Sathee, Long term nitrogen deficiency alters expression of miRNAs and alters nitrogen metabolism and root architecture in Indian dwarf wheat ( Triticum sphaerococcum Perc.) genotypes, Sci Rep 13(1) (2023) 5002. H. Zhang, X. Chen, C. Wang, Z. Xu, Y. Wang, X. Liu, Z. Kang, W. Ji, Long non-coding genes implicated in response to stripe rust pathogen stress in wheat ( Triticum aestivum L.), Mol Biol Rep 40(11) (2013) 6245–53. B. Liu, X. Xue, S. Cui, X. Zhang, Q. Han, L. Zhu, X. Liang, X. Wang, L. Huang, X. Chen, Z. Kang, Cloning and characterization of a wheat β-1,3-glucanase gene induced by the stripe rust pathogen Puccinia striiformis f. sp. tritici , Molecular Biology Reports 37(2) (2010) 1045–1052. L. Gao, S. Wang, Y. Zhang, X. Li, H. Wang, D. Liu, Identification and characterization of a β-1, 3-glucanase gene, TcLr19Glu, involved in wheat resistance against Puccinia triticina , J. Plant Biochem. Biotechnol. 25 (2016) 319–326. A. Amo, J.M. Soriano, Unravelling consensus genomic regions conferring leaf rust resistance in wheat via meta-QTL analysis, Plant Genome 15(1) (2022) e20185. S. Liu, J. Lan, B. Zhou, Y. Qin, Y. Zhou, X. Xiao, J. Yang, J. Gou, J. Qi, Y. Huang, C. Tang, HbNIN2, a cytosolic alkaline/neutral-invertase, is responsible for sucrose catabolism in rubber-producing laticifers of Hevea brasiliensis (para rubber tree), New Phytol 206(2) (2015) 709–25. T.E. Coram, X. Huang, G. Zhan, M.L. Settles, X. Chen, Meta-analysis of transcripts associated with race-specific resistance to stripe rust in wheat demonstrates common induction of blue copper-binding protein, heat-stress transcription factor, pathogen-induced WIR1A protein, and ent-kaurene synthase transcripts, Funct Integr Genomics 10(3) (2010) 383–92. P. Prasad, S. Savadi, S.C. Bhardwaj, P.L. Kashyap, O.P. Gangwar, H. Khan, S. Kumar, R. Kumar, V. Patil, Stage-specific reprogramming of defense responsive genes during Lr24-mediated leaf rust resistance in wheat, Journal of Plant Pathology 111(2) (2019) 283–293. M.S. Mafa, N. Lebusa, T.F. Gumani, G. Kemp, B. Visser, W.H.P. Boshoff, H.D. Castelyn, Accumulation of complex oligosaccharides and CAZymes activity under acid conditions constitute the Thatcher + Lr9 defence responses to Puccinia triticina , Biologia 78 (2023). E.C. Henningsen, V. Omidvar, R. Della Coletta, J.M. Michno, E. Gilbert, F. Li, M.E. Miller, C.L. Myers, S.P. Gordon, J.P. Vogel, B.J. Steffenson, S.F. Kianian, C.D. Hirsch, M. Figueroa, Identification of Candidate Susceptibility Genes to, Front Plant Sci 12 (2021) 657796. Z. Zhao, T. Chen, J. Yue, N. Pu, J. Liu, L. Luo, M. Huang, T. Guo, W. Xiao, regulates heading date in rice, Mol Breed 43(8) (2023) 62. Y. Loarce, E. Navas, C. Paniagua, A. Fominaya, J.L. Manjón, E. Ferrer, Identification of Genes in a Partially Resistant Genotype of Avena sativa Expressed in Response to Puccinia coronata Infection, Front Plant Sci 7 (2016) 731. S. Li, C. Castillo-González, B. Yu, X. Zhang, The functions of plant small RNAs in development and in stress responses, Plant J 90(4) (2017) 654–670. S. Bhogireddy, S.K. Mangrauthia, R. Kumar, A.K. Pandey, S. Singh, A. Jain, H. Budak, R.K. Varshney, H. Kudapa, Regulatory non-coding RNAs: a new frontier in regulation of plant biology, Funct Integr Genomics 21(3–4) (2021) 313–330. S.G. Krattinger, E.S. Lagudah, W. Spielmeyer, R.P. Singh, J. Huerta-Espino, H. McFadden, E. Bossolini, L.L. Selter, B. Keller, A putative ABC transporter confers durable resistance to multiple fungal pathogens in wheat, Science 323(5919) (2009) 1360–3. Y. Han, Q. Zhu, Z. Zhang, K. Meng, Y. Hou, Q. Ban, J. Suo, J. Rao, Analysis of xyloglucan endotransglycosylase/hydrolase (XTH) genes and diverse roles of isoenzymes during persimmon fruit development and postharvest softening, PLoS One 10(4) (2015) e0123668. E. Miedes, E.P. Lorences, The implication of xyloglucan endotransglucosylase/hydrolase (XTHs) in tomato fruit infection by Penicillium expansum Link. A, J Agric Food Chem 55(22) (2007) 9021–6. H.H. Flor, Current Status of the Gene-For-Gene Concept, Annual Review of Phytopathology 9(1) (1971) 275–296. Y. Duan, S. Duan, M.R. Armstrong, J. Xu, J. Zheng, J. Hu, X. Chen, I. Hein, G. Li, L. Jin, Comparative Transcriptome Profiling Reveals Compatible and Incompatible Patterns of Potato Toward, G3 (Bethesda) 10(2) (2020) 623–634. B. Chandrasekar, A. Wanke, S. Wawra, P. Saake, L. Mahdi, N. Charura, M. Neidert, G. Poschmann, M. Malisic, M. Thiele, K. Stühler, M. Dama, M. Pauly, A. Zuccaro, Fungi hijack a ubiquitous plant apoplastic endoglucanase to release a ROS scavenging β-glucan decasaccharide to subvert immune responses, Plant Cell 34(7) (2022) 2765–2784. M. Müller, S. Munné-Bosch, Ethylene Response Factors: A Key Regulatory Hub in Hormone and Stress Signaling, Plant Physiol 169(1) (2015) 32–41. F. Dunker, A. Trutzenberg, J.S. Rothenpieler, S. Kuhn, R. Pröls, T. Schreiber, A. Tissier, A. Kemen, E. Kemen, R. Hückelhoven, A. Weiberg, Oomycete small RNAs bind to the plant RNA-induced silencing complex for virulence, Elife 9 (2020). H. Dubey, K. Kiran, R. Jaswal, P. Jain, A.M. Kayastha, S.C. Bhardwaj, T.K. Mondal, T.R. Sharma, Discovery and profiling of small RNAs from Puccinia triticina by deep sequencing and identification of their potential targets in wheat. J. Mapuranga, J. Chang, L. Zhang, N. Zhang, W. Yang, Fungal Secondary Metabolites and Small RNAs Enhance Pathogenicity during Plant-Fungal Pathogen Interactions, Journal of Fungi 9(1) (2023) 4. X. Zhang, Y. Liu, A. Ayaz, H. Zhao, S. Lü, The Plant Fatty Acyl Reductases, Int J Mol Sci 23(24) (2022). J. Cheng, N. Song, J. Wu, A patatin-like protein synergistically regulated by jasmonate and ethylene signaling pathways plays a negative role in, Plant Divers 41(1) (2019) 7–12. R. Xue, M. Feng, J. Chen, W. Ge, M.W. Blair, A methyl esterase 1 (PvMES1) promotes the salicylic acid pathway and enhances Fusarium wilt resistance in common beans, Theor Appl Genet 134(8) (2021) 2379–2398. M. Hasegawa, I. Mitsuhara, S. Seo, T. Imai, J. Koga, K. Okada, H. Yamane, Y. Ohashi, Phytoalexin Accumulation in the Interaction Between Rice and the Blast Fungus, Molecular Plant-Microbe Interactions 23(8) (2010) 1000–1011. J.S. Ramirez-Prado, S.J.M. Piquerez, A. Bendahmane, H. Hirt, C. Raynaud, M. Benhamed, Modify the Histone to Win the Battle: Chromatin Dynamics in Plant-Pathogen Interactions, Front Plant Sci 9 (2018) 355. A. Sharma, V. Kumar, B. Shahzad, M. Tanveer, G.P.S. Sidhu, N. Handa, S.K. Kohli, P. Yadav, A.S. Bali, R.D. Parihar, O.I. Dar, K. Singh, S. Jasrotia, P. Bakshi, M. Ramakrishnan, S. Kumar, R. Bhardwaj, A.K. Thukral, Worldwide pesticide usage and its impacts on ecosystem, SN Appl. Sci. 1 (2019) 1446. H. Sang, J.-I. Kim, Advanced strategies to control plant pathogenic fungi by host-induced gene silencing (HIGS) and spray-induced gene silencing (SIGS), Plant Biotechnology Reports 14 (2020) 1–8. B.T.L. Hoang, S.J. Fletcher, C.A. Brosnan, A.B. Ghodke, N. Manzie, N. Mitter, RNAi as a Foliar Spray: Efficiency and Challenges to Field Applications, Int J Mol Sci 23(12) (2022). N. Mitter, E.A. Worrall, K.E. Robinson, P. Li, R.G. Jain, C. Taochy, S.J. Fletcher, B.J. Carroll, G.Q. Lu, Z.P. Xu, Clay nanosheets for topical delivery of RNAi for sustained protection against plant viruses, Nat Plants 3 (2017) 16207. Additional Declarations The authors declare no competing interests. Supplementary Files TableS1.xlsx TableS2.xlsx TableS3.xlsx Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-3875675","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":267827584,"identity":"ce27b15a-1601-4e60-b9ec-f1cd9c2a257f","order_by":0,"name":"Marek Daniel Koter","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA/ElEQVRIiWNgGAWjYJACxgaGBDD9ACpgAMQWhLXwMDAwGyBpkSBKCxtMGX4tuu1nHz6cwZAmZ8/e+6zyx6/axAb25m0SjDtwazE7k25suIEhx5iH57jZbd6+44kNPMfKJBjP4NFyII1N8gFDRWKPRBrbbcaeY7kNEjlmEoxteLScf8b+E6xF/hlb4U+QFvk3BLTcSGNjBDoMaAsbGwPPjxqgLTyEtDxjlgR635jnTBqzNG/Dgfo2nrRii0R8fjmfxvixhyFZjr39GOPHH3/qjPnZD2+88XGHDU4tYMD4D8ZoO8zABmIkNuDXgQT+1EH1Eq9lFIyCUTAKhj8AABqsUW6D1madAAAAAElFTkSuQmCC","orcid":"https://orcid.org/0000-0002-9821-8475","institution":"Warsaw University of Life Sciences","correspondingAuthor":true,"prefix":"","firstName":"Marek","middleName":"Daniel","lastName":"Koter","suffix":""},{"id":267827948,"identity":"3d7b4244-f412-4adc-b10c-de571cf257e4","order_by":1,"name":"Mateusz Matuszkiewicz","email":"","orcid":"https://orcid.org/0000-0002-4320-7103","institution":"Warsaw University of Life Sciences","correspondingAuthor":false,"prefix":"","firstName":"Mateusz","middleName":"","lastName":"Matuszkiewicz","suffix":""},{"id":267828197,"identity":"6a5561c4-3824-421e-809a-f3a6caddee36","order_by":2,"name":"Monika Rakoczy-Trojanowska","email":"","orcid":"https://orcid.org/0000-0001-6891-0471","institution":"Warsaw University of Life Sciences","correspondingAuthor":false,"prefix":"","firstName":"Monika","middleName":"","lastName":"Rakoczy-Trojanowska","suffix":""}],"badges":[],"createdAt":"2024-01-18 11:52:30","currentVersionCode":1,"declarations":{"humanSubjects":false,"vertebrateSubjects":false,"conflictsOfInterestStatement":false,"humanSubjectEthicalGuidelines":false,"humanSubjectConsent":false,"humanSubjectClinicalTrial":false,"humanSubjectCaseReport":false,"vertebrateSubjectEthicalGuidelines":false},"doi":"10.21203/rs.3.rs-3875675/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-3875675/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":49855374,"identity":"d2368540-5c0f-4dfe-89d3-726ec9f74554","added_by":"auto","created_at":"2024-01-19 07:14:00","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":70799,"visible":true,"origin":"","legend":"\u003cp\u003eBioinformatics pipeline for the detection of different sRNA classes. Sequence reads were trimmed using Cutadapt and mapped to the Lo7 genome using Bowtie. The edgeR package was used to detect differentially expressed sRNAs, whereas psRNATarget was used to identify their mRNA targets in the Lo7 transcriptome.\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-3875675/v1/62e6b2e6dad4a2425202b2ee.png"},{"id":49855375,"identity":"278295d9-1221-4e58-8993-c7f212f809d3","added_by":"auto","created_at":"2024-01-19 07:14:00","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":67035,"visible":true,"origin":"","legend":"\u003cp\u003eAverage number of all reads and unique reads (18–35 nucleotides long) in sequence libraries. Among all reads, the most common reads were sRNAs consisting of 21, 22, or 24 nt and tRFs consisting of 33 nt.\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-3875675/v1/24cbe91ef1dca38edd3fe1df.png"},{"id":49855376,"identity":"10154afb-8666-45f9-82d6-e5bc7d8ced7b","added_by":"auto","created_at":"2024-01-19 07:14:00","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":188380,"visible":true,"origin":"","legend":"\u003cp\u003eVenn diagram of all sRNA sequences from the PmiREN and miRbase databases and the sRNAs identified using miRDeep2 and ShortStack and from a published report [4].\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-3875675/v1/8a3b0f51781c84f0efc90742.png"},{"id":49855669,"identity":"155aaa0f-ebae-4d12-ac90-79a421009b00","added_by":"auto","created_at":"2024-01-19 07:22:01","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":49991,"visible":true,"origin":"","legend":"\u003cp\u003eNumber of differentially expressed sRNAs per experiment.\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-3875675/v1/91f1ecf5cabe886b43cb3e17.png"},{"id":49855385,"identity":"7d9988af-4eaf-493d-ae3b-f3e801c23fb1","added_by":"auto","created_at":"2024-01-19 07:14:01","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":107261,"visible":true,"origin":"","legend":"\u003cp\u003eExamples of different sRNA classes affected by the Prs infection. The threshold for identifying differentially expressed transcripts was as follows: |log2(fold change)| \u0026gt;1 (fold-change) ≥ −2 or 2 (A) or |log2(fold change)| \u0026gt;1 (B). Both sRNAs and miRNAs were identified by ShortStack.\u003c/p\u003e","description":"","filename":"5.png","url":"https://assets-eu.researchsquare.com/files/rs-3875675/v1/3498ac7e1b9f93cd07f7701a.png"},{"id":49856302,"identity":"e438ddf3-b43b-41aa-8cc6-e20d36535580","added_by":"auto","created_at":"2024-01-19 07:38:01","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":132774,"visible":true,"origin":"","legend":"\u003cp\u003eNumber of differentially expressed host sRNAs (20–24 nt) and miRNAs in the CP and ICP host–pathogen interactions (A). Number of differentially expressed host sRNAs (20–24 nt) and miRNAs at 20 and 36 h post-treatment (hpt) (B).\u003c/p\u003e","description":"","filename":"6.png","url":"https://assets-eu.researchsquare.com/files/rs-3875675/v1/ae473bcb57aa834e9d8d7877.png"},{"id":49855673,"identity":"3629d04d-71bd-4c86-8d36-69d798be3523","added_by":"auto","created_at":"2024-01-19 07:22:01","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":223760,"visible":true,"origin":"","legend":"\u003cp\u003e(A) IsomiRs, which are sRNAs mostly modified at their 3′ or 5′ ends, leading to differences in their expression profiles. (B) Their sequence diversity substantially affects their targets. The Venn diagram presents the differences in the potential isomiR targets. The numbers represent the specific and shared targets of selected isomiRs.\u003c/p\u003e","description":"","filename":"7.png","url":"https://assets-eu.researchsquare.com/files/rs-3875675/v1/8db0bfad2f67ad5d5ec30f42.png"},{"id":49856791,"identity":"e79297fe-e74c-49a6-8540-5d89afa4dbdd","added_by":"auto","created_at":"2024-01-19 07:46:01","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":61482,"visible":true,"origin":"","legend":"\u003cp\u003eFunctional enrichment of the genes targeted by the differentially expressed sRNAs. The targets active in the response to pathogens were assigned to the molecular function (MF) Gene Ontology (GO) groups.\u003c/p\u003e","description":"","filename":"8.png","url":"https://assets-eu.researchsquare.com/files/rs-3875675/v1/756066e4181de71ffb941259.png"},{"id":49855383,"identity":"d9ed2149-b4b1-40dd-be84-39df1965ad74","added_by":"auto","created_at":"2024-01-19 07:14:01","extension":"png","order_by":9,"title":"Figure 9","display":"","copyAsset":false,"role":"figure","size":78186,"visible":true,"origin":"","legend":"\u003cp\u003eAll milRNAs (A) and differentially expressed milRNAs (B) in the CP and ICP host–pathogen interactions involving three lines (D39, D33, and L318). The potential targets for\u003cstrong\u003e \u003c/strong\u003eall milRNAs (C) and differentially expressed milRNAs (D) in the CP and ICP host–pathogen interactions involving the three lines are presented.\u003c/p\u003e","description":"","filename":"9.png","url":"https://assets-eu.researchsquare.com/files/rs-3875675/v1/6cb2630681e753521a5073f4.png"},{"id":49992676,"identity":"dbc69d0c-af08-44d6-9315-84e52f5caeac","added_by":"auto","created_at":"2024-01-22 18:58:24","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1653242,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-3875675/v1/a9f0f416-cb5e-41a9-aeb0-c7d0418bed41.pdf"},{"id":49855378,"identity":"ee156fee-a3b9-48d7-a5cd-79e66df88be0","added_by":"auto","created_at":"2024-01-19 07:14:01","extension":"xlsx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":124361,"visible":true,"origin":"","legend":"","description":"","filename":"TableS1.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-3875675/v1/5ca8eaed9976bbe57d1f24df.xlsx"},{"id":49855377,"identity":"556caa83-ba09-43e6-bfb6-a5d7d93c08f4","added_by":"auto","created_at":"2024-01-19 07:14:01","extension":"xlsx","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":673893,"visible":true,"origin":"","legend":"","description":"","filename":"TableS2.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-3875675/v1/e821c7f95d129146f6358e82.xlsx"},{"id":49856056,"identity":"a2b0931b-a9fd-4ac0-959a-7ca75fccc4ae","added_by":"auto","created_at":"2024-01-19 07:30:01","extension":"xlsx","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":40049,"visible":true,"origin":"","legend":"","description":"","filename":"TableS3.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-3875675/v1/fcdcf5ecadb749e164f6d494.xlsx"}],"financialInterests":"The authors declare no competing interests.","formattedTitle":"\u003cp\u003eRole of sRNA molecules in the rye–leaf rust\u003cem\u003e \u003c/em\u003einteraction\u003c/p\u003e","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003eDespite being considered a cereal that is highly resistant to various environmental stresses, rye (\u003cem\u003eSecale cereale\u003c/em\u003e L.) is susceptible to leaf rust (LR), which can lead to yield losses of up to 40%. Leaf rust is caused by an airborne pathogen, namely the obligate biotrophic basidiomycete \u003cem\u003eP.\u003c/em\u003e f. sp. \u003cem\u003esecalis\u003c/em\u003e (\u003cem\u003ePrs\u003c/em\u003e) [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. The genetic basis of the interaction between rye and the LR pathogen has been studied for over 20 years. A total of 17 resistance genes (\u003cem\u003ePr1\u0026ndash;5\u003c/em\u003e, \u003cem\u003ePr-d\u0026ndash;f\u003c/em\u003e, \u003cem\u003ePr-i\u0026ndash;l\u003c/em\u003e, \u003cem\u003ePr-n\u003c/em\u003e, \u003cem\u003ePr-p\u003c/em\u003e, \u003cem\u003ePr-r\u003c/em\u003e, and \u003cem\u003ePr-t\u003c/em\u003e) located on chromosomes 1R, 2R, 4R, 6R, and 7R have been identified in studies that applied a Mendelian genetics approach [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. The recent sequencing of the rye genome allowed for increasingly precise analyses at the molecular level, resulting in the identification of the genes responsive to LR [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. Additionally, a recent genome-wide association study mapped five LR resistance-associated quantitative trait loci on chromosome arms 1RS (short arm of chromosome 1R), 1RL (long arm of chromosome 1R), 2RL, 5RL, and 7RS [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. The most important resistance-associated marker on chromosome arm 1RS was physically co-localized with molecular markers delimiting \u003cem\u003ePr3\u003c/em\u003e, which is a previously characterized gene. The second region on 7RS comprised numerous nucleotide-binding leucine-rich repeat (NLR)-encoding genes, one of which, provisionally denoted \u003cem\u003ePr6\u003c/em\u003e, was similar (at the protein level) to the wheat LR response-related \u003cem\u003eLr1\u003c/em\u003e gene on chromosome 5DL. Krępski et al. (2023) recently showed that several rye orthologs of wheat \u003cem\u003eLr\u003c/em\u003e genes (\u003cem\u003eScLr1_3\u003c/em\u003e, \u003cem\u003eScLr1_4\u003c/em\u003e, \u003cem\u003eScLr1_8\u003c/em\u003e, and \u003cem\u003eScRga2_6\u003c/em\u003e) may be critical for the rye response to LR [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe expression of many plant genes is regulated by small RNAs (sRNAs), which are non-coding sequences consisting of 20\u0026ndash;32 nucleotides (nt). The sRNAs have been divided into a few main classes according to the differences in their biogenesis and function. One of the main sRNA classes includes short interfering RNAs (siRNAs) and microRNAs (miRNAs). These molecules can bind to specific proteins in the RNA-induced silencing complex, which then recognizes and targets complementary mRNA sequences, leading to translational inhibition or transcript degradation [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. The results of an analysis of miRNAs classified according to new biosynthesis-based criteria have been published [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe large-scale miRNome sequencing of different plant and animal species has revealed distinct miRNA sequences originating from a single miRNA locus. These isoforms, which are more frequently referred to as isomiRs, represent variants with heterogenous ends, sequences, and lengths [\u003cspan additionalcitationids=\"CR11\" citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. IsomiRs are variants of canonical miRNAs that differ in their length or sequence. They can be classified into two groups: templated isomiRs, which are encoded in their respective genes and result from alternative DCL cleavage, and non-templated isomiRs, which are modified by enzymes after transcription and may have nucleotide additions, deletions, or substitutions. Moreover, isomiRs originating from the same gene may be active in different plant tissues or developmental stages [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. Studies on miRNA expression revealed fold-change dynamics, where the addition or loss of two or three nucleotides may result in completely different expression profiles [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. Relatively little is known about the role of isomiRs in biotic stress responses. Recent studies have confirmed the distinct differential expression of individual isomiRs from a single sequence during the complex response of tomato to the potato cyst nematode [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eSmall RNAs also exist in fungi. They were first detected in the filamentous fungus \u003cem\u003eNeurospora crassa\u003c/em\u003e in a study in which the transformation with homologous sequences led to the transient inactivation of endogenous transcripts [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Earlier research on tomato infected with \u003cem\u003eBotrytis cinerea\u003c/em\u003e identified fungal miRNA-like RNAs (milRNAs) targeting the host transcripts of many genes related to pathogen resistance [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. Since then, milRNAs have been identified in many fungal species, including plant parasites [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e, \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e]. During an infection, the sRNA from the host plant can enter the cells of the pathogen and silence the pathogen genes and vice versa [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e, \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. Small RNAs are a component of the fungal effectome [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e]. Several researchers have explored the interactions between the sRNAs of different \u003cem\u003ePuccinia\u003c/em\u003e fungal species and defense-related plant genes. For example, Mueth et al. (2015) identified \u003cem\u003ePuccinia striiformis\u003c/em\u003e RNAs and predicted the host target genes [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. Of the 22 target genes homologous to known plant stress resistance genes, seven encode nucleotide-binding site leucine-rich repeat (NBS-LRR) proteins and one encodes an ATP-binding cassette (ABC) transporter C family member. Wang et al. (2017) subsequently identified a novel sRNA (Pst-milR1) produced by \u003cem\u003eP. striiformis\u003c/em\u003e f. sp. \u003cem\u003etritici\u003c/em\u003e infecting wheat [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. Because the transcript of the pathogenesis-related gene \u003cem\u003ePr2\u003c/em\u003e is targeted by Pst-milR1, the authors concluded that Pst-milR1 functions as an effector that suppresses the wheat immune response to stripe rust.\u003c/p\u003e \u003cp\u003eStudies conducted over the last 10 years have shown that miRNAs induced by pathogens can be effective regulators of plant resistance (\u003cem\u003eR\u003c/em\u003e) genes encoding diverse proteins, including the following: NBS-LRR proteins; pathogenesis-related proteins; proteins associated with amino acid metabolism, replication, transport, and chelation of reactive oxygen species; cysteine-rich receptor-like protein kinases; ABC transporters; and other proteins related to plant immune responses [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e, \u003cspan additionalcitationids=\"CR25 CR26 CR27\" citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e]. Many \u003cem\u003eR\u003c/em\u003e genes expressed specifically in response to rusts have recently been identified in wheat and rye [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e, \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eIn terms of rust fungi, miRNAs regulate the defense responses of wheat infected with \u003cem\u003ePuccinia graminis\u003c/em\u003e f. sp. \u003cem\u003etritici\u003c/em\u003e, which causes stem rust [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e]. The authors suggested that different miRNAs might independently regulate the response of infected plants. In another study, Kumar et al. (2014) analyzed wheat inoculated with \u003cem\u003ePuccinia triticina\u003c/em\u003e (causing wheat LR) and detected 52 miRNAs, of which 22 were differentially expressed between the susceptible and resistant near-isogenic lines (NILs) [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. Additionally, there were more upregulated miRNAs in the susceptible NILs than in the resistant NILs. The authors identified 1,306 potential target genes, including genes encoding universal stress-responsive proteins (e.g., Pr1) or ABC transporters. However, there were no NLR-encoding genes targeted by miRNAs. In a later study, Kumar et al. (2017) identified 1,056 mature miRNAs (including 559 novel miRNAs) [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e]. Following the treatment with the pathogen, there were more miRNAs in the resistant plants than in the susceptible plants. However, the miRNA counts increased in the susceptible line, but decreased in the resistant line. The degradome analysis identified 701 predicted target genes associated with defense responses, signal transduction, development, metabolism, and transcriptional regulation. Interestingly, Feng et al. (2016) detected 1,316 miRNA candidates activated by \u003cem\u003eP. striiformis\u003c/em\u003e f. sp. \u003cem\u003etritici\u003c/em\u003e, which is responsible for yellow rust [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e]. For the upregulated miRNAs, the authors detected ata-miR1432-5p-targeted genes encoding proteins from the calcium ion-binding family and PC-3p-7484 (wheat specific miRNA)-targeted genes encoding various proteins (e.g., ubiquillin, mitochondrial-processing peptidase, tubby-like F-box protein, and other hypothetical proteins) [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e]. Nair et al. (2022) identified multiple miRNAs associated with \u003cem\u003eSr36\u003c/em\u003e and \u003cem\u003eLr45\u003c/em\u003e, which confer resistance to stem rust and LR, respectively [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e]. The mining for miRNAs revealed 26 known miRNAs and seven novel miRNAs in cultivars infected with stem rust as well as 22 known miRNAs and four novel miRNAs in LR-infected samples. The expression of various miRNAs originating from different subgenomes may reflect a potential relationship between the mechanisms underlying the defense responses to stem rust and LR mediated by specific \u003cem\u003eR\u003c/em\u003e genes.\u003c/p\u003e \u003cp\u003eIn plants, isomiRs are prospective host factors responsive to pathogen regulators. Specifically, they target genes involved in developmental processes, such as leucine catabolism, and genes encoding proteins that protect against stress conditions (e.g., GST). One of the isomiRs (isomiR523-4) detected in an earlier study reportedly targets \u003cem\u003ePuccinia\u003c/em\u003e mRNAs from the SunT superfamily, which is associated with defense responses [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. There is considerable information regarding the effects of rust infections on sRNAs during plant\u0026ndash;pathogen interactions, but there has been relatively little related research on rye.\u003c/p\u003e"},{"header":"2. Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1. Plant materials, growth conditions, and treatment with the rust pathogen\u003c/h2\u003e \u003cp\u003eThis study was conducted using the following three rye inbred lines: L318, which was bred by the Department of Plant Genetics, Breeding and Biotechnology, Warsaw University of Life Sciences (Warsaw, Poland), and D33 and D39, which were developed by Danko Plant Breeding, Ltd. (Poland). The inbred lines were selected according to the previously reported benzoxazinoid (BX) contents of field-grown plants after a natural vernalization period [\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e] and differential seedling stage responses to an artificial infection with a partially compatible \u003cem\u003ePrs\u003c/em\u003e isolate [\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eFour \u003cem\u003ePrs\u003c/em\u003e isolates (1/1.6, 83/2/2.2_5x, 81/r/5_5x, and 88/o/1_5x) that differed in terms of their interactions with host plants were used to inoculate lines D33, D39, and L318 (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eInteractions between rye inbred lines and \u003cem\u003ePrs\u003c/em\u003e isolates\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003ePrs\u003c/em\u003e isolate\u003c/p\u003e \u003cp\u003eRye line\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1/1.6\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003e83/2/2.2_5x\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003e81/r/5_5x\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003e88/o/1_5x\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD33\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eincompatible\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ecompatible\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD39\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eincompatible\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ecompatible\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eL318\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eincompatible\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ecompatible\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eRye seeds were sown on moistened tissue paper in Petri dishes, which were incubated in a darkened room for 2 days at 22\u0026deg;C. The germinated seeds were transferred to plastic pots (12 cm diameter) filled with sterilized peat substrate (10 germinated seeds per pot). The seedlings were cultivated for 10 days in a growth chamber at 22\u0026deg;C with a 16-h light (60 \u0026micro;mol m\u003csup\u003e\u0026minus;\u0026thinsp;2\u003c/sup\u003e s\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e)/8-h dark cycle. At 12 days after sowing, the plants were inoculated with a \u003cem\u003ePrs\u003c/em\u003e spore suspension or the control solution. The plants were then covered with black boxes to maintain darkness and 100% humidity and incubated for 24 h at 18\u0026deg;C. They were subsequently transferred to a phytotron with a 16 h (22\u0026deg;C)/8 h (18\u0026deg;C) cycle.\u003c/p\u003e \u003cp\u003eFor the treatment, spores from a single-spore \u003cem\u003ePrs\u003c/em\u003e isolate were suspended in Novec\u0026trade; 7100 engineered fluid (3M, Maplewood, USA) at a density of 1 mg/ml. The control (mock-inoculated) plants were inoculated with Novec\u0026trade; 7100 engineered fluid lacking \u003cem\u003ePrs\u003c/em\u003e spores. The treatments were performed using leaf glass diffusers (Carl Roth, Karlsruhe, Germany).\u003c/p\u003e \u003cp\u003eFor each pot, leaves were collected from five plants at 20 hpt and from the other five plants at 36 hpt. All leaf samples were collected in tubes, frozen immediately in liquid nitrogen, and stored at \u0026minus;\u0026thinsp;80\u0026deg;C. The leaves were weighed and then 100 mg leaf material was ground in a mortar for the RNA extraction using the mirVana\u0026trade; miRNA Isolation Kit (ThermoFisher, Waltham, USA). The RNA condition and concentration were determined using the Qubit 4 fluorometer (ThermoFisher) prior to the next-generation sequencing (NGS) analysis, which was completed by Genomed S.A. (Warsaw, Poland) as previously described [\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e].\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2. Bioinformatics analyses\u003c/h2\u003e \u003cp\u003eSequencing data in FASTQ files were checked for quality using FastQC (version 0.11.8), while sequencing adapters were trimmed using Cutadapt (version 3.0) [\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e, \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e]. The reads were mapped to the rye genome with Bowtie (version 1.3) [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e]. In the next step, miRDeep2 (version 0.1.3) was used to detect known miRNAs (i.e., deposited in miRbase) and novel miRNAs [\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e]. The quantitative analysis was performed using the DESeq2 package (version 1.32.0) from the Bioconductor repository [\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e]. The ShortStack software (version 3.5.8) was used to separate miRNAs from among the sRNAs according to the latest guidelines [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e]. The sRNAs were quantitatively analyzed using the edgeR package (version 3.34.1) from the Bioconductor repository [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe isomiRs were detected manually using miRNA sequences aligned by Bowtie (version 1.3). The \u003cem\u003ePuccinia\u003c/em\u003e-specific milRNAs were selected as follows. Sequencing reads from the mock-inoculated and \u003cem\u003ePrs\u003c/em\u003e-infected rye leaves were mapped to the \u003cem\u003ePrs\u003c/em\u003e genome as well as to the rye Lo7 genome. The Venn algorithm was used to select \u003cem\u003ePrs\u003c/em\u003e-specific sRNA sequences. The detected miRNAs were compared with those in the miRBase and PmiREN 2.0 databases [\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e, \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e]. The psRNATarget web server was used to identify potential sRNA targets; the default parameters were applied, with the exception of maximum expectation, which was set to 3 [\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e, \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e]. For the functional enrichment analysis, the standard parameters of g:Profiler were used [\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e, \u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e].\u003c/p\u003e \u003c/div\u003e"},{"header":"3. Results","content":"\u003cdiv id=\"Sec6\" class=\"Section2\"\u003e\n \u003ch2\u003e3.1. Identification of sRNAs in rye\u003c/h2\u003e\n \u003cp\u003eA comprehensive sRNA analysis pipeline (Fig. \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e) was developed to identify sRNAs in rye. The Illumina sRNA sequencing generated 642,040,931 reads, of which 12.01% (77,123,956) were unique reads 18\u0026ndash;35 nt long (Fig. \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e). The most common read length was 32 nt, but the most diverse sRNAs consisted of 24 nt. The fact the reads consisting of 32 nt were the most abundant was likely due to the tRFs/tsRNAs, which are regulatory RNAs originating from tRNA segments. The sequences comprising 21\u0026ndash;24 nt, which were mostly miRNAs and other sRNAs, were also over-represented. The rye sRNAs were recently surveyed in the Lo7 sequencing project, during which in-house scripts were used for identifying and annotating 1,005 different sRNA sequences [\u003cspan class=\"CitationRef\"\u003e4\u003c/span\u003e]. In the current study, two algorithms in the miRDeep2 and ShortStack programs detected 65 and 462 sRNAs, respectively. Next, the sRNAs were compared with the Lo7 sRNA sequences and with all other sRNAs in the miRBase and PmiREN databases. A Venn diagram was constructed to separate experiment- and database-specific sRNA sequences (Fig. \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e). In total, 43 and 390 novel sRNA sequences were detected using miRDeep2 and ShortStack, respectively.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec7\" class=\"Section2\"\u003e\n \u003ch2\u003e3.2. Detecting differentially expressed sRNAs at different time-points during the rye\u0026ndash;rust interaction\u003c/h2\u003e\n \u003cp\u003eThis study initially focused on the dynamic expression of sRNAs in three rye lines infected with LR. Two algorithms detected 316 sequences, with miRDeep2 and ShortStack detecting 14 and 302 sequences, respectively. The subsequent analysis focused on the differentially expressed sRNAs and whether they decrease or increase the expression of genes important for plant defenses against fungal pathogens. The following threshold was applied for the expression analysis: |log\u003csub\u003e2\u003c/sub\u003e(fold change)| \u0026gt;1 (Fig. \u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e).\u003c/p\u003e\n \u003cp\u003eNext, the compatible (CP) and incompatible (ICP) host\u0026ndash;pathogen interactions were analyzed and compared to detect the sRNAs influencing gene expression during these two interactions. For the CP interaction, there were 225 sRNAs and seven miRNAs, whereas for the ICP interaction, there were only 57 sRNAs and one miRNA. Notably, 12 sRNAs were common to both interactions. The sRNA expression dynamics varied among the time-points, especially at 20 and 36 h post-treatment (hpt). During this period, the number of differentially expressed sRNAs increased by 84 (from 104 to 188), while the number of differentially expressed miRNAs decreased from five to two. Additionally, similar to the previous findings, there were several sequences that were common between the two time-points, including two sRNAs and two miRNAs (Fig. \u003cspan class=\"InternalRef\"\u003e5\u003c/span\u003eA and B).\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec8\" class=\"Section2\"\u003e\n \u003ch2\u003e3.3. Host isomiRs responsive to the pathogen\u003c/h2\u003e\n \u003cp\u003eDifferentially expressed miRNAs and their highly expressed cognate sequences (isomiRs) were manually selected (Fig. \u003cspan class=\"InternalRef\"\u003e7\u003c/span\u003eA). They differed regarding their sequences as well as in terms of the number of specific target sequences (i.e., 10, 10, 19, and 72; Fig. \u003cspan class=\"InternalRef\"\u003e7\u003c/span\u003eB). The differences in the isomiR sequences influence target recognition, thereby increasing the range of targeted transcripts important for plant defenses against pathogens.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec9\" class=\"Section2\"\u003e\n \u003ch2\u003e3.4. Predicted targets for the rye sRNAs\u003c/h2\u003e\n \u003cp\u003eThe psRNATarget tool and the rye Lo7 transcriptome were used for predicting sRNA targets. The resulting hits were enriched among the highly expressed |log\u003csub\u003e2\u003c/sub\u003e(fold change)| \u0026gt; 2and strongly expressed (baseMean 50) transcripts (Table \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e). Multiple targets with known potential roles in host defenses against rust infections were detected (Table S1), including targets encoding NBS-LRR disease resistance proteins, receptor kinases, enzymes associated with sugar transport and metabolism, and ABC transporters. More specifically, some of the targets encoded a glutamate receptor, which mediate electrical signaling in response to fungal infections or inhibit fungal growth [\u003cspan class=\"CitationRef\"\u003e48\u003c/span\u003e].\u003c/p\u003e\n \u003cp\u003eTo highlight the functional diversity among the predicted targets of the rye sRNAs, we performed a gene functional enrichment analysis using g:profiler [\u003cspan class=\"CitationRef\"\u003e46\u003c/span\u003e] (Supplementary Table S1)\u003c/p\u003e\n \u003cp\u003eTable 2. Selected sRNAs and their highly expressed targets or targets expressed in all three rye lines (L318, D33, and D39)\u003c/p\u003e\n \u003cp\u003e\u003cimg src=\"data:image/png;base64,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\" width=\"756\" height=\"232\"\u003e\u003cbr\u003e\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec10\" class=\"Section2\"\u003e\n \u003ch2\u003e3.5. milRNA dynamics and their predicted targets in rye\u003c/h2\u003e\n \u003cp\u003eA total of 53 LR-specific sequences were detected. The CP and ICP rust\u0026ndash;host interactions were due to different \u003cem\u003ePsr\u003c/em\u003e isolates. Additionally, the control rye lines were free of \u003cem\u003ePsr\u003c/em\u003e. Thus, milRNA dynamics were examined at 20 and 36 h posttreatment (hpt). There were more differentially expressed milRNAs in the CP interactions than in the ICP interactions. Line D39 had the most differentially expressed milRNAs (20), followed by lines L318 (five) and D33 (three) (Fig. \u003cspan class=\"InternalRef\"\u003e9\u003c/span\u003eA). For the ICP interactions, there was only one specific sequence in lines L318 and D33. There were many stably expressed milRNAs in all lines and interactions, but there were more in the CP interactions (106, 118, and 133 in D39, L318, and D33, respectively) than in the ICP interactions (77, 58, and 90 in D39, L318, and D33, respectively) (Fig. \u003cspan class=\"InternalRef\"\u003e9\u003c/span\u003eB). Not all active milRNAs have to be differentially expressed and have important regulatory effects on host gene expression. There were more potential milRNA targets in the CP interactions than in the ICP interactions. The potential targets for all and differentially expressed milRNAs were compared (Fig. \u003cspan class=\"InternalRef\"\u003e9\u003c/span\u003eC and D). The full list of targets is provided in supplementary Table S2 and S3.\u003c/p\u003e\n\u003c/div\u003e"},{"header":"4. Discussion","content":"\u003cp\u003eIn this study, sRNAs, miRNAs, and milRNAs were detected and analyzed in terms of their expression dynamics in rye\u0026ndash;\u003cem\u003ePrs\u003c/em\u003e interactions, with a particular focus on their potential targets and regulatory effects on the host transcriptome. There are reports describing the mobilization of sRNAs due to LR infections of wheat [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e, \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e, \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e]. Unfortunately, similar studies have not been conducted for rye, even though it is the closest relative of wheat.\u003c/p\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003e4.1. Comparison of the effectiveness of different algorithms for identifying sRNAs\u003c/h2\u003e \u003cp\u003eThe recently published genomes and transcriptomes of two rye varieties, Chinese Weining and European Lo7 [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e, \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e], significantly accelerated the molecular research on rye. The study conducted by Rabanus-Wallace et al. (2021) explored many aspects of rye transcriptomics, including sRNAs. The authors used their own in-house script to detect sRNAs, which were imprecisely referred to as miRNAs, and identified 13,238 sequences belonging to 90 families. The process underlying miRNA biogenesis is relatively well known and was used to accurately identify miRNAs [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. A software package called ShortStack was developed on the basis of a published article [\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e]. However, one of the most popular tools for detecting sRNAs is the miRDeep2 software package [\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e]. Since its release, it has been used in multiple studies on sRNAs in different organisms. In the current study, the rye sRNAome was analyzed using both ShortStack and miRDeep2, which involve different algorithms, and the resulting sequences were compared with rye transcriptome sequencing data and the sequences deposited in the largest sRNA databases (miRBase and PmiREN). Using this approach, 433 novel sRNA sequences were identified in rye. In the subsequent sRNA transcriptomic analysis, milRNAs synthesized only in response to LR were examined. Because of the lack of an available whole \u003cem\u003ePrs\u003c/em\u003e transcriptome, the sRNAome was filtered and screened for milRNAs. This strategy involved dual genome mapping. More specifically, the sequencing reads generated for the LR-infected and mock-inoculated rye leaves were preliminarily mapped to both the \u003cem\u003ePrs\u003c/em\u003e and rye genomes. In addition, a Venn diagram was constructed to separate the resulting sRNA sequences. ShortStack was used to identify milRNAs.\u003c/p\u003e \u003cp\u003eUsing both algorithms to detect sRNAs enabled us to identify more differentially expressed sRNAs than if only one algorithm was used and to analyze their target interactions. Eight miRNAs and 293 sRNAs were detected using ShortStack, whereas 14 sRNAs were detected using miRDeep2. Clearly distinguishing between sRNA and miRNA sequences is conducive to accurately predicting their functions.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003e4.2. Different interactions between plant gene and s/miRNA transcripts\u003c/h2\u003e \u003cp\u003eThe thorough investigation of the interplay between sRNAs and the targeted mRNAs in plants indicated whole sRNA sequences interact with compatible parts of mRNAs. There are many reports describing the interaction between one sRNA and many targets and vice versa. A single target sequence may have many so-called miRNA-responsive elements (MREs) and interact with many different sRNAs. This phenomenon involving complex multidimensional interactions between different molecules, which is common in diverse biochemical pathways, enhances the regulation of gene expression and the coordination of host responses to pathogen attacks. This interplay among sRNAs, long non-coding RNAs (lncRNAs), and mRNAs results in diverse sRNA\u0026ndash;target dynamics (upregulated or downregulated) [\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e]. The lncRNAs interact with miRNAs either as target mimics that form competing endogenous RNAs (ceRNAs) or as target lncRNAs that are spliced by miRNAs. Additionally, miRNAs can bind to transcribed pseudogenes and lncRNAs through MREs, which compete with these miRNAs for mRNA-binding sites. These RNAs function as molecular sponges or decoys and suppress the targeting of mRNAs by miRNAs, which explains why they are referred to as ceRNAs. To be able to serve as a ceRNA, the lncRNA must contain an MRE that is partially complementary to the miRNA sequence [\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e]. The effect of the interplay between miRNAs and lncRNAs has been investigated in many host\u0026ndash;pathogen interactions, including the interaction between wheat and the pathogens responsible for rusts, including LR [\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e], yellow rust [\u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e], and stripe rust [\u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eIn the present study, the following distinct interactions between the sRNAs and their targets were detected in the same comparison (i.e., same time-point): (i) sRNAs and their targets were upregulated (i.e., +/+); (ii) sRNAs and their targets were downregulated (i.e., \u0026minus;/\u0026minus;); (iii) sRNAs and their targets had the opposite changes (i.e., +/\u0026minus; or \u0026minus;/+). In all models, many of the targets were for genes with confirmed roles in the immune response to rust fungi, including genes encoding CYP450 [\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e], beta-glucanase [\u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e55\u003c/span\u003e, \u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e], glycosyltransferase [\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e], acid invertase [\u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e], and kaurene synthase [\u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e59\u003c/span\u003e] in the +/+ model; chlorophyll \u003cem\u003ea\u003c/em\u003e/\u003cem\u003eb\u003c/em\u003e-binding protein [\u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e60\u003c/span\u003e], beta-glucosidase [\u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e61\u003c/span\u003e], fatty acid hydroxylase [\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e], and SAUR auxin-responsive protein [\u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e63\u003c/span\u003e] in the \u0026minus;/\u0026minus; model; and glutamate receptor in oat [\u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e64\u003c/span\u003e] and ammonium transporter [\u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e65\u003c/span\u003e] in the +/\u0026minus; model. No immune response-specific genes were identified in the \u0026minus;/+ model. However, the biological significance of these different types of interactions is unclear and may reflect the considerable complexity of the systems regulating various types of non-coding RNA molecules [\u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e66\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe interactions between differentially expressed sRNAs and their targets described in the previous paragraph were for the same time-point, but their interactions can be more dynamic. For example, high or low sRNA levels at one time-point can influence the expression of their targets at a later time-point. Thus, the sRNAs and their targets were examined at different time-points. The downregulated sRNAs at 20 hpt affected the upregulated transcripts of important biotic stress-responsive genes at 36 hpt. Earlier research indicated some of these genes encode receptor-like kinases, proteins involved in detecting pathogens, and ABC transporters belonging to a large family with known roles in the resistance to multiple fungal pathogens in cereals [\u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e67\u003c/span\u003e]. In the reverse configuration, upregulated sRNAs at 20 hpt affected downregulated targets at 36 hpt where there were fewer target transcripts, one of which encodes a longifolia protein that regulates xyloglucan endotransglycosylases/hydrolases (XTHs). The XTHs and xyloglucan endotransglycosylases (XETs) are enzymes that repair hemicellulose. Decreases in the expression of the corresponding genes lead to decreases in XET activity and xyloglucan endotransglucosylation, ultimately altering the xyloglucan structure [\u003cspan citationid=\"CR68\" class=\"CitationRef\"\u003e68\u003c/span\u003e]. These changes might promote fruit softening and wall disassembly, facilitating fungal colonizations and infections [\u003cspan citationid=\"CR69\" class=\"CitationRef\"\u003e69\u003c/span\u003e]. Another enzyme encoded by a transcript in the reverse configuration group was identified as a strictosidine synthase family member that regulates the secondary metabolism associated with plant responses to pathogens.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003e4.3. Identification of s/miRNAs in CP and ICP interactions\u003c/h2\u003e \u003cp\u003eA new era in plant\u0026ndash;pathogen interaction-based research was initiated after Flor determined the flax response to a flax rust (\u003cem\u003eMelampsora lini\u003c/em\u003e) infection and described his gene-for-gene theory in his seminal paper [\u003cspan citationid=\"CR70\" class=\"CitationRef\"\u003e70\u003c/span\u003e]. The gene-for-gene theory explains the genetic basis of the CP and ICP interactions between plants and pathogens. The theory states that a plant is resistant to a pathogen if it has a resistance (\u003cem\u003eR\u003c/em\u003e) gene that matches an avirulence (\u003cem\u003eAvr\u003c/em\u003e) gene from the pathogen. For wheat, which is closely related to rye, more than 200 \u003cem\u003eR\u003c/em\u003e genes conferring resistance to rusts (including LR) have been identified (Catalogue of Gene Symbols for Wheat; \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://wheat.pw.usda.gov/GG3/wgc\u003c/span\u003e\u003cspan address=\"https://wheat.pw.usda.gov/GG3/wgc\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e). The expression of many of these genes may be regulated by sRNAs.\u003c/p\u003e \u003cp\u003eThe changes induced by two \u003cem\u003ePrs\u003c/em\u003e isolates associated with CP and ICP interactions were compared. There were more differentially expressed sRNAs and miRNAs in the CP interactions (225 and seven, respectively) than in the ICP interactions (seven and one, respectively), suggesting the response to \u003cem\u003ePrs\u003c/em\u003e is more complex during CP interactions than during ICP interactions. The greater number of regulatory sRNAs in CP interactions than in ICP interactions may indicate an increase in the number of targeted transcripts and a more dynamic and complex regulation of the host response during CP interactions. These findings are inconsistent with the results of an earlier study on the potato response to two \u003cem\u003ePhytophthora infestans\u003c/em\u003e isolates, in which there were more differentially expressed transcripts in the ICP interaction than in the CP interaction. The reason for this discrepancy might be related to the differences in the selected time-points between the earlier study [\u003cspan citationid=\"CR71\" class=\"CitationRef\"\u003e71\u003c/span\u003e] and the current study.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003e4.4. Host genotype-specific targets\u003c/h2\u003e \u003cp\u003eThe three rye lines included in the present study (L318, D33, and D39) differ regarding their response to LR under field and phytotron conditions [\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e, \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e]. Lines L318 and D39 are respectively the most susceptible and resistant to LR. In addition, 73, 107, and 55 sRNAs targeting 182, 544, and 116 potential mRNAs were detected in lines L318, D33, and D39, respectively. All but one of the sRNAs were specific to a particular rye line. Interestingly, the most susceptible line (L318) had the most upregulated and downregulated sRNA sequences (62 and 12, respectively). In contrast, in the most resistant line (D39), there were 32 and 23 upregulated and downregulated sRNAs, respectively (i.e., relatively even number of altered sRNAs). This may be related to the number of targets detected in L318 and D39 (i.e. 182 and 116, respectively). Compared with the resistant line, the susceptible line may require the activation of more genes following an LR infection.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003e4.5. IsomiRs: the influence of small sequence differences on target selection\u003c/h2\u003e \u003cp\u003eThe identified isomiRs in rye infected with a fungal pathogen included 20 and 21 nt sequences with 14 nt in common. The isomiRs that differed by only one nucleotide were observed to target different mRNAs in the host. The longer sequence (UGUGUUCUCAGGUCGCCCCCG) targeted the transcripts of 11 endoglucanase genes and two superoxide dismutase genes, which are well known for their roles in plant defenses. Endoglucanases are required for the formation of cellulose microfibrils, but recent research indicated fungi can hijack a ubiquitous plant apoplastic endoglucanase, leading to the release of a reactive oxygen species-scavenging β-glucan decasaccharide that subverts immune responses [\u003cspan citationid=\"CR72\" class=\"CitationRef\"\u003e72\u003c/span\u003e]. The shorter sequence (UCAGGUCGCCCCCGCUGGAG) targeted the transcripts of 16 transcription factor genes (e.g., MADS box gene). Some of the proteins encoded by the targets contribute to plant stress responses. SPIRAL1 plays a vital role in stress-induced microtubule disassembly, which is required for cell wall rearrangements and anisotropic cell growth. The AP2-like ethylene-responsive transcription factor is a member of a protein group that functions at a hub in hormone and stress signaling pathways [\u003cspan citationid=\"CR73\" class=\"CitationRef\"\u003e73\u003c/span\u003e]. Chloroplastic chlorophyll \u003cem\u003ea\u003c/em\u003e/\u003cem\u003eb\u003c/em\u003e-binding proteins decrease oxidative stress, while CsAtPR5 belongs to a pathogen-responsive protein family.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec17\" class=\"Section2\"\u003e \u003ch2\u003e4.6. Role of cross-kingdom RNA interference in CP and ICP interactions\u003c/h2\u003e \u003cp\u003eCross-kingdom RNA interference (ck-RNAi) is an sRNA-mediated gene silencing mechanism affecting hosts and pathogens [\u003cspan citationid=\"CR74\" class=\"CitationRef\"\u003e74\u003c/span\u003e]. Fungi and oomycetes lack typical miRNAs, but similar sRNA sequences were identified in the fungus \u003cem\u003eN. crassa\u003c/em\u003e and designated as milRNAs. This new class of small non-coding RNAs was subsequently detected in diverse fungi and oomycetes, including plant pathogenic species. In fact, the first example of a non-protein effector influencing the response of tomato to gray mold caused by \u003cem\u003eB. cinerea\u003c/em\u003e was a fungal milRNA that silenced the transcripts of host defense response-related genes [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. Since then, similar examples of pathogen factors silencing host genes have been reported for various agriculturally important crops, including wheat, barley, rice, and tomato [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e, \u003cspan citationid=\"CR75\" class=\"CitationRef\"\u003e75\u003c/span\u003e, \u003cspan citationid=\"CR76\" class=\"CitationRef\"\u003e76\u003c/span\u003e]. The host targets encoded several proteins involved in responses to pathogen attacks, including pathogenesis-related proteins with NB-ARC domains. The transcripts targeted by milRNAs encoded lipid metabolism-related enzymes crucial for plant responses to biotic stress. For example, fatty acyl reductase catalyzes the NADPH-dependent reduction of fatty acyl-CoA or acyl-ACP substrates to primary fatty alcohols, which are intermediate metabolites or metabolic end products that contribute to the formation of plant extracellular protective lipid barriers (e.g., cuticular wax, sporopollenin, suberin, and taproot wax) [\u003cspan citationid=\"CR77\" class=\"CitationRef\"\u003e77\u003c/span\u003e]. Notably, one of the targets was a transcript encoding patatin. Both patatin and patatin-like proteins (PLPs) are involved in plant responses to pathogens, especially fungi. A recent study showed that in \u003cem\u003eNicotiana attenuata\u003c/em\u003e, \u003cem\u003eNaPLP\u003c/em\u003e expression increases substantially following an infection by \u003cem\u003eAlternaria alternata\u003c/em\u003e [\u003cspan citationid=\"CR78\" class=\"CitationRef\"\u003e78\u003c/span\u003e]. Another targeted transcript, which encodes a methyl esterase, was downregulated by the upregulated milRNAs. This finding is in accordance with the results of an earlier study in which methyl esterase 1 (PvMES1) promoted the salicylic acid pathway and enhanced the resistance of common bean to Fusarium wilt [\u003cspan citationid=\"CR79\" class=\"CitationRef\"\u003e79\u003c/span\u003e]. The basic helix-loop-helix (bHLH) DNA-binding proteins are involved in the biosynthesis of diterpenoid phytoalexins, which might be induced in blast-infected rice[\u003cspan citationid=\"CR80\" class=\"CitationRef\"\u003e80\u003c/span\u003e]. Interestingly, milRNA can reportedly target transcripts encoding histone 3, which is post-transcriptionally modified in response to different biotic stresses [\u003cspan citationid=\"CR81\" class=\"CitationRef\"\u003e81\u003c/span\u003e].\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec18\" class=\"Section2\"\u003e \u003ch2\u003e4.7. Utility of sRNAs for protecting crops and promoting sustainable agriculture\u003c/h2\u003e \u003cp\u003eThe global application of pesticides on crops reached 2.66 tons in 2020. Their harmful effects on the environment are well known [\u003cspan citationid=\"CR82\" class=\"CitationRef\"\u003e82\u003c/span\u003e]. The research community has proposed many environmentally friendly alternatives to pesticides, one of which involves host-induced gene silencing (HIGS). In this technique, a plant is transformed with a target gene (partial or complete sequence) originating from a pathogen. The expression of the transgene triggers RNA interference, thereby effectively silencing the expression of an essential pathogen gene. Recent research confirmed that HIGS is effective for controlling hemibiotrophic or necrotrophic fungal pathogens (e.g., \u003cem\u003eVerticillium dahliae\u003c/em\u003e and \u003cem\u003eB. cinerea\u003c/em\u003e) as well as the obligate biotroph \u003cem\u003eP. triticina\u003c/em\u003e and \u003cem\u003eFusarium\u003c/em\u003e species. A simpler technique that does not require the production of transgenic plants is spray-induced gene silencing (SIGS), which involves spraying host plants with the RNA of pathogen genes. After the SIGS technique was first developed in 2016, it was used to protect certain crops from pathogens [\u003cspan citationid=\"CR83\" class=\"CitationRef\"\u003e83\u003c/span\u003e], including canola infected with \u003cem\u003eB. cinerea\u003c/em\u003e and \u003cem\u003eSclerotinia sclerotiorum\u003c/em\u003e, barley infected with \u003cem\u003eFusarium\u003c/em\u003e species, and wheat infected with \u003cem\u003eRhizoctonia cerealis\u003c/em\u003e. The RNAs used to silence genes were sequences targeting transcripts related to thioredoxin reductase and ergosterol biosynthesis. Interestingly, the earlier study also provided evidence that SIGS may have the same effects as a semi-systemic fungicide. When plants were sprayed with CYP3-dsRNAs, they were protected from the disease even in parts that were not sprayed (e.g., distal leaf parts). Moreover, CYP3-dsRNAs were detected in the sprayed and unsprayed leaf parts. The subsequent analysis of dsRNA localization using green fluorescent dye ATTO 488-labeled dsRNAs indicated that the dsRNAs at the leaf surface were taken up by plant cells and then moved to different parts of the plant, including the stomata, xylem cells, companion cells, phloem parenchyma cells, mesophyll cells, and trichomes [\u003cspan citationid=\"CR84\" class=\"CitationRef\"\u003e84\u003c/span\u003e]. Accordingly, these dsRNAs are eco-friendly fungicides that are safe for the environment and humans. There has been research conducted to address some of the issues that may need to be addressed, including the stability and efficient delivery of sRNAs to pathogens. To increase RNA stability, researchers docked an antiviral dsRNA to double hydroxide clay nanosheets, which resulted in enhanced protection against phytopathogenic viruses [\u003cspan citationid=\"CR85\" class=\"CitationRef\"\u003e85\u003c/span\u003e]. In addition, the use of artificial vesicles or liposomes to protect RNAs may improve SIGS techniques for protecting plants and developing antifungal compounds, especially for fungi that can efficiently take up extracellular vesicles (EVs) [\u003cspan citationid=\"CR84\" class=\"CitationRef\"\u003e84\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eIn conclusion, we detected rye and \u003cem\u003ePrs\u003c/em\u003e sRNAs that target vital transcripts for host\u0026ndash;pathogen interactions, including transcripts encoding glucanases, proteins with NBS-LRR domains, peroxidases, and different transcription factors. Furthermore, newly detected \u003cem\u003ePrs\u003c/em\u003e milRNAs targeting host resistance-related transcripts (e.g., those encoding NBS-LRR proteins, peroxidases, ABC transporters, and pathogenesis-related proteins) may be silenced or eliminated using Short Tandem Target Mimic (STTMs) as part of a SIGS-based strategy to prevent rust infections. This approach may protect rye against fungal infections (at least LR) and is fully compliant with the current EU regulations that favor this type of agro-technical solution over the genetic modification of crop plants.\u003c/p\u003e\u003c/div\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eCRediT authorship contribution statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMarek Daniel Koter:\u003c/strong\u003e Data analysis, Writing. \u003cstrong\u003eMateusz Matuszkiewicz\u003c/strong\u003e: Data analysis. \u003cstrong\u003eMonika Rakoczy-Trojanowska\u003c/strong\u003e: Conceptualization, Writing - review \u0026amp; editing, Funding acquisition.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDeclaration of competing interest\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis work was financially supported by the Polish National Science Centre (grant number 2018/31/B/NZ9/00439).\u003c/p\u003e\n\u003cp\u003eWe thank Edanz (www.edanz.com/ac) for editing a draft of this manuscript.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eK. Peksa, B. Bankina, Characterization of Puccinia recondita, the causal agent of brown rust: A review, Research for Rural Development 2 (2019) 70\u0026ndash;76.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eO.V. Solodukhina, V.D. Kobylyanskij, Problems of winter rye breeding for resistance to leaf and stem rusts, Plant Breed Seed Sci. 48(2/2) (2003) 87\u0026ndash;96.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eS.R. Roux, B. Hackauf, B. Ruge-Wehling, A. Linz, P. Wehling, Exploitation and comprehensive characterization of leaf rust resistance in rye, Vortr. Pflanzenz\u0026uuml;chtung 71 (2007) 144\u0026ndash;150.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM.T. Rabanus-Wallace, B. Hackauf, M. Mascher, T. Lux, T. Wicker, H. Gundlach, M. Baez, A. Houben, K.F.X. Mayer, L. Guo, J. Poland, C.J. Pozniak, S. Walkowiak, J. Melonek, C.R. Praz, M. Schreiber, H. Budak, M. Heuberger, B. Steuernagel, B. Wulff, A. B\u0026ouml;rner, B. Byrns, J. Č\u0026iacute;žkov\u0026aacute;, D.B. Fowler, A. Fritz, A. Himmelbach, G. Kaithakottil, J. Keilwagen, B. Keller, D. Konkin, J. Larsen, Q. Li, B. Myśk\u0026oacute;w, S. Padmarasu, N. Rawat, U. Sesiz, S. Biyiklioglu-Kaya, A. Sharpe, H. Šimkov\u0026aacute;, I. Small, D. Swarbreck, H. Toegelov\u0026aacute;, N. Tsvetkova, A.V. Voylokov, J. Vr\u0026aacute;na, E. Bauer, H. Bolibok-Bragoszewska, J. Doležel, A. Hall, J. Jia, V. Korzun, A. Laroche, X.F. Ma, F. Ordon, H. \u0026Ouml;zkan, M. Rakoczy-Trojanowska, U. Scholz, A.H. Schulman, D. Siekmann, S. Stojałowski, V.K. Tiwari, M. Spannagl, N. Stein, Chromosome-scale genome assembly provides insights into rye biology, evolution and agronomic potential, Nat Genet 53(4) (2021) 564\u0026ndash;573.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eN.M. Vendelbo, K. Mahmood, P. Sarup, M.S. Hovm\u0026oslash;ller, A.F. Justesen, P.S. Kristensen, J. Orabi, A. Jahoor, Discovery of a Novel Leaf Rust (Puccinia recondita) Resistance Gene in Rye (Secale cereale L.) Using Association Genomics, Cells 11(1) (2021) 64.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eN.M. Vendelbo, K. Mahmood, B. Steuernagel, B.B.H. Wulff, P. Sarup, M.S. Hovm\u0026oslash;ller, A.F. Justesen, P.S. Kristensen, J. Orabi, A. Jahoor, Discovery of Resistance Genes in Rye by Targeted Long-Read Sequencing and Association Genetics, Cells 11(8) (2022).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eT. Krępski, M. Olechowski, I. Samborska-Skutnik, M. Święcicka, A. Grądzielewska, M. Rakoczy-Trojanowska, Identification and characteristics of wheat Lr orthologs in three rye inbred lines, PLoS One 18(7) (2023) e0288520.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eP. Guleria, M. Mahajan, J. Bhardwaj, S.K. Yadav, Plant Small RNAs: Biogenesis, Mode of Action and Their Roles in Abiotic Stresses, Genomics, Proteomics \u0026amp; Bioinformatics 9(6) (2011) 183\u0026ndash;199.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM.J. Axtell, B.C. Meyers, Revisiting Criteria for Plant MicroRNA Annotation in the Era of Big Data, Plant Cell 30(2) (2018) 272\u0026ndash;284.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eH. Budak, M. Kantar, R. Bulut, B.A. Akpinar, Stress responsive miRNAs and isomiRs in cereals, Plant Sci 235 (2015) 1\u0026ndash;13.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eE.M. Fard, S. Moradi, N.N. Salekdeh, B. Bakhshi, M.R. Ghaffari, M. Zeinalabedini, G.H. Salekdeh, Plant isomiRs: origins, biogenesis, and biological functions, Genomics 112(5) (2020) 3382\u0026ndash;3395.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eC.T. Neilsen, G.J. Goodall, C.P. Bracken, IsomiRs\u0026ndash;the overlooked repertoire in the dynamic microRNAome, Trends Genet 28(11) (2012) 544\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eC. Lelandais-Bri\u0026egrave;re, L. Naya, E. Sallet, F. Calenge, F. Frugier, C. Hartmann, J. Gouzy, M. Crespi, Genome-wide Medicago truncatula small RNA analysis revealed novel microRNAs and isoforms differentially regulated in roots and nodules, Plant Cell 21(9) (2009) 2780\u0026ndash;96.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM.D. Koter, M. Święcicka, M. Matuszkiewicz, A. Pacak, N. Derebecka, M. Filipecki, The miRNAome dynamics during developmental and metabolic reprogramming of tomato root infected with potato cyst nematode, Plant Sci 268 (2018) 18\u0026ndash;29.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM. Święcicka, W. Skowron, P. Cieszyński, J. Dąbrowska-Bronk, M. Matuszkiewicz, M. Filipecki, M.D. Koter, The suppression of tomato defence response genes upon potato cyst nematode infection indicates a key regulatory role of miRNAs, Plant Physiol Biochem 113 (2017) 51\u0026ndash;55.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eN. Romano, G. Macino, Quelling: transient inactivation of gene expression in Neurospora crassa by transformation with homologous sequences, Mol Microbiol 6(22) (1992) 3343\u0026ndash;53.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eA. Weiberg, M. Wang, M. Bellinger, H. Jin, Small RNAs: a new paradigm in plant-microbe interactions, Annu Rev Phytopathol 52 (2014) 495\u0026ndash;516.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eB. Wang, Y. Sun, N. Song, M. Zhao, R. Liu, H. Feng, X. Wang, Z. Kang, \u003cem\u003ePuccinia striiformis\u003c/em\u003e f. sp. \u003cem\u003etritici\u003c/em\u003e microRNA-like RNA 1 (Pst-milR1), an important pathogenicity factor of Pst, impairs wheat resistance to Pst by suppressing the wheat pathogenesis-related 2 gene, New Phytologist 215(1) (2017) 338\u0026ndash;350.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eB. Wang, Y. Sun, N. Song, M. Zhao, R. Liu, H. Feng, X. Wang, Z. Kang, \u003cem\u003ePuccinia striiformis\u003c/em\u003e f. sp. \u003cem\u003etritici\u003c/em\u003e microRNA-like RNA 1 (Pst-milR1), an important pathogenicity factor of Pst, impairs wheat resistance to Pst by suppressing the wheat pathogenesis-related 2 gene, New Phytol 215(1) (2017) 338\u0026ndash;350.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM. Xu, G. Li, Y. Guo, Y. Gao, L. Zhu, Z. Liu, R. Tian, C. Gao, P. Han, N. Wang, F. Guo, J. Bao, C. Jia, H. Feng, L. Huang, A fungal microRNA-like RNA subverts host immunity and facilitates pathogen infection by silencing two host receptor‐like kinase genes, New Phytologist 233(6) (2022) 2503\u0026ndash;2519.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eD. Kumar, D. Singh, P. Kanodia, K.V. Prabhu, M. Kumar, K. Mukhopadhyay, Discovery of Novel Leaf Rust Responsive microRNAs in Wheat and Prediction of Their Target Genes, J Nucleic Acids 2014 (2014) 570176.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eN.A. Mueth, S.R. Ramachandran, S.H. Hulbert, Small RNAs from the wheat stripe rust fungus (\u003cem\u003ePuccinia striiformis\u003c/em\u003e f.sp. \u003cem\u003etritici\u003c/em\u003e), BMC Genomics 16(1) (2015) 718.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eR.S. Poethig, Small RNAs and developmental timing in plants, Curr Opin Genet Dev 19(4) (2009) 374\u0026ndash;8.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eO.P. Gupta, V. Permar, V. Koundal, U.D. Singh, S. Praveen, MicroRNA regulated defense responses in \u003cem\u003eTriticum aestivum\u003c/em\u003e L. during \u003cem\u003ePuccinia graminis\u003c/em\u003e f.sp. \u003cem\u003etritici\u003c/em\u003e infection, Mol Biol Rep 39(2) (2012) 817\u0026ndash;24.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eS. Dutta, M. Kumar, K. Mukhopadhyay, Identification and Validation of Leaf Rust Responsive Wheat isomiRs and their Target Genes in both Wheat and Puccinia triticina, (2019).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eS. Dutta, D. Kumar, S. Jha, K.V. Prabhu, M. Kumar, K. Mukhopadhyay, Identification and molecular characterization of a trans-acting small interfering RNA producing locus regulating leaf rust responsive gene expression in wheat (\u003cem\u003eTriticum aestivum\u003c/em\u003e L.), Planta 246(5) (2017) 939\u0026ndash;957.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003e\u0026Ouml;. Bilir, D. G\u0026ouml;l, Y. Hong, J.M. McDowell, M. T\u0026ouml;r, Small RNA-based plant protection against diseases, Front Plant Sci 13 (2022) 951097.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM.M. Nair, S.H.K. Kumar, S. Jyothsna, K.T. Sundaram, C. Manjunatha, M. Sivasamy, M. Alagu, Stem and leaf rust-induced miRNAome in bread wheat near-isogenic lines and their comparative analysis, Appl Microbiol Biotechnol 106(24) (2022) 8211\u0026ndash;8232.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eP. Gruner, A. Witzke, K. Flath, J. Eifler, B. Schmiedchen, M. Schmidt, A. Gordillo, D. Siekmann, F.J. Fromme, S. Koch, H.P. Piepho, T. Miedaner, Studying Stem Rust and Leaf Rust Resistances of Self-Fertile Rye Breeding Populations, Int J Mol Sci 23(22) (2022).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eK. Kumar, I. Jan, G. Saripalli, P.K. Sharma, R.R. Mir, H.S. Balyan, P.K. Gupta, An Update on Resistance Genes and Their Use in the Development of Leaf Rust Resistant Cultivars in Wheat, Front Genet 13 (2022) 816057.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eD. Kumar, S. Dutta, D. Singh, K.V. Prabhu, M. Kumar, K. Mukhopadhyay, Uncovering leaf rust responsive miRNAs in wheat (\u003cem\u003eTriticum aestivum\u003c/em\u003e L.) using high-throughput sequencing and prediction of their targets through degradome analysis, Planta 245(1) (2017) 161\u0026ndash;182.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eH. Feng, T. Wang, C. Feng, Q. Zhang, X. Zhang, L. Huang, X. Wang, Z. Kang, Identification of microRNAs and their corresponding targets involved in the susceptibility interaction of wheat response to \u003cem\u003ePuccinia striiformis\u003c/em\u003e f. sp. \u003cem\u003etritici\u003c/em\u003e, Physiol Plant 157(1) (2016) 95\u0026ndash;107.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM. Rakoczy-Trojanowska, P. Krajewski, J. Bocianowski, M. Schollenberger, W. Wakuliński, P. Milczarski, P. Masojć, M. Targońska-Karasek, Z. Banaszak, K. Banaszak, W. Brukwiński, W. Orczyk, A. Kilian, Identification of Single Nucleotide Polymorphisms Associated with Brown Rust Resistance, α-Amylase Activity and Pre-harvest Sprouting in Rye (Plant Mol Biol Report 35(3) (2017) 366\u0026ndash;378.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM. Święcicka, M. Dmochowska-Boguta, W. Orczyk, A. Grądzielewska, A. Stochmal, M. Kowalczyk, L. Bolibok, M. Rakoczy-Trojanowska, Changes in benzoxazinoid contents and the expression of the associated genes in rye (\u003cem\u003eSecale cereale\u003c/em\u003e L.) due to brown rust and the inoculation procedure, PLoS One 15(5) (2020) e0233807.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM. Martin, Cutadapt removes adapter sequences from high-throughput sequencing reads, EMBnet.journal 17(1) (2011) 10.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eS. Andrews, FASTQC. A quality control tool for high throughput sequence data, 2010.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eB. Langmead, C. Trapnell, M. Pop, S.L. Salzberg, Ultrafast and memory-efficient alignment of short DNA sequences to the human genome, Genome Biol 10(3) (2009) R25.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eS.D. Mackowiak, Identification of novel and known miRNAs in deep-sequencing data with miRDeep2, Curr Protoc Bioinformatics Chap. 12 (2011) Unit 12.10.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM.I. Love, W. Huber, S. Anders, Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2, Genome Biol 15(12) (2014) 550.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM.J. Axtell, ShortStack: comprehensive annotation and quantification of small RNA genes, RNA 19(6) (2013) 740\u0026ndash;51.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM.D. Robinson, D.J. McCarthy, G.K. Smyth, edgeR: a Bioconductor package for differential expression analysis of digital gene expression data, Bioinformatics 26(1) (2010) 139\u0026ndash;40.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eA. Kozomara, M. Birgaoanu, S. Griffiths-Jones, miRBase: from microRNA sequences to function, Nucleic Acids Res 47(D1) (2019) D155-D162.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZ. Guo, Z. Kuang, Y. Wang, Y. Zhao, Y. Tao, C. Cheng, J. Yang, X. Lu, C. Hao, T. Wang, X. Cao, J. Wei, L. Li, X. Yang, PmiREN: a comprehensive encyclopedia of plant miRNAs, Nucleic Acids Res 48(D1) (2020) D1114-D1121.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eX. Dai, Z. Zhuang, P.X. Zhao, psRNATarget: a plant small RNA target analysis server (2017 release), Nucleic Acids Research 46(W1) (2018) W49-W54.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eL. Moyo, S.V. Ramesh, M. Kappagantu, N. Mitter, V. Sathuvalli, H.R. Pappu, The effects of potato virus Y-derived virus small interfering RNAs of three biologically distinct strains on potato (\u003cem\u003eSolanum tuberosum\u003c/em\u003e) transcriptome, Virology Journal 14(1) (2017).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eU. Raudvere, L. Kolberg, I. Kuzmin, T. Arak, P. Adler, H. Peterson, J. Vilo, g:Profiler: a web server for functional enrichment analysis and conversions of gene lists (2019 update), Nucleic Acids Res 47(W1) (2019) W191-W198.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eL. Kolberg, U. Raudvere, I. Kuzmin, J. Vilo, H. Peterson, gprofiler2 -- an R package for gene list functional enrichment analysis and namespace conversion toolset g:Profiler, F1000Res 9 (2020).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eS. Feng, C. Pan, S. Ding, Q. Ma, C. Hu, P. Wang, K. Shi, The Glutamate Receptor Plays a Role in Defense against \u003cem\u003eBotrytis cinerea\u003c/em\u003e through Electrical Signaling in Tomato, Applied Sciences 11(23) (2021) 11217.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eG. Li, L. Wang, J. Yang, H. He, H. Jin, X. Li, T. Ren, Z. Ren, F. Li, X. Han, X. Zhao, L. Dong, Y. Li, Z. Song, Z. Yan, N. Zheng, C. Shi, Z. Wang, S. Yang, Z. Xiong, M. Zhang, G. Sun, X. Zheng, M. Gou, C. Ji, J. Du, H. Zheng, J. Doležel, X.W. Deng, N. Stein, Q. Yang, K. Zhang, D. Wang, A high-quality genome assembly highlights rye genomic characteristics and agronomically important genes, Nat Genet 53(4) (2021) 574\u0026ndash;584.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eX. Yang, L. Zhang, Y. Yang, M. Schmid, Y. Wang, miRNA Mediated Regulation and Interaction between Plants and Pathogens, Int J Mol Sci 22(6) (2021).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eA. Biswas, B. Sen, S. Bandyopadhyay, C. Mal, Co-regulatory functions of miRNA and lncRNA in adapting biotic and abiotic stress in economically important dicot plants, Plant Gene 26 (2021).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eN. Jain, S. Rani, C. Sharma, N. Sinha, A. Singh, J.B. Sharma, P. Prasad, G. Saripalli, P.K. Sharma, H.S. Balyan, P.K. Gupta, K.V. Prabhu, Large-scale stage-specific regulation of gene expression during host-pathogen interactions in CSP44 bread wheat carrying APR gene Lr48, Funct Plant Biol 47(3) (2020) 203\u0026ndash;225.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eS. Das, D. Singh, H.S. Meena, S.K. Jha, J. Kumari, V. Chinnusamy, L. Sathee, Long term nitrogen deficiency alters expression of miRNAs and alters nitrogen metabolism and root architecture in Indian dwarf wheat (\u003cem\u003eTriticum sphaerococcum\u003c/em\u003e Perc.) genotypes, Sci Rep 13(1) (2023) 5002.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eH. Zhang, X. Chen, C. Wang, Z. Xu, Y. Wang, X. Liu, Z. Kang, W. Ji, Long non-coding genes implicated in response to stripe rust pathogen stress in wheat (\u003cem\u003eTriticum aestivum\u003c/em\u003e L.), Mol Biol Rep 40(11) (2013) 6245\u0026ndash;53.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eB. Liu, X. Xue, S. Cui, X. Zhang, Q. Han, L. Zhu, X. Liang, X. Wang, L. Huang, X. Chen, Z. Kang, Cloning and characterization of a wheat β-1,3-glucanase gene induced by the stripe rust pathogen \u003cem\u003ePuccinia striiformis\u003c/em\u003e f. sp. \u003cem\u003etritici\u003c/em\u003e, Molecular Biology Reports 37(2) (2010) 1045\u0026ndash;1052.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eL. Gao, S. Wang, Y. Zhang, X. Li, H. Wang, D. Liu, Identification and characterization of a β-1, 3-glucanase gene, TcLr19Glu, involved in wheat resistance against \u003cem\u003ePuccinia triticina\u003c/em\u003e, J. Plant Biochem. Biotechnol. 25 (2016) 319\u0026ndash;326.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eA. Amo, J.M. Soriano, Unravelling consensus genomic regions conferring leaf rust resistance in wheat via meta-QTL analysis, Plant Genome 15(1) (2022) e20185.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eS. Liu, J. Lan, B. Zhou, Y. Qin, Y. Zhou, X. Xiao, J. Yang, J. Gou, J. Qi, Y. Huang, C. Tang, HbNIN2, a cytosolic alkaline/neutral-invertase, is responsible for sucrose catabolism in rubber-producing laticifers of \u003cem\u003eHevea brasiliensis\u003c/em\u003e (para rubber tree), New Phytol 206(2) (2015) 709\u0026ndash;25.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eT.E. Coram, X. Huang, G. Zhan, M.L. Settles, X. Chen, Meta-analysis of transcripts associated with race-specific resistance to stripe rust in wheat demonstrates common induction of blue copper-binding protein, heat-stress transcription factor, pathogen-induced WIR1A protein, and ent-kaurene synthase transcripts, Funct Integr Genomics 10(3) (2010) 383\u0026ndash;92.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eP. Prasad, S. Savadi, S.C. Bhardwaj, P.L. Kashyap, O.P. Gangwar, H. Khan, S. Kumar, R. Kumar, V. Patil, Stage-specific reprogramming of defense responsive genes during Lr24-mediated leaf rust resistance in wheat, Journal of Plant Pathology 111(2) (2019) 283\u0026ndash;293.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM.S. Mafa, N. Lebusa, T.F. Gumani, G. Kemp, B. Visser, W.H.P. Boshoff, H.D. Castelyn, Accumulation of complex oligosaccharides and CAZymes activity under acid conditions constitute the Thatcher\u0026thinsp;+\u0026thinsp;Lr9 defence responses to \u003cem\u003ePuccinia triticina\u003c/em\u003e, \u003cem\u003eBiologia\u003c/em\u003e 78 (2023).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eE.C. Henningsen, V. Omidvar, R. Della Coletta, J.M. Michno, E. Gilbert, F. Li, M.E. Miller, C.L. Myers, S.P. Gordon, J.P. Vogel, B.J. Steffenson, S.F. Kianian, C.D. Hirsch, M. Figueroa, Identification of Candidate Susceptibility Genes to, Front Plant Sci 12 (2021) 657796.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZ. Zhao, T. Chen, J. Yue, N. Pu, J. Liu, L. Luo, M. Huang, T. Guo, W. Xiao, regulates heading date in rice, Mol Breed 43(8) (2023) 62.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eY. Loarce, E. Navas, C. Paniagua, A. Fominaya, J.L. Manj\u0026oacute;n, E. Ferrer, Identification of Genes in a Partially Resistant Genotype of Avena sativa Expressed in Response to \u003cem\u003ePuccinia coronata\u003c/em\u003e Infection, Front Plant Sci 7 (2016) 731.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eS. Li, C. Castillo-Gonz\u0026aacute;lez, B. Yu, X. Zhang, The functions of plant small RNAs in development and in stress responses, Plant J 90(4) (2017) 654\u0026ndash;670.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eS. Bhogireddy, S.K. Mangrauthia, R. Kumar, A.K. Pandey, S. Singh, A. Jain, H. Budak, R.K. Varshney, H. Kudapa, Regulatory non-coding RNAs: a new frontier in regulation of plant biology, Funct Integr Genomics 21(3\u0026ndash;4) (2021) 313\u0026ndash;330.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eS.G. Krattinger, E.S. Lagudah, W. Spielmeyer, R.P. Singh, J. Huerta-Espino, H. McFadden, E. Bossolini, L.L. Selter, B. Keller, A putative ABC transporter confers durable resistance to multiple fungal pathogens in wheat, Science 323(5919) (2009) 1360\u0026ndash;3.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eY. Han, Q. Zhu, Z. Zhang, K. Meng, Y. Hou, Q. Ban, J. Suo, J. Rao, Analysis of xyloglucan endotransglycosylase/hydrolase (XTH) genes and diverse roles of isoenzymes during persimmon fruit development and postharvest softening, PLoS One 10(4) (2015) e0123668.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eE. Miedes, E.P. Lorences, The implication of xyloglucan endotransglucosylase/hydrolase (XTHs) in tomato fruit infection by \u003cem\u003ePenicillium expansum\u003c/em\u003e Link. A, J Agric Food Chem 55(22) (2007) 9021\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eH.H. Flor, Current Status of the Gene-For-Gene Concept, Annual Review of Phytopathology 9(1) (1971) 275\u0026ndash;296.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eY. Duan, S. Duan, M.R. Armstrong, J. Xu, J. Zheng, J. Hu, X. Chen, I. Hein, G. Li, L. Jin, Comparative Transcriptome Profiling Reveals Compatible and Incompatible Patterns of Potato Toward, G3 (Bethesda) 10(2) (2020) 623\u0026ndash;634.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eB. Chandrasekar, A. Wanke, S. Wawra, P. Saake, L. Mahdi, N. Charura, M. Neidert, G. Poschmann, M. Malisic, M. Thiele, K. St\u0026uuml;hler, M. Dama, M. Pauly, A. Zuccaro, Fungi hijack a ubiquitous plant apoplastic endoglucanase to release a ROS scavenging β-glucan decasaccharide to subvert immune responses, Plant Cell 34(7) (2022) 2765\u0026ndash;2784.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM. M\u0026uuml;ller, S. Munn\u0026eacute;-Bosch, Ethylene Response Factors: A Key Regulatory Hub in Hormone and Stress Signaling, Plant Physiol 169(1) (2015) 32\u0026ndash;41.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eF. Dunker, A. Trutzenberg, J.S. Rothenpieler, S. Kuhn, R. Pr\u0026ouml;ls, T. Schreiber, A. Tissier, A. Kemen, E. Kemen, R. H\u0026uuml;ckelhoven, A. Weiberg, Oomycete small RNAs bind to the plant RNA-induced silencing complex for virulence, Elife 9 (2020).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eH. Dubey, K. Kiran, R. Jaswal, P. Jain, A.M. Kayastha, S.C. Bhardwaj, T.K. Mondal, T.R. Sharma, Discovery and profiling of small RNAs from \u003cem\u003ePuccinia triticina\u003c/em\u003e by deep sequencing and identification of their potential targets in wheat.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJ. Mapuranga, J. Chang, L. Zhang, N. Zhang, W. Yang, Fungal Secondary Metabolites and Small RNAs Enhance Pathogenicity during Plant-Fungal Pathogen Interactions, Journal of Fungi 9(1) (2023) 4.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eX. Zhang, Y. Liu, A. Ayaz, H. Zhao, S. L\u0026uuml;, The Plant Fatty Acyl Reductases, Int J Mol Sci 23(24) (2022).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJ. Cheng, N. Song, J. Wu, A patatin-like protein synergistically regulated by jasmonate and ethylene signaling pathways plays a negative role in, Plant Divers 41(1) (2019) 7\u0026ndash;12.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eR. Xue, M. Feng, J. Chen, W. Ge, M.W. Blair, A methyl esterase 1 (PvMES1) promotes the salicylic acid pathway and enhances Fusarium wilt resistance in common beans, Theor Appl Genet 134(8) (2021) 2379\u0026ndash;2398.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eM. Hasegawa, I. Mitsuhara, S. Seo, T. Imai, J. Koga, K. Okada, H. Yamane, Y. Ohashi, Phytoalexin Accumulation in the Interaction Between Rice and the Blast Fungus, Molecular Plant-Microbe Interactions 23(8) (2010) 1000\u0026ndash;1011.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJ.S. Ramirez-Prado, S.J.M. Piquerez, A. Bendahmane, H. Hirt, C. Raynaud, M. Benhamed, Modify the Histone to Win the Battle: Chromatin Dynamics in Plant-Pathogen Interactions, Front Plant Sci 9 (2018) 355.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eA. Sharma, V. Kumar, B. Shahzad, M. Tanveer, G.P.S. Sidhu, N. Handa, S.K. Kohli, P. Yadav, A.S. Bali, R.D. Parihar, O.I. Dar, K. Singh, S. Jasrotia, P. Bakshi, M. Ramakrishnan, S. Kumar, R. Bhardwaj, A.K. Thukral, Worldwide pesticide usage and its impacts on ecosystem, SN Appl. Sci. 1 (2019) 1446.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eH. Sang, J.-I. Kim, Advanced strategies to control plant pathogenic fungi by host-induced gene silencing (HIGS) and spray-induced gene silencing (SIGS), Plant Biotechnology Reports 14 (2020) 1\u0026ndash;8.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eB.T.L. Hoang, S.J. Fletcher, C.A. Brosnan, A.B. Ghodke, N. Manzie, N. Mitter, RNAi as a Foliar Spray: Efficiency and Challenges to Field Applications, Int J Mol Sci 23(12) (2022).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eN. Mitter, E.A. Worrall, K.E. Robinson, P. Li, R.G. Jain, C. Taochy, S.J. Fletcher, B.J. Carroll, G.Q. Lu, Z.P. Xu, Clay nanosheets for topical delivery of RNAi for sustained protection against plant viruses, Nat Plants 3 (2017) 16207.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[{"identity":"a16c6e9f-b7af-49e9-9e48-6423017874ce","identifier":"10.13039/501100004281","name":"Narodowe Centrum Nauki","awardNumber":"2018/31/B/NZ9/00439","order_by":0}],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"National Science Centre","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"sRNA, leaf rust, isomiR, milRNA, ck-RNAi","lastPublishedDoi":"10.21203/rs.3.rs-3875675/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-3875675/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eLeaf rust (LR) caused by \u003cem\u003ePuccinia recondita\u003c/em\u003e f. sp. \u003cem\u003esecalis\u003c/em\u003e (\u003cem\u003ePrs\u003c/em\u003e) is one of the most damaging diseases of rye. However, the genetic basis of the rye response to LR remains relatively unknown. Specifically, the involvement of small RNAs (sRNAs) in the rye\u0026ndash;\u003cem\u003ePrs\u003c/em\u003e interaction has not been characterized. In this study, the changes in various sRNAs in response to LR were revealed in three rye lines (D33, D39, and L318) that vary regarding their immune responses to LR and two \u003cem\u003ePrs\u003c/em\u003e isolates that cause compatible (CP) or incompatible (ICP) interactions with the host. The sRNAs were analyzed using miRDeep2 and ShortStack algorithms, with true miRNAs detected on the basis of strict miRNA biosynthesis-related parameters. The differential expression of all sRNAs and miRNAs was analyzed and potential targets (i.e., mRNA) were detected. Additionally, the targets in the CP and ICP host\u0026ndash;pathogen interactions were compared. Moreover, different isoforms of miRNAs originating from the same miRNA gene (isomiRs) were manually identified and the diversity in their potential targets were examined. The isomiRs can broaden the range of targets related to plant defense responses to pathogens. For example, UGUGUUCUCAGGUCGCCCCCG targets the transcripts of 11 endoglucanase genes and two superoxide dismutase genes, which encode proteins with plant defense-related functions. A shorter sequence (UCAGGUCGCCCCCGCUGGAG) was revealed to target 16 mRNAs encoding transcription factors, including those from the MADS box family. Finally, miRNA-like RNAs (milRNAs) from \u003cem\u003ePrs\u003c/em\u003e and their predicted targets in rye were identified. To examine the effects of the experimental design, the differences in the number of milRNAs between CP and ICP interactions as well as the milRNA expression dynamics at two time-points were analyzed. Compared with the ICP interactions, the CP interactions resulted in more milRNAs, which affected many more genes, including some related to biotic stress responses.\u003c/p\u003e","manuscriptTitle":"Role of sRNA molecules in the rye–leaf rust interaction","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-01-19 07:13:56","doi":"10.21203/rs.3.rs-3875675/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"e9e186e2-b311-4d92-b173-b0f90e8c7c0d","owner":[],"postedDate":"January 19th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":28221795,"name":"Molecular Genetics"}],"tags":[],"updatedAt":"2024-01-19T07:13:56+00:00","versionOfRecord":[],"versionCreatedAt":"2024-01-19 07:13:56","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-3875675","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-3875675","identity":"rs-3875675","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}