Genome-wide identification and expression analysis of the monosaccharide transporter (MST) gene family in wheat (Triticum aestivum L.)

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Jun Zhang, Bo Zhang, Shuang Zhou, Wenzhong Tian, Rong Zhang, Yiren Chen, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7561375/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 21 Jan, 2026 Read the published version in Genetica → Version 1 posted 4 You are reading this latest preprint version Abstract Carbohydrates function as both energy sources and signaling molecules in various critical physiological processes. Monosaccharide transporters (MSTs) are a class of membrane-bound carrier proteins in crops that mediate the transmembrane transport of monosaccharides, thereby playing a central role in crop growth and development, resource allocation, and responses to environmental stimuli. In this study, a total of 200 MST family genes were identified in wheat and categorized into seven subfamilies. Twenty conserved motifs were detected within the TaMST family, with each subfamily exhibiting similar conserved motif patterns. The TaMST gene family was evenly distributed across the three wheat subgenomes, with both segmental and tandem duplications contributing to gene family expansion. The TaMST gene family was found to contain numerous cis-regulatory elements associated with growth and development, hormone signaling, and abiotic stress responses. Expression analysis revealed that most TaMSTs were expressed at low levels in wheat grains, whereas 69, 66, 67, and 64 genes exhibited high expression levels in leaves, buds, roots, and spikes, respectively. Following exogenous sugar treatments, the expression of all TaMSTs in roots was down-regulated, while 4, 2, and 3 genes showed up-regulated expression in leaves after treatment with fructose, glucose, and sucrose, respectively. Subcellular localization displayed TaERD3, TaPMT29 and TaSTP18 were all located on the cell membrane. These findings suggest that MSTs play essential roles not only in wheat organ development but also in the perception and response to sugar signaling. This study provides valuable insights for future investigations into the functional divergence of the MST gene family. Wheat Monosaccharide transporter Functional divergence Expression analysis Subcellular localization Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 Introduction Carbohydrate serves as a fundamental substance for the growth, development, and energy metabolism of higher plants and constitutes a key component of carbon skeletons(Ma et al., 2024 ). It regulates various physiological processes such as flowering, seed germination, root architecture, senescence, and stress responses through mechanisms including energy storage, osmotic adjustment, and signal transduction(Ruan, 2014 ). Carbohydrate is synthesized and exported from “source” organs and accumulated or converted in “sink” organs. The “flow” between these organs primarily consists of fructose, glucose, and sucrose, which are transported over short or long distances to connect source and sink tissues. This intricate process requires the coordinated action of multiple sugar transporters for precise regulation(Eom et al., 2015 ). In plants, major sugar transporter families include Sugar Will Eventually Be Exported Transporters (SWEETs), Sucrose Transporters (SUTs), and Monosaccharide Transporters (MSTs)(Deng et al., 2019 ; Eom et al., 2015 ). MST proteins belong to the family of intrinsic membrane proteins and are primarily responsible for the transmembrane transport of various monosaccharides[4]. Wheat is classified as a “sugary leaf” plant, with sugar content accounting for up to 95% of the dry weight of its leaves. Even minor fluctuations in sugar levels can trigger significant changes in gene expression (Ma et al., 2024 ). Therefore, identifying the MSTs gene family within the wheat genome and analyzing its expression patterns across different tissues, organs, and under exogenous sugar treatments will provide a foundation for further functional studies of MSTs in wheat and offer valuable insights for high-yield wheat breeding programs. Genome-wide identification of the MST gene family has been conducted in several species, including Arabidopsis (Büttner, 2007 ; Büttner, 2010 ; Wormit et al., 2007 ), grape(Afoufa-Bastien et al., 2010 ), tobacco(Okubo-Kurihara et al., 2011 ), tomato(Hackel et al., 2006 ; Mccurdy et al., 2010 ), alfalfa(Doidy et al., 2012b ), and rose(Henry et al., 2011 ). Based on sequence characteristics and substrate specificity, the MST family can be categorized into seven subfamilies: hexose transporters (Sugar Transport Protein, STP), polyol/monosaccharide transporters (Polyol/Monosaccharide Transporter, PMT), early-responsive to dehydration six-like (ERD6), vacuolar membrane monosaccharide transporters (Tonoplast Membrane Transporter, TMT), inositol transporters (Inositol Transporter, INT), plastidic glucose transporters (Plastidic Glucose Transporter, pGlcT), and vacuolar glucose transporters (Vacuolar Glucose Transporter, VGT)(Johnson et al., 2006 ; Johnson and Thomas, 2007 ). In Arabidopsis, identified STP proteins function as H + /hexose co-transporters localized on the plasma membrane. Most STPs exhibit broad substrate transport capabilities(Büttner, 2010 ; Zheng et al., 2014 ). For example, AtSTP1, AtSTP2, AtSTP3, AtSTP4, AtSTP6, and AtSTP11 can transport glucose, xylose, mannose, and galactose with varying affinities but do not transport fructose (Cho et al., 2010 ). Conversely, AtSTP6, AtSTP13, and OsMST4 are capable of transporting fructose but not pentoses such as xylose and ribose(Afoufa-Bastien et al., 2010 ; Büttner, 2010 ; Wang et al., 2007 ). However, certain STP proteins demonstrate substrate specificity in transport. For instance, AtSTP9 exclusively transports glucose, while AtSTP14 specifically transports galactose(Poschet et al., 2010 ; Schneider et al., 2008 ). Additionally, PLT-like proteins have been shown to transport both polyols and monosaccharides(Klepek et al., 2005 ; Klepek et al., 2010 ). Among the XTPH (also known as VGT) proteins, AtVGT1 facilitates glucose transport but not xylose. Both AtVGT1 and AtVGT2 are H + /glucose antiporters located on the vacuolar membrane and play roles in the transport and storage of monosaccharides within the vacuole (Büttner, 2007 ; Wormit et al., 2007 ). Similarly, AZT (also known as TMT) proteins are localized on the vacuolar membrane. Studies have demonstrated that in the attmt1/attmt2 double mutant of Arabidopsis, the vacuole's capacity to take up sucrose is significantly reduced, indicating that TMT proteins participate in the transmembrane transport of sucrose across the vacuolar membrane (Schulz et al., 2011 ). Meanwhile, pGlcT proteins are involved in the transport of glucose (Cho et al., 2011 ). Within the MST family in Arabidopsis, ERD6 (also referred to as SFPs) represents the largest subfamily. The first identified ERD6 protein, AtERD6, is induced by drought and low temperature(Quirino et al., 2001 ). Notably, AtSFP1 and AtSFP2 are stress-induced auxiliary diffusion transporters exhibiting distinct spatiotemporal expression patterns (Yamada et al., 2009 ). Beyond model species like Arabidopsis, several MST genes have also been characterized in rice. For example, Toyofuku et al. cloned and analyzed three MST genes, OsMST1 , OsMST2 , and OsMST3 , and found that OsMST3 mediates the transport of specific monosaccharides via energy-dependent H + co-transport(Kyoko et al., 2000 ). Functional analysis through heterologous expression confirmed that OsMST5 plays a role in regulating pollen development in rice(Ngampanya et al., 2003 ). Moreover, OsMST4 , which exhibits constitutive expression, can transport fructose, galactose, mannose, and glucose(Poschet et al., 2010 ). Wang et al. reported that OsMST6 functions as a broad-spectrum monosaccharide transporter whose expression is induced by salt stress and sugar treatment(Wang et al., 2008 ). Furthermore, OsTMTs are involved in vacuolar sugar transport(Cho et al., 2010 ). Additionally, OsGMST1 is up-regulated under salt stress conditions, and knocking down this gene significantly reduces the salt stress resistance of rice(Cao et al., 2011 ). In summary, the MST family genes exert an essential role in plant growth and development as well as the response to biological and abiotic stresses. This study was motivated by the importance of wheat as a globally significant staple crop and the increasing use of wheat genomic data for gene family identification. Previous studies have reported on gene families such as FDL(Kan et al., 2025 ), NHX(Sharma et al., 2023 ), GH13(Yin et al., 2024 ), and SNARE(Wang et al., 2021 ). However, no comprehensive identification of the MST gene family in wheat has been reported to date, which has hindered functional studies of MST genes in this species. To address this gap, the present study conducted a systematic bioinformatics analysis of the wheat MSTs gene family, focusing on physicochemical properties, gene structure, intraspecific collinearity, promoter cis-regulatory elements, expression profiles, subcellular localization, and protein interaction networks. Additionally, the expression patterns of MST genes across different wheat tissues were analyzed, and the effects of exogenous sugar signaling on MST gene expression were investigated. These findings provide a foundational reference for future functional studies of MST genes in wheat. Materials and Methods Genome-wide identification of the TaMST family in wheat The genome data of wheat was obtained from the EnsemblPlants database (http://plants.ensembl.org/index.html). The MST domain (PF00083) was retrieved from the Pfam database. HMMER 3.0 was employed to perform conserved domain searches for identifying members of the TaMSTs gene family. Additionally, the protein sequence of AtMSTs from Arabidopsis thaliana was used as a query for BLASTP analysis (E-value < 1E -5 ) to identify homologous sequences. Candidate TaMST genes were selected based on combined results from BLAST and HMMER analyses. Basic physicochemical and phylogenetic analysis of the TaMSTs family in wheat The amino acid length, isoelectric point, and molecular weight of TaMST proteins were analyzed using the ExPASy ProtParam tool (https://web.expasy.org/protparam/). Subcellular localization prediction was conducted using ProtComp 9.0 software (https://linux1.softberry.com/berry.phtml). Transmembrane helices were predicted via the DTU Health Tech server (https://services.healthtech.dtu.dk/services/TMHMM-2.0/). The neighbor joining (NJ) phylogenetic tree was constructed by using mega7 tool with adjacency method (bootstrap value was set to 1000), and it was optimized by online software ChiPlot (https://www.chiplot.online/). Gene structure and conserved motif analysis of the TaMST family in wheat Gene structures were determined based on the gff annotation file. Conserved motifs among TaMST family members were identified using the MEME Suite (http://meme-suite.org/tools/meme), with the maximum number of motifs set to 20 and other parameters at default settings. Visualization of gene structures and motifs was performed using TBtools. Chromosome localization and collinearity analysis of the TaMSTs family in wheat Chromosomal positions of TaMST genes were extracted from the IWGSC wheat genome annotation database (http://wheat-urgi.versailles.inra.fr/). Chromosomal localization was visualized by TBtools. Collinearity analysis was carried out through MCScanX and visualized via Advanced Circos. Ka/Ks analysis of the TaMSTs family in wheat Nucleotide sequences of TaMST genes were downloaded from the EnsemblPlants database. MCScanX was used to identify collinear gene pairs. The Ka/Ks ratio was calculated by the TBtools implementation of the Ka/Ks Calculator program to assess selection pressures acting on duplicated gene pairs. Promoter cis-regulatory element analysis of the TaMSTs family in wheat The 2000-bp upstream promoter regions of TaMST genes were extracted via TBtools. Potential cis-regulatory elements within these promoter sequences were predicted by the PlantCARE database (http://bioinformatics.psb.ugent.be/webtools/plantcare/html/), and the results were visualized according to TBtools. Expression patterns of the wheat TaMSTs family across different tissues Expression levels of TaMST genes in root, grain, spike, leaf, and bud tissues of the Chinese Spring wheat variety were obtained from the ExpVIP wheat expression database (wheat-expression.com). Data were analyzed and visualized as a heatmap using TBtools. Expression analysis of the wheat TaMSTs family under exogenous sugar treatments Seeds of the YM2 wheat cultivar with uniform size were surface-sterilized, rinsed thoroughly with distilled water, and soaked in sterile water for 12 hours. After germination, seedlings were transferred onto floating nets for hydroponic growth. At the two-leaf and one-heart stage, plants were subjected to four treatments: CK (distilled water), T1 (2% sucrose), T2 (2% fructose), and T3 (2% glucose). Each treatment had three biological replicates. Leaf samples were collected 24 hours after treatment and immediately frozen in liquid nitrogen. Total RNA was extracted using TRIZOL reagent (TaKaRa, 9108Q), reverse-transcribed into cDNA (TaKaRa, RR047Q), and analyzed by RT-qPCR (Vazyme, 221). Relative gene expression levels were calculated using the 2 ⁻ΔΔCt method. The primers used in this study were listed in Table S1. Gene cloning, vector construction, and subcellular localization Full-length coding sequences of TaERD3 , TaPMT2 and TaSTP18 were amplified from YM 2 cDNA using specific primers (Table S1). PCR amplification was performed using 2× Phanta Flash Master Mix (Dye Plus) with high-fidelity enzyme (P520, Vazyme). The reaction mixture consisted of 1 μL template, 1 μL each of forward and reverse primers, 10 μL master mix, and 7 μL ddH₂O. Amplification was carried out according to the manufacturer’s recommended thermal cycling conditions. After electrophoresis, the gel was recovered through the gel recovery kit (CW2302M, Cwbio). The sequences that were sequenced correctly were truncated at the terminators and added with restriction sites, and then subcloned into the pCAMBIA1300-GFP vector. The recombinant plasmid was transformed into Agrobacterium GV3101. The leaves of Nicotiana benthamiana were injected on the back side and cultured for 48 hours. The subcellular localization signals were observed via a laser confocal microscope (LSM710, Zeiss). Results and discussion Genome-wide identification of the TaMSTs family in wheat Based on the protein sequences of 53 AtMSTs from Arabidopsis , a total of 260 TaMST candidate genes were identified in wheat through BLASTP and HMM searches. After filtering out genes with incomplete domains and redundant sequences using NCBI-CD, a final set of 200 TaMST genes was obtained. These genes were systematically renamed according to their chromosomal positions and corresponding protein sequences, for example, from TaSTP1 to TaSTP74 . The physicochemical properties and subcellular localization data are summarized in Supplementary Table S2. Analysis of the primary structure of all TaMST proteins revealed that the amino acid length ranged from 393 to 591 residues, molecular weights varied between 41.91 and 63.04 kDa, theoretical isoelectric points (pI) ranged from 4.90 to 10.05, with 40 acidic amino acids, 1 neutral amino acid, and the rest being alkaline amino acids. The protein instability index ranged from 28.99 to 54.12, with those below 40 being stable proteins. There were a total of 131 proteins classified as stable proteins. The aliphatic index ranged from 94.67 to 119.27, while the average hydrophobicity index ranged from 0.368 to 0.803. Among these, 36 proteins were predicted to be amphipathic, and the rest were hydrophobic. Subcellular localization predictions indicated that two TaMST proteins were localized to thylakoid membranes, eight to the endoplasmic reticulum, 38 to vacuolar membranes, and the remaining to the plasma membrane. Transmembrane helix prediction analysis showed that TaMST proteins contain approximately 8 to 12 transmembrane helices. Monosaccharide transporters (MSTs) play essential roles in model plant species such as Arabidopsis and rice(Büttner, 2007; Cho et al., 2010). However, in other crops, including wheat, the identification and expression profiling of MST gene families remain limited. In 2020, researchers from the Crop Development Centre at the University of Saskatchewan, in collaboration with several leading international wheat genome research teams, published the genome sequences of 16 representative wheat varieties, representing the most comprehensive wheat genome resource available to date. This milestone has significantly advanced wheat genomics and provided a solid foundation for the identification and functional analysis of the wheat MST gene family(Walkowiak et al., 2020). In this study, a comprehensive analysis of the gene structure, phylogeny, and conserved protein motifs of wheat MST family members was conducted for the first time. This discrepancy may be attributed to the hexaploid nature of wheat. Most MST proteins were basic (pI>7), likely due to their high content of basic amino acids, suggesting that these transporters predominantly function in alkaline environments. Using BLAST searches and HMMER analysis, a total of 200 TaMST genes were identified in the wheat genome, which is substantially higher than the 53 in Arabidopsis(Büttner, 2007), 65 in rice(Johnson and Thomas, 2007), 59 in grape(Afoufa-Bastien et al., 2010), and 58 in Medicago truncatula (Doidy et al., 2012a). Phylogenetic analysis, gene structure, and conserved motif analysis of the TaMSTs family in wheat Phylogenetic analysis of the TaMSTs amino acid sequences was conducted using MEGAX software (Fig. 1). In terms of the resulting phylogenetic tree, the TaMSTs family can be classified into seven distinct subfamilies: TaERD, TaINT, TapGlcT, TaPMT, TaSTP, TaTMT, and TaVGT. The TaSTPs subfamily contained the largest number of members (74), whereas the TaVGTs subfamily had the fewest (6). Other subfamilies included TaERDs (34), TaINTs (9), TapGlcTs (9), TaPMTs (48), and TaTMTs (20). The exon-intron structures of TaMST s were determined based on gene annotation data (Fig. 2). The analysis results indicated that among the 200 TaMST family members, 196 contained both exons and introns, with a notable variation observed in the number of exons and introns. The number of exons ranged from 1 to 18, while the number of introns varied from 1 to 17. Furthermore, four TaMST members were found to consist solely of a single exon without any introns. Members within the same subfamily generally exhibited similar exon-intron structures. Notably, most members of the TaERD , TapGlcT , and TaVGT subfamilies contained approximately 16 exons. In contrast, TaPMT and 90% of TaTMT members involved only 2~3 introns, and the remaining subfamilies had fewer than six exons per gene. Additionally, 16 genes lacked upstream or downstream untranslated regions (UTRs), which may affect their regulatory functions. Conserved motifs among the 200 TaMST family members were analyzed using the online tool MEME (Fig. 2, Fig. 3), identifying a total of 20 distinct motifs (Motif 1 to Motif 20). Within each subfamily, the composition and sequence of conserved motifs were relatively consistent. The TaSTP subfamily exhibited the highest number of motifs (approximately 16), while the TaTMT subfamily had the fewest (an average of 12). Motifs 2, 5, 8, and 13 were present in all genes, whereas Motif 16 was exclusive to the TaSTP subfamily and Motif 17 was unique to the TaTMT subfamily. These differences in motif distribution suggest that the TaMSTs gene family has undergone functional divergence during evolution. Phylogenetic analysis classified the TaMST family into seven distinct subfamilies: TaERD , TaINT , TapGlcT , TaPMT , TaSTP , TaTMT , and TaVGT . Among these, the TaSTP subfamily contains the highest number of members (74), whereas the TaVGT subfamily contains the fewest (6). Members of the TaERD , TapGlcT , and TaVGT subfamilies possess a relatively high number of exons (approximately 16), whereas the remaining subfamilies contain fewer than six exons. These structural variations may result from evolutionary events such as exon and intron insertions or deletions, exonization, and pseudo-exonization, reflecting the structural diversity of MST genes(Xu et al., 2012). Chromosome Distribution and Co-linearity Analysis of the TaMSTs Family in Wheat According to the chromosome location map of the TaMST genes (Fig. 4), a total of 200 TaMST genes are distributed across 22 wheat chromosomes, with a relatively even distribution among the three sub-genomes. Specifically, the A, B, and D genomes contain 67, 67, and 63 genes, respectively. Chromosomes 2 and 5 exhibit relatively higher gene densities, particularly chromosome 2, which harbors 68 TaMST genes. In contrast, chromosomes 3, 6, and 7 display lower gene counts, with 12, 11, and 14 genes, respectively. Co-linearity analysis of the TaMST gene family was performed using MCScanX (Fig. 5), revealing that 55 TaMST genes formed 23 tandem repeat gene pairs. Among these, the TaERD , TaPMT , TaSTP , and TaTMT sub-families contained 4, 8, 7, and 4 tandem repeat gene pairs, respectively, and no tandem repeats were identified in other sub-families. A total of 81,798 co-linear regions were detected in the wheat genome, with 130 TaMST genes located within these regions. These genes were primarily generated through segmental duplication events. The TaSTP sub-family exhibited the highest number of segmental duplication genes, accounting for 56 genes, whereas the TaVGT sub-family had the fewest, with only 4 genes. The significant expansion of the TaSTP sub-family suggests that both tandem and segmental duplication mechanisms have played crucial roles in the evolutionary expansion of the TaMST gene family. Furthermore, Ka/Ks ratio analysis revealed that all values were less than 1, indicating that these genes have undergone purifying selection and have been evolutionarily conserved (Supplementary Table S3). Phylogenetic and evolutionary analyses revealed that MST family members across various species, including Arabidopsis, rice, and grape, can be classified into seven subfamilies. The subfamily classification of wheat MST genes aligns with that of other species, suggesting that wheat, despite being a hexaploid organism, has not developed lineage-specific MST genes. Within the wheat MST gene family, all seven subfamilies contain four conserved motifs, whereas in maize, only three conserved motifs are shared among MST members(Zhu et al., 2024). The number of conserved motifs varies across subfamilies, indicating both structural conservation and functional divergence among MST subfamilies(Ding et al., 2023). Gene structure analysis further revealed that MST members within the same subfamily exhibit similar exon numbers and conserved motifs, implying potential functional similarity among subfamily members. Whole genome duplication events significantly contribute to the expansion and diversification of plant gene families, influencing both gene structure and function. Polyploidization, a key component of whole genome duplication in wheat, plays a major role in the expansion of gene families(Pozo and Ramirez-Parra, 2015). In Arabidopsis, the genes AtERD6-like4 , AtINT2 , AtpGlcT4 , AtPMT4 , AtSTP1 , AtTMT2 , and AtVGT3 represent the ancestral members of the seven MST subfamilies. The wheat MST subfamilies identified in this study show direct orthologous relationships with these ancestral Arabidopsis genes, indicating that substantial gene duplication events have occurred within the wheat MST gene family. Research by Johnson et al. further supports this, demonstrating that following the divergence of monocot and dicot plants, distinct mechanisms of whole genome duplication occurred in each lineage, collectively contributing to the evolutionary divergence of the wheat MST subfamilies(Johnson and Thomas, 2007). The collinearity analysis results revealed that 130 wheat MST genes were located within collinear regions, and these genes were primarily generated through segmental duplication. Among them, the TaSTP subfamily exhibited the highest number of segmentally duplicated genes, encompassing 56 related members. Furthermore, 55 wheat MST genes constituted 23 pairs of tandem repeats, indicating that both tandem and segmental duplication mechanisms have played vital roles in the expansion of the MST gene family. In addition, all Ka/Ks ratios of these duplicated gene pairs were less than 1, suggesting that these MST genes have undergone purifying selection and have been evolutionarily conserved. Analysis of cis -acting elements in the TaMST family of wheat To further elucidate the functional characteristics of the TaMST gene family, cis -acting elements within the 2 kb upstream promoter regions of these genes were analyzed. Eighteen representative elements were selected for visualization (Fig. 6). The results indicated the presence of three major categories of cis -acting elements: those associated with growth and development, hormone responsiveness, and abiotic stress responses. Within the growth and development-related category, several photosynthesis-related elements, such as G-Box, C-Box, MRE, and Sp1, were commonly identified across all members. Additionally, root-specific regulatory elements, meristem tissue-specific elements, and palisade mesophyll cell differentiation-related elements were also detected. Regarding abiotic stress response elements, drought-inducible elements, defense/stress-responsive elements, cold stress-responsive elements, and hypoxia-inducible elements were identified. Notably, 77 and 66 genes contained drought- and cold-responsive elements, respectively. With respect to hormone-responsive elements, 60.5%, 27.5%, and 90.0% of the genes contained auxin-, gibberellin-, and abscisic acid-responsive elements, respectively. The gene TaERD25 exhibited the lowest number of regulatory elements, with only seven, and lacked any hormone-responsive motifs. In contrast, TaSTP28 possessed the highest number of regulatory elements (55), including motifs related to abscisic acid, auxin, gibberellin, light regulation, and hypoxia. Statistical analysis of all TaMST genes revealed that 97.5%, 99.5%, and 100% of the members contained at least one hormone-responsive element, abiotic stress-related element, and growth/development-related regulatory element, respectively. These findings suggest that the TaMST gene family plays a pivotal role in regulating wheat growth and development, hormone response, and in mediating responses to abiotic stress. Expression analysis of the TaMST families in different tissues To study the expression levels of the TaMST genes in different tissues and organs, the RNA-Seq data of wheat in roots, leaves, buds, grains and spikes were analyzed using the public wheat database (Fig. 7). The results showed that the expression levels of TaMST family members were higher in leaves, buds, roots and spike types, accounting for 34.5%, 33.0%, 33.5% and 32.0% respectively (log 2 TPM+1 ＞2). The expression levels of genes in grains were overall lower, with 87.5% of genes having low expression levels (log 2 TPM+1 ＜2), and 70.5% of them were not expressed (log 2 TPM+1 ＝0). The distribution of gene expression levels in the TaMST subfamilies was uneven. For example, in the TaINT subfamily, 4 genes did not express in these tissues, while other genes had higher expression levels. The overall expression level of the TaSTP subfamily was very low, with 49.54% of TaSTP genes having almost no expression in these organs. TaPMT had one-third of its genes expressed only in roots, while TapGlcT had half of its genes not expressed in roots. The overall expression levels of TaERD and TaVGT subfamilies were relatively high. The uneven distribution of gene expression levels in different tissues indicated the functional differentiation of the TaMST gene family. Expression pattern analysis of wheat TaMST family in response to exogenous sugars In order to explore the expression of TaMST genes at the RNA level in response to different exogenous sugars, 8 TaMST genes were selected for RT-qPCR detection based on their expression levels and classification in roots and leaves (Fig. 8 and Fig. 9). It was found that the expression patterns of TaMST genes in roots and leaves were significantly different. In leaves, TaERD3 and TaTMT9 had decreased expression levels under the treatment of three sugars, but the decrease was not significant under fructose treatment. TaINT2 , TaPMT27 , TaSTP5 and TaSTP60 had increased expression levels under fructose treatment, with TaSTP59 and TaPMT27 having the highest increase, rising by 2.6 times and 1.6 times respectively. The expression levels of TaINT2 and TapGlcT1 had increased under glucose treatment, with TaINT2 having the most significant increase, rising by 1.2 times. TaPMT27 and TaSTP59 had significantly decreased under glucose treatment, while the expression levels of the other 4 genes did not change dramatically. TaPMT27 , TaSTP59 and TaVGT2 had up-regulated expression under sucrose treatment, with TaPMT27 having the most obvious increase, up by 7.2 times compared to CK. TaVGT2 only had increased under sucrose treatment, and there was no evidence change under the other two exogenous sugar treatments. In roots, all genes had noticeably decreased expression levels after treatment with three exogenous sugars, with 6 genes expressing a lower degree of down-regulation compared to other two sugars, and 6 genes having the highest degree after glucose treatment. The down-regulation trend of TaERD3 , TapGlcT , TaSTP59 and TaSTP60 was similar under the treatment of three exogenous sugars, with expression levels from high to low being fructose treatment, sucrose treatment and glucose treatment. TaPMT9 had a 4.0-fold and 19.6-fold higher expression level than other two exogenous sugars under exogenous sucrose treatment, consistent with the expression trend in leaves. Analyzing gene expression profiles in conjunction with evolutionary relationships facilitates the understanding of gene function(Sun et al., 2011). The first plant MST gene identified was AtSTP1 in Arabidopsis thaliana , which exhibited high expression levels in guard cells of (Sauer et al., 1990; Stadler et al., 2003). OsSTP3 was the first MST gene identified in rice and was found to be highly expressed in leaves, leaf sheaths, roots, and callus tissues, playing a role in the transport of monosaccharides essential for cell wall synthesis(Kyoko et al., 2000). TaSTP13 was the first MST gene cloned and characterized in wheat. Expression analysis revealed that TaSTP13 was predominantly expressed in source tissues such as leaves and exhibited significant responses to mechanical injury, low temperature, exogenous MeJA, and stripe rust infection, suggesting its potential involvement in the MeJA signaling pathway(Huai et al., 2020). With the exception of STP and TMT subfamilies, most members of the other five wheat MST subfamilies exhibited tissue-specific expression patterns in organs such as buds, leaves, spikes, grains, and roots, indicating their potential roles in wheat growth and development. These findings were consistent with previous studies in other species(Büttner, 2010; Mamun et al., 2006; Qin et al., 2018). TaPMT27 exhibited relatively high expression across all analyzed tissues and organs, suggesting its potentially broad functional role in wheat growth and development, possibly as a broad-spectrum monosaccharide transporter. TaSTP18 was highly expressed in both aerial parts and roots, indicating its involvement in both vegetative growth and carbohydrate allocation between leaf "source" and root "sink" tissues. Significant functional divergence existed among MST family members in terms of sugar transport specificity. For instance, exogenous glucose, fructose, and sucrose can remarkably induce the expression of OsSTP1 in rice, suggesting its role in the transport of these sugars. OsSTP4 was a constitutively expressed gene in rice, showing high expression in leaves, leaf sheaths, and embryos, and was capable of transporting fructose, mannose, glucose, and galactose. OsSTP13 and OsSTP27 were predominantly expressed in roots and aerial parts, respectively, and responded to exogenous hormones and sugars(Johnson and Thomas, 2007; Mamun et al., 2006). Notably, compared with Arabidopsis, the STP subfamily in rice has undergone significant expansion, and interspecies differences in expression patterns suggest distinct functional roles in plant development(Johnson et al., 2006). Research on other MST subfamilies remains limited. Exogenous fructose and glucose have been shown to regulate the expression of certain MST genes, indicating their potential roles in sugar sensing and transport. Subcellular localization analysis of wheat TaMST proteins To verify the subcellular localization of wheat TaMST proteins, three genes were selected for localization analysis. The results illustrated that the empty vector GFP was widely expressed in cells, and the green fluorescence signal was mainly distributed on the cell nucleus and cell membrane. While TaERD3, TaPMT29 and TaSTP18 were all located on the cell membrane, which was consistent with the results predicted through bioinformatics analysis previously (Fig. 10). Conclusion This study identified 200 members of the monosaccharide transporter (MST) gene family across the entire wheat genome. Based on gene structure and the topological arrangement of the phylogenetic tree, these members were classified into seven subfamilies: TaERD , TaINT , TapGlcT , TaPMT , TaSTP , TaTMT , and TaVGT . The TaSTP subfamily contained the largest number of members, with a total of 74, whereas the TaVGT subfamily had the fewest, with only 6. The 200 MST gene family members were evenly distributed across the 22 chromosomes of wheat. Motifs 2, 5, 8, and 13 were identified as conserved motifs shared among MST family members. Gene structure analysis revealed significant variation in the number of exons and introns among MST genes, ranging from 1 to 18 exons and 0 to 17 introns, respectively. Gene duplication events, including fragment duplication and tandem duplication, were found to play crucial roles in the expansion of the MST gene family. All TaMST genes were found to contain cis-regulatory elements associated with growth and development regulation. Expression analysis indicated that TaMST genes were most highly expressed in leaves and least expressed in grains. Furthermore, different MST family members exhibited complex response patterns to exogenous fructose, glucose, and sucrose, suggesting that MST genes primarily function in "source" organs and individual members may exhibit diverse sugar transport mechanisms. Declarations Author contributions Jun Zhang performed the bioinformatics analysis of the MSTs gene family in wheat ( Triticum aestivum L.), and was a major contributor in writing the manuscript. Bo Zhang wrote, reviewed and edited. Shuang Zhou analyzed the transcriptome data. Wenzhong Tian constructed the phylogenetic tree and analyzed the protein structural domains. Yiren Chen completed qPCR. Binbin Guo contributed to subcellular localization. Chao Ma provided conceptualization, project administration, and funding acquisition. All authors read and approved the final manuscript. Funding This work is supported by the National Natural Science Foundation of China (32372227), the Natural Science Foundation of Henan Province (242300421319), Henan Provincial Science and Technology Key Research Project (252102111075). Undergraduate Innovation and Entrepreneurship Training Program of Henan University of Science and Technology (2025424, 2025426, 2025435) Data availability The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request. Competing interests The authors declare that they have no competing interests. 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Zhu J, Li T, Ma J, Li W, Zhang H, Nadezhda T, Zhu Y, Dong X, Li C, Fan J (2024) Genome-wide identification and investigation of monosaccharide transporter gene family based on their evolution and expression analysis under abiotic stress and hormone treatments in maize ( Zea mays L.). BMC Plant Biol 24(1): 496. https://doi.org/10.1186/s12870-024-05186-2. Additional Declarations No competing interests reported. Supplementary Files SupplementaryTable.docx Cite Share Download PDF Status: Published Journal Publication published 21 Jan, 2026 Read the published version in Genetica → Version 1 posted Editorial decision: Revision requested 18 Sep, 2025 Editor assigned by journal 18 Sep, 2025 Submission checks completed at journal 09 Sep, 2025 First submitted to journal 08 Sep, 2025 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. 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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-7561375","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":516933859,"identity":"5c5f2de1-40ac-4893-9aa4-9abb4943008b","order_by":0,"name":"Jun Zhang","email":"","orcid":"","institution":"Henan University of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Jun","middleName":"","lastName":"Zhang","suffix":""},{"id":516933860,"identity":"8072b92d-5af8-4e8b-9c98-23353aec9784","order_by":1,"name":"Bo Zhang","email":"","orcid":"","institution":"Henan University of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Bo","middleName":"","lastName":"Zhang","suffix":""},{"id":516933861,"identity":"08b2eb4f-a91c-4e80-b442-05dca5b50e6a","order_by":2,"name":"Shuang Zhou","email":"","orcid":"","institution":"Henan University of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Shuang","middleName":"","lastName":"Zhou","suffix":""},{"id":516933866,"identity":"3b640c5a-8d46-4bd1-bc28-434079ac1f62","order_by":3,"name":"Wenzhong Tian","email":"","orcid":"","institution":"Luoyang Academy of Agriculture and Forestry Sciences","correspondingAuthor":false,"prefix":"","firstName":"Wenzhong","middleName":"","lastName":"Tian","suffix":""},{"id":516933868,"identity":"144ce3a2-958c-4d79-8984-be3b5dac4e55","order_by":4,"name":"Rong Zhang","email":"","orcid":"","institution":"Henan University of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Rong","middleName":"","lastName":"Zhang","suffix":""},{"id":516933871,"identity":"39904a59-9579-453a-a650-b2ce33b629e7","order_by":5,"name":"Yiren Chen","email":"","orcid":"","institution":"Henan University of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Yiren","middleName":"","lastName":"Chen","suffix":""},{"id":516933872,"identity":"e8a95629-7450-4745-9c57-2e8179b6614b","order_by":6,"name":"Binbin Guo","email":"","orcid":"","institution":"Henan University of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Binbin","middleName":"","lastName":"Guo","suffix":""},{"id":516933873,"identity":"b5c80e7d-64cd-4fcf-8ab1-48f81bbce79b","order_by":7,"name":"Chao Ma","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA5klEQVRIie3RsYrCQBCA4ZWFTRNNm7AH9wQHsyxEi8WXCUy1haXdmSbX5AHyGD5CvOGsAraCKQJCqhSWFgpnryS57or9YLr5i2EYc5z/STzGcPG+S5sLmOXYBL0g5KSKFSZjEwqiQqD0L9+TzdD+xxe1zXWNEk651gZKzjz62fYlcYVzlVdGQ12ps4V6xnzEY29SWhFOM0ygtFpbaDkL/bg/OXQiumf0uS1tLBdAk81gcrRCTjPiUYEo2bikjeVbhTzwiVQOmIjBWw5JG3Xrxyu9NG2uN7MMPNr3Js/E39Ydx3GcV34BP+FMGD4yMsgAAAAASUVORK5CYII=","orcid":"","institution":"Henan University of Science and Technology","correspondingAuthor":true,"prefix":"","firstName":"Chao","middleName":"","lastName":"Ma","suffix":""}],"badges":[],"createdAt":"2025-09-08 07:53:20","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7561375/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7561375/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1007/s10709-026-00257-8","type":"published","date":"2026-01-21T15:57:07+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":94825029,"identity":"8d67c32c-ac79-4c94-9f1f-3b22c50f9c81","added_by":"auto","created_at":"2025-10-31 06:49:44","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":1046403,"visible":true,"origin":"","legend":"\u003cp\u003ePhylogenetic relationships among members of TaMSTs family in wheat\u003c/p\u003e","description":"","filename":"Fig1.png","url":"https://assets-eu.researchsquare.com/files/rs-7561375/v1/feab92c03806c9cc1b130068.png"},{"id":94767758,"identity":"7a9678e6-ad98-46b1-9553-8e2c25436209","added_by":"auto","created_at":"2025-10-30 13:14:29","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":1587663,"visible":true,"origin":"","legend":"\u003cp\u003ePhylogenetic relationships among members of TaMST family in wheat\u003c/p\u003e\n\u003cp\u003eA:\u003cem\u003e \u003c/em\u003ePhylogenetic tree of the sugar transporter protein family; B: Conserved motifs of the \u003cem\u003eTaMST\u003c/em\u003e family; C：Gene structure of the\u003cem\u003e TaMST\u003c/em\u003efamily. CDS: Coding sequences; UTR: Untranslated regions.\u003c/p\u003e","description":"","filename":"Fig2.png","url":"https://assets-eu.researchsquare.com/files/rs-7561375/v1/45aa4995b0deab3d69e69e94.png"},{"id":94824963,"identity":"aa34b636-ed1e-4edf-b0fb-2bab99dbeed5","added_by":"auto","created_at":"2025-10-31 06:49:38","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":1342171,"visible":true,"origin":"","legend":"\u003cp\u003eIdentification of conserved sequence of \u003cem\u003eTaMST\u003c/em\u003efamily members in wheat\u003c/p\u003e","description":"","filename":"Fig3.png","url":"https://assets-eu.researchsquare.com/files/rs-7561375/v1/ba2cd8f7ab65c27c204c866e.png"},{"id":94767761,"identity":"6483b939-1d3a-4a8c-97f3-c493ea5c5c42","added_by":"auto","created_at":"2025-10-30 13:14:30","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":682721,"visible":true,"origin":"","legend":"\u003cp\u003eChromosomal distribution of \u003cem\u003eTaMST\u003c/em\u003egenes in wheat\u003c/p\u003e\n\u003cp\u003eThe red curve represents tandem duplication between genes; The color of the chromosome bands indicates the density of the genes.\u003c/p\u003e","description":"","filename":"Fig4.png","url":"https://assets-eu.researchsquare.com/files/rs-7561375/v1/8a50b6052255c316314f7b52.png"},{"id":94824844,"identity":"b5241615-4d7a-487c-aa17-b9531dd0123f","added_by":"auto","created_at":"2025-10-31 06:49:25","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":1260324,"visible":true,"origin":"","legend":"\u003cp\u003eRegional collinearity relationships of wheat \u003cem\u003eTaMST\u003c/em\u003e genes\u003c/p\u003e\n\u003cp\u003eDifferent colored lines represent chromosomal regions with homology in different subfamily genes; The bar and lines of rings 1 and 2 indicate the location of base deletions; Ring 3 is the gene density heatmap.\u003c/p\u003e","description":"","filename":"Fig5.png","url":"https://assets-eu.researchsquare.com/files/rs-7561375/v1/37e84db9a8e5a78f605588e7.png"},{"id":94767767,"identity":"8948132d-a83f-41b6-9d06-91ff4dfaf659","added_by":"auto","created_at":"2025-10-30 13:14:30","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":1318739,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003eCis\u003c/em\u003e-acting element of \u003cem\u003eTaMST\u003c/em\u003e genes promoter in wheat\u003c/p\u003e\n\u003cp\u003eDifferent promoter \u003cem\u003ecis\u003c/em\u003e-acting elements are represented by the dots of different colors.\u003c/p\u003e","description":"","filename":"Fig6.png","url":"https://assets-eu.researchsquare.com/files/rs-7561375/v1/53c2d48fe92551ff80dcf0e6.png"},{"id":94767765,"identity":"72812314-02c6-4c0d-8527-4883e0ec675e","added_by":"auto","created_at":"2025-10-30 13:14:30","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":1402512,"visible":true,"origin":"","legend":"\u003cp\u003eRelative expression pattern of wheat \u003cem\u003eTaMST\u003c/em\u003egenes in different tissues\u003c/p\u003e","description":"","filename":"Fig7.png","url":"https://assets-eu.researchsquare.com/files/rs-7561375/v1/ed853af5f7fc8fbf99f7f817.png"},{"id":94767763,"identity":"9fb2708c-f9c6-49e9-bc36-6aaa4df51ce6","added_by":"auto","created_at":"2025-10-30 13:14:30","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":214419,"visible":true,"origin":"","legend":"\u003cp\u003eRelative expression pattern of \u003cem\u003eTaMST\u003c/em\u003e genes family members under exogenous fructose(Fru), glucose(Glu), and sucrose(Suc) in roots (R)\u003c/p\u003e","description":"","filename":"Fig8.png","url":"https://assets-eu.researchsquare.com/files/rs-7561375/v1/6fd80a38361ef979b866f0c6.png"},{"id":94767766,"identity":"d33da9c3-69a3-4223-9fbe-ccac671fde6e","added_by":"auto","created_at":"2025-10-30 13:14:30","extension":"png","order_by":9,"title":"Figure 9","display":"","copyAsset":false,"role":"figure","size":216086,"visible":true,"origin":"","legend":"\u003cp\u003eRelative expression pattern of \u003cem\u003eTaMST\u003c/em\u003e genes family members under exogenous fructose(Fru), glucose(Glu), and sucrose(Suc) in leaves (L)\u003c/p\u003e","description":"","filename":"Fig9.png","url":"https://assets-eu.researchsquare.com/files/rs-7561375/v1/65c5ab22f8ae73fdaec88df3.png"},{"id":94767768,"identity":"4e57bfe3-dc56-49a5-9c64-d598c8e520d5","added_by":"auto","created_at":"2025-10-30 13:14:30","extension":"png","order_by":10,"title":"Figure 10","display":"","copyAsset":false,"role":"figure","size":895360,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSubcellular localization analysis of empty vector, TaERD3, TaPMT29, and TaSTP18\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Fig10.png","url":"https://assets-eu.researchsquare.com/files/rs-7561375/v1/eb8703cd2c99d50d088c7006.png"},{"id":101151658,"identity":"83698612-8431-41a2-9868-1fc0730055b5","added_by":"auto","created_at":"2026-01-26 16:00:54","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":10912084,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7561375/v1/ce6ba767-ef95-45ff-9c11-425e03976688.pdf"},{"id":94824822,"identity":"e153edf0-7bfe-4b3a-832c-fe27ba04a32b","added_by":"auto","created_at":"2025-10-31 06:49:23","extension":"docx","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":109231,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryTable.docx","url":"https://assets-eu.researchsquare.com/files/rs-7561375/v1/47e0b4269cee15e815923519.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Genome-wide identification and expression analysis of the monosaccharide transporter (MST) gene family in wheat (Triticum aestivum L.)","fulltext":[{"header":"Introduction","content":"\u003cp\u003eCarbohydrate serves as a fundamental substance for the growth, development, and energy metabolism of higher plants and constitutes a key component of carbon skeletons(Ma et al., \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). It regulates various physiological processes such as flowering, seed germination, root architecture, senescence, and stress responses through mechanisms including energy storage, osmotic adjustment, and signal transduction(Ruan, \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). Carbohydrate is synthesized and exported from \u0026ldquo;source\u0026rdquo; organs and accumulated or converted in \u0026ldquo;sink\u0026rdquo; organs. The \u0026ldquo;flow\u0026rdquo; between these organs primarily consists of fructose, glucose, and sucrose, which are transported over short or long distances to connect source and sink tissues. This intricate process requires the coordinated action of multiple sugar transporters for precise regulation(Eom et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). In plants, major sugar transporter families include Sugar Will Eventually Be Exported Transporters (SWEETs), Sucrose Transporters (SUTs), and Monosaccharide Transporters (MSTs)(Deng et al., \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Eom et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). MST proteins belong to the family of intrinsic membrane proteins and are primarily responsible for the transmembrane transport of various monosaccharides[4]. Wheat is classified as a \u0026ldquo;sugary leaf\u0026rdquo; plant, with sugar content accounting for up to 95% of the dry weight of its leaves. Even minor fluctuations in sugar levels can trigger significant changes in gene expression (Ma et al., \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). Therefore, identifying the MSTs gene family within the wheat genome and analyzing its expression patterns across different tissues, organs, and under exogenous sugar treatments will provide a foundation for further functional studies of MSTs in wheat and offer valuable insights for high-yield wheat breeding programs.\u003c/p\u003e\u003cp\u003eGenome-wide identification of the MST gene family has been conducted in several species, including Arabidopsis (B\u0026uuml;ttner, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2007\u003c/span\u003e; B\u0026uuml;ttner, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; Wormit et al., \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2007\u003c/span\u003e), grape(Afoufa-Bastien et al., \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2010\u003c/span\u003e), tobacco(Okubo-Kurihara et al., \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2011\u003c/span\u003e), tomato(Hackel et al., \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2006\u003c/span\u003e; Mccurdy et al., \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2010\u003c/span\u003e), alfalfa(Doidy et al., \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2012b\u003c/span\u003e), and rose(Henry et al., \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). Based on sequence characteristics and substrate specificity, the MST family can be categorized into seven subfamilies: hexose transporters (Sugar Transport Protein, STP), polyol/monosaccharide transporters (Polyol/Monosaccharide Transporter, PMT), early-responsive to dehydration six-like (ERD6), vacuolar membrane monosaccharide transporters (Tonoplast Membrane Transporter, TMT), inositol transporters (Inositol Transporter, INT), plastidic glucose transporters (Plastidic Glucose Transporter, pGlcT), and vacuolar glucose transporters (Vacuolar Glucose Transporter, VGT)(Johnson et al., \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2006\u003c/span\u003e; Johnson and Thomas, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). In Arabidopsis, identified STP proteins function as H\u003csup\u003e+\u003c/sup\u003e/hexose co-transporters localized on the plasma membrane. Most STPs exhibit broad substrate transport capabilities(B\u0026uuml;ttner, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; Zheng et al., \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). For example, AtSTP1, AtSTP2, AtSTP3, AtSTP4, AtSTP6, and AtSTP11 can transport glucose, xylose, mannose, and galactose with varying affinities but do not transport fructose (Cho et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). Conversely, AtSTP6, AtSTP13, and OsMST4 are capable of transporting fructose but not pentoses such as xylose and ribose(Afoufa-Bastien et al., \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; B\u0026uuml;ttner, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; Wang et al., \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). However, certain STP proteins demonstrate substrate specificity in transport. For instance, AtSTP9 exclusively transports glucose, while AtSTP14 specifically transports galactose(Poschet et al., \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; Schneider et al., \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2008\u003c/span\u003e). Additionally, PLT-like proteins have been shown to transport both polyols and monosaccharides(Klepek et al., \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2005\u003c/span\u003e; Klepek et al., \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). Among the XTPH (also known as VGT) proteins, AtVGT1 facilitates glucose transport but not xylose. Both AtVGT1 and AtVGT2 are H\u003csup\u003e+\u003c/sup\u003e/glucose antiporters located on the vacuolar membrane and play roles in the transport and storage of monosaccharides within the vacuole (B\u0026uuml;ttner, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2007\u003c/span\u003e; Wormit et al., \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). Similarly, AZT (also known as TMT) proteins are localized on the vacuolar membrane. Studies have demonstrated that in the \u003cem\u003eattmt1/attmt2\u003c/em\u003e double mutant of Arabidopsis, the vacuole's capacity to take up sucrose is significantly reduced, indicating that TMT proteins participate in the transmembrane transport of sucrose across the vacuolar membrane (Schulz et al., \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). Meanwhile, pGlcT proteins are involved in the transport of glucose (Cho et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). Within the MST family in Arabidopsis, ERD6 (also referred to as SFPs) represents the largest subfamily. The first identified ERD6 protein, AtERD6, is induced by drought and low temperature(Quirino et al., \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2001\u003c/span\u003e). Notably, AtSFP1 and AtSFP2 are stress-induced auxiliary diffusion transporters exhibiting distinct spatiotemporal expression patterns (Yamada et al., \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e2009\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eBeyond model species like Arabidopsis, several \u003cem\u003eMST\u003c/em\u003e genes have also been characterized in rice. For example, Toyofuku et al. cloned and analyzed three \u003cem\u003eMST\u003c/em\u003e genes, \u003cem\u003eOsMST1\u003c/em\u003e, \u003cem\u003eOsMST2\u003c/em\u003e, and \u003cem\u003eOsMST3\u003c/em\u003e, and found that \u003cem\u003eOsMST3\u003c/em\u003e mediates the transport of specific monosaccharides via energy-dependent H\u003csup\u003e+\u003c/sup\u003e co-transport(Kyoko et al., \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2000\u003c/span\u003e). Functional analysis through heterologous expression confirmed that \u003cem\u003eOsMST5\u003c/em\u003e plays a role in regulating pollen development in rice(Ngampanya et al., \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2003\u003c/span\u003e). Moreover, \u003cem\u003eOsMST4\u003c/em\u003e, which exhibits constitutive expression, can transport fructose, galactose, mannose, and glucose(Poschet et al., \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). Wang et al. reported that \u003cem\u003eOsMST6\u003c/em\u003e functions as a broad-spectrum monosaccharide transporter whose expression is induced by salt stress and sugar treatment(Wang et al., \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2008\u003c/span\u003e). Furthermore, OsTMTs are involved in vacuolar sugar transport(Cho et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). Additionally, \u003cem\u003eOsGMST1\u003c/em\u003e is up-regulated under salt stress conditions, and knocking down this gene significantly reduces the salt stress resistance of rice(Cao et al., \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2011\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eIn summary, the MST family genes exert an essential role in plant growth and development as well as the response to biological and abiotic stresses. This study was motivated by the importance of wheat as a globally significant staple crop and the increasing use of wheat genomic data for gene family identification. Previous studies have reported on gene families such as FDL(Kan et al., \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2025\u003c/span\u003e), NHX(Sharma et al., \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2023\u003c/span\u003e), GH13(Yin et al., \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e2024\u003c/span\u003e), and SNARE(Wang et al., \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). However, no comprehensive identification of the MST gene family in wheat has been reported to date, which has hindered functional studies of \u003cem\u003eMST\u003c/em\u003e genes in this species. To address this gap, the present study conducted a systematic bioinformatics analysis of the wheat MSTs gene family, focusing on physicochemical properties, gene structure, intraspecific collinearity, promoter cis-regulatory elements, expression profiles, subcellular localization, and protein interaction networks. Additionally, the expression patterns of MST genes across different wheat tissues were analyzed, and the effects of exogenous sugar signaling on MST gene expression were investigated. These findings provide a foundational reference for future functional studies of MST genes in wheat.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cp\u003e\u003cstrong\u003eGenome-wide identification of the \u003cem\u003eTaMST\u003c/em\u003e family in wheat \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe genome data of wheat was obtained from the EnsemblPlants database (http://plants.ensembl.org/index.html). The MST domain (PF00083) was retrieved from the Pfam database. HMMER 3.0 was employed to perform conserved domain searches for identifying members of the \u003cem\u003eTaMSTs\u003c/em\u003e gene family. Additionally, the protein sequence of \u003cem\u003eAtMSTs\u003c/em\u003e from \u003cem\u003eArabidopsis thaliana\u003c/em\u003e was used as a query for BLASTP analysis (E-value \u0026lt; 1E\u003csup\u003e-5\u003c/sup\u003e) to identify homologous sequences. Candidate \u003cem\u003eTaMST\u003c/em\u003e genes were selected based on combined results from BLAST and HMMER analyses. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eBasic physicochemical and phylogenetic analysis of the TaMSTs family in wheat \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe amino acid length, isoelectric point, and molecular weight of TaMST proteins were analyzed using the ExPASy ProtParam tool (https://web.expasy.org/protparam/). Subcellular localization prediction was conducted using ProtComp 9.0 software (https://linux1.softberry.com/berry.phtml). Transmembrane helices were predicted via the DTU Health Tech server (https://services.healthtech.dtu.dk/services/TMHMM-2.0/). The neighbor joining (NJ) phylogenetic tree was constructed by using mega7 tool with adjacency method (bootstrap value was set to 1000), and it was optimized by online software ChiPlot (https://www.chiplot.online/). \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eGene structure and conserved motif analysis of the TaMST family in wheat \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eGene structures were determined based on the gff annotation file. Conserved motifs among TaMST family members were identified using the MEME Suite (http://meme-suite.org/tools/meme), with the maximum number of motifs set to 20 and other parameters at default settings. Visualization of gene structures and motifs was performed using TBtools. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eChromosome localization and collinearity analysis of the \u003cem\u003eTaMSTs\u003c/em\u003e family in wheat \u0026nbsp;\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eChromosomal positions of \u003cem\u003eTaMST\u0026nbsp;\u003c/em\u003egenes were extracted from the IWGSC wheat genome annotation database (http://wheat-urgi.versailles.inra.fr/). Chromosomal localization was visualized by TBtools. Collinearity analysis was carried out through MCScanX and visualized via Advanced Circos. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eKa/Ks analysis of the\u003cem\u003e\u0026nbsp;TaMSTs\u003c/em\u003e family in wheat \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNucleotide sequences of \u003cem\u003eTaMST\u003c/em\u003e genes were downloaded from the EnsemblPlants database. MCScanX was used to identify collinear gene pairs. The Ka/Ks ratio was calculated by the TBtools implementation of the Ka/Ks Calculator program to assess selection pressures acting on duplicated gene pairs. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePromoter cis-regulatory element analysis of the \u003cem\u003eTaMSTs\u003c/em\u003e family in wheat \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe 2000-bp upstream promoter regions of \u003cem\u003eTaMST\u003c/em\u003e genes were extracted via TBtools. Potential cis-regulatory elements within these promoter sequences were predicted by the PlantCARE database (http://bioinformatics.psb.ugent.be/webtools/plantcare/html/), and the results were visualized according to TBtools. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eExpression patterns of the wheat \u003cem\u003eTaMSTs\u003c/em\u003e family across different tissues \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eExpression levels of \u003cem\u003eTaMST\u003c/em\u003e genes in root, grain, spike, leaf, and bud tissues of the Chinese Spring wheat variety were obtained from the ExpVIP wheat expression database (wheat-expression.com). Data were analyzed and visualized as a heatmap using TBtools. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eExpression analysis of the wheat \u003cem\u003eTaMSTs\u003c/em\u003e family under exogenous sugar treatments \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSeeds of the YM2 wheat cultivar with uniform size were surface-sterilized, rinsed thoroughly with distilled water, and soaked in sterile water for 12 hours. After germination, seedlings were transferred onto floating nets for hydroponic growth. At the two-leaf and one-heart stage, plants were subjected to four treatments: CK (distilled water), T1 (2% sucrose), T2 (2% fructose), and T3 (2% glucose). Each treatment had three biological replicates. Leaf samples were collected 24 hours after treatment and immediately frozen in liquid nitrogen. Total RNA was extracted using TRIZOL reagent (TaKaRa, 9108Q), reverse-transcribed into cDNA (TaKaRa, RR047Q), and analyzed by RT-qPCR (Vazyme, 221). Relative gene expression levels were calculated using the \u003cem\u003e2\u003csup\u003e⁻\u0026Delta;\u0026Delta;Ct\u003c/sup\u003e\u003c/em\u003e\u003csup\u003e\u0026nbsp;\u003c/sup\u003emethod. The primers used in this study were listed in Table S1.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eGene cloning, vector construction, and subcellular localization \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFull-length coding sequences of \u003cem\u003eTaERD3\u003c/em\u003e, \u003cem\u003eTaPMT2\u003c/em\u003e and \u003cem\u003eTaSTP18\u003c/em\u003e were amplified from YM 2 cDNA using specific primers (Table S1). PCR amplification was performed using 2\u0026times; Phanta Flash Master Mix (Dye Plus) with high-fidelity enzyme (P520, Vazyme). The reaction mixture consisted of 1 \u0026mu;L template, 1 \u0026mu;L each of forward and reverse primers, 10 \u0026mu;L master mix, and 7 \u0026mu;L ddH₂O. Amplification was carried out according to the manufacturer\u0026rsquo;s recommended thermal cycling conditions. After electrophoresis, the gel was recovered through the gel recovery kit (CW2302M, Cwbio). The sequences that were sequenced correctly were truncated at the terminators and added with restriction sites, and then subcloned into the \u003cem\u003epCAMBIA1300-GFP\u0026nbsp;\u003c/em\u003evector. The recombinant plasmid was transformed into Agrobacterium GV3101. The leaves of \u003cem\u003eNicotiana benthamiana\u003c/em\u003e were injected on the back side and cultured for 48 hours. The subcellular localization signals were observed via a laser confocal microscope (LSM710, Zeiss).\u003c/p\u003e"},{"header":"Results and discussion ","content":"\u003cp\u003e\u003cstrong\u003eGenome-wide identification of the TaMSTs family in wheat\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eBased on the protein sequences of 53 AtMSTs from \u003cem\u003eArabidopsis\u003c/em\u003e, a total of 260 \u003cem\u003eTaMST\u003c/em\u003e candidate genes were identified in wheat through BLASTP and HMM searches. After filtering out genes with incomplete domains and redundant sequences using NCBI-CD, a final set of 200 \u003cem\u003eTaMST\u003c/em\u003e genes was obtained. These genes were systematically renamed according to their chromosomal positions and corresponding protein sequences, for example, from \u003cem\u003eTaSTP1\u003c/em\u003e to \u003cem\u003eTaSTP74\u003c/em\u003e. The physicochemical properties and subcellular localization data are summarized in Supplementary Table S2. Analysis of the primary structure of all TaMST proteins revealed that the amino acid length ranged from 393 to 591 residues, molecular weights varied between 41.91 and 63.04 kDa, theoretical isoelectric points (pI) ranged from 4.90 to 10.05, with 40 acidic amino acids, 1 neutral amino acid, and the rest being alkaline amino acids. The protein instability index ranged from 28.99 to 54.12, with those below 40 being stable proteins. There were a total of 131 proteins classified as stable proteins. The aliphatic index ranged from 94.67 to 119.27, while the average hydrophobicity index ranged from 0.368 to 0.803. Among these, 36 proteins were predicted to be amphipathic, and the rest were hydrophobic. Subcellular localization predictions indicated that two TaMST proteins were localized to thylakoid membranes, eight to the endoplasmic reticulum, 38 to vacuolar membranes, and the remaining to the plasma membrane. Transmembrane helix prediction analysis showed that TaMST proteins contain approximately 8 to 12 transmembrane helices.\u003c/p\u003e\n\u003cp\u003eMonosaccharide transporters (MSTs) play essential roles in model plant species such as Arabidopsis and rice(B\u0026uuml;ttner, 2007; Cho et al., 2010). However, in other crops, including wheat, the identification and expression profiling of MST gene families remain limited. In 2020, researchers from the Crop Development Centre at the University of Saskatchewan, in collaboration with several leading international wheat genome research teams, published the genome sequences of 16 representative wheat varieties, representing the most comprehensive wheat genome resource available to date. This milestone has significantly advanced wheat genomics and provided a solid foundation for the identification and functional analysis of the wheat MST gene family(Walkowiak et al., 2020). In this study, a comprehensive analysis of the gene structure, phylogeny, and conserved protein motifs of wheat MST family members was conducted for the first time. This discrepancy may be attributed to the hexaploid nature of wheat. Most MST proteins were basic (pI\u0026gt;7), likely due to their high content of basic amino acids, suggesting that these transporters predominantly function in alkaline environments. Using BLAST searches and HMMER analysis, a total of 200\u003cem\u003e\u0026nbsp;TaMST\u003c/em\u003e genes were identified in the wheat genome, which is substantially higher than the 53 in Arabidopsis(B\u0026uuml;ttner, 2007), 65 in rice(Johnson and Thomas, 2007), 59 in grape(Afoufa-Bastien et al., 2010), and 58 in \u003cem\u003eMedicago truncatula\u003c/em\u003e (Doidy et al., 2012a).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePhylogenetic analysis, gene structure, and conserved motif analysis of the TaMSTs family in wheat\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePhylogenetic analysis of the TaMSTs amino acid sequences was conducted using MEGAX software (Fig. 1). In terms of the resulting phylogenetic tree, the TaMSTs family can be classified into seven distinct subfamilies: TaERD, TaINT, TapGlcT, TaPMT, TaSTP, TaTMT, and TaVGT. The TaSTPs subfamily contained the largest number of members (74), whereas the TaVGTs subfamily had the fewest (6). Other subfamilies included TaERDs (34), TaINTs (9), TapGlcTs (9), TaPMTs (48), and TaTMTs (20).\u003c/p\u003e\n\u003cp\u003eThe exon-intron structures of \u003cem\u003eTaMST\u003c/em\u003es were determined based on gene annotation data (Fig. 2). The analysis results indicated that among the 200 \u003cem\u003eTaMST\u003c/em\u003e family members, 196 contained both exons and introns, with a notable variation observed in the number of exons and introns. The number of exons ranged from 1 to 18, while the number of introns varied from 1 to 17. Furthermore, four \u003cem\u003eTaMST\u003c/em\u003e members were found to consist solely of a single exon without any introns. Members within the same subfamily generally exhibited similar exon-intron structures. Notably, most members of the \u003cem\u003eTaERD\u003c/em\u003e, \u003cem\u003eTapGlcT\u003c/em\u003e, and \u003cem\u003eTaVGT\u003c/em\u003e subfamilies contained approximately 16 exons. In contrast, \u003cem\u003eTaPMT\u003c/em\u003e and 90% of \u003cem\u003eTaTMT\u003c/em\u003e members involved only 2~3 introns, and the remaining subfamilies had fewer than six exons per gene. Additionally, 16 genes lacked upstream or downstream untranslated regions (UTRs), which may affect their regulatory functions.\u003c/p\u003e\n\u003cp\u003eConserved motifs among the 200 TaMST family members were analyzed using the online tool MEME (Fig. 2, Fig. 3), identifying a total of 20 distinct motifs (Motif 1 to Motif 20). Within each subfamily, the composition and sequence of conserved motifs were relatively consistent. The TaSTP subfamily exhibited the highest number of motifs (approximately 16), while the TaTMT subfamily had the fewest (an average of 12). Motifs 2, 5, 8, and 13 were present in all genes, whereas Motif 16 was exclusive to the TaSTP subfamily and Motif 17 was unique to the TaTMT subfamily. These differences in motif distribution suggest that the TaMSTs gene family has undergone functional divergence during evolution.\u003c/p\u003e\n\u003cp\u003ePhylogenetic analysis classified the \u003cem\u003eTaMST\u003c/em\u003e family into seven distinct subfamilies: \u003cem\u003eTaERD\u003c/em\u003e, \u003cem\u003eTaINT\u003c/em\u003e, \u003cem\u003eTapGlcT\u003c/em\u003e, \u003cem\u003eTaPMT\u003c/em\u003e, \u003cem\u003eTaSTP\u003c/em\u003e, \u003cem\u003eTaTMT\u003c/em\u003e, and \u003cem\u003eTaVGT\u003c/em\u003e. Among these, the \u003cem\u003eTaSTP\u003c/em\u003e subfamily contains the highest number of members (74), whereas the \u003cem\u003eTaVGT\u003c/em\u003e subfamily contains the fewest (6). Members of the \u003cem\u003eTaERD\u003c/em\u003e, \u003cem\u003eTapGlcT\u003c/em\u003e, and \u003cem\u003eTaVGT\u003c/em\u003e subfamilies possess a relatively high number of exons (approximately 16), whereas the remaining subfamilies contain fewer than six exons. These structural variations may result from evolutionary events such as exon and intron insertions or deletions, exonization, and pseudo-exonization, reflecting the structural diversity of \u003cem\u003eMST\u003c/em\u003e genes(Xu et al., 2012).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eChromosome Distribution and Co-linearity Analysis of the \u003cem\u003eTaMSTs\u003c/em\u003e Family in Wheat\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAccording to the chromosome location map of the \u003cem\u003eTaMST\u003c/em\u003e genes (Fig. 4), a total of 200 \u003cem\u003eTaMST\u003c/em\u003e genes are distributed across 22 wheat chromosomes, with a relatively even distribution among the three sub-genomes. Specifically, the A, B, and D genomes contain 67, 67, and 63 genes, respectively. Chromosomes 2 and 5 exhibit relatively higher gene densities, particularly chromosome 2, which harbors 68 \u003cem\u003eTaMST\u003c/em\u003e genes. In contrast, chromosomes 3, 6, and 7 display lower gene counts, with 12, 11, and 14 genes, respectively.\u003c/p\u003e\n\u003cp\u003eCo-linearity analysis of the\u003cem\u003e\u0026nbsp;TaMST\u003c/em\u003e gene family was performed using MCScanX (Fig. 5), revealing that 55 \u003cem\u003eTaMST\u003c/em\u003e genes formed 23 tandem repeat gene pairs. Among these, the \u003cem\u003eTaERD\u003c/em\u003e, \u003cem\u003eTaPMT\u003c/em\u003e, \u003cem\u003eTaSTP\u003c/em\u003e, and \u003cem\u003eTaTMT\u003c/em\u003e sub-families contained 4, 8, 7, and 4 tandem repeat gene pairs, respectively, and no tandem repeats were identified in other sub-families. A total of 81,798 co-linear regions were detected in the wheat genome, with 130 \u003cem\u003eTaMST\u003c/em\u003e genes located within these regions. These genes were primarily generated through segmental duplication events. The \u003cem\u003eTaSTP\u003c/em\u003e sub-family exhibited the highest number of segmental duplication genes, accounting for 56 genes, whereas the \u003cem\u003eTaVGT\u003c/em\u003e sub-family had the fewest, with only 4 genes. The significant expansion of the \u003cem\u003eTaSTP\u003c/em\u003e sub-family suggests that both tandem and segmental duplication mechanisms have played crucial roles in the evolutionary expansion of the \u003cem\u003eTaMST\u003c/em\u003e gene family. Furthermore, Ka/Ks ratio analysis revealed that all values were less than 1, indicating that these genes have undergone purifying selection and have been evolutionarily conserved (Supplementary Table S3).\u003c/p\u003e\n\u003cp\u003ePhylogenetic and evolutionary analyses revealed that MST family members across various species, including Arabidopsis, rice, and grape, can be classified into seven subfamilies. The subfamily classification of wheat \u003cem\u003eMST\u003c/em\u003e genes aligns with that of other species, suggesting that wheat, despite being a hexaploid organism, has not developed lineage-specific \u003cem\u003eMST\u003c/em\u003e genes. Within the wheat \u003cem\u003eMST\u003c/em\u003e gene family, all seven subfamilies contain four conserved motifs, whereas in maize, only three conserved motifs are shared among MST members(Zhu et al., 2024). The number of conserved motifs varies across subfamilies, indicating both structural conservation and functional divergence among MST subfamilies(Ding et al., 2023). Gene structure analysis further revealed that MST members within the same subfamily exhibit similar exon numbers and conserved motifs, implying potential functional similarity among subfamily members. Whole genome duplication events significantly contribute to the expansion and diversification of plant gene families, influencing both gene structure and function. Polyploidization, a key component of whole genome duplication in wheat, plays a major role in the expansion of gene families(Pozo and Ramirez-Parra, 2015). In Arabidopsis, the genes \u003cem\u003eAtERD6-like4\u003c/em\u003e, \u003cem\u003eAtINT2\u003c/em\u003e, \u003cem\u003eAtpGlcT4\u003c/em\u003e, \u003cem\u003eAtPMT4\u003c/em\u003e, \u003cem\u003eAtSTP1\u003c/em\u003e, \u003cem\u003eAtTMT2\u003c/em\u003e, and \u003cem\u003eAtVGT3\u0026nbsp;\u003c/em\u003erepresent the ancestral members of the seven \u003cem\u003eMST\u003c/em\u003e subfamilies. The wheat \u003cem\u003eMST\u0026nbsp;\u003c/em\u003esubfamilies identified in this study show direct orthologous relationships with these ancestral Arabidopsis genes, indicating that substantial gene duplication events have occurred within the wheat \u003cem\u003eMST\u0026nbsp;\u003c/em\u003egene family. Research by Johnson et al. further supports this, demonstrating that following the divergence of monocot and dicot plants, distinct mechanisms of whole genome duplication occurred in each lineage, collectively contributing to the evolutionary divergence of the wheat \u003cem\u003eMST\u003c/em\u003e subfamilies(Johnson and Thomas, 2007). The collinearity analysis results revealed that 130 wheat \u003cem\u003eMST\u003c/em\u003e genes were located within collinear regions, and these genes were primarily generated through segmental duplication. Among them, the \u003cem\u003eTaSTP\u003c/em\u003e subfamily exhibited the highest number of segmentally duplicated genes, encompassing 56 related members. Furthermore, 55 wheat \u003cem\u003eMST\u003c/em\u003e genes constituted 23 pairs of tandem repeats, indicating that both tandem and segmental duplication mechanisms have played vital roles in the expansion of the \u003cem\u003eMST\u003c/em\u003e gene family. In addition, all Ka/Ks ratios of these duplicated gene pairs were less than 1, suggesting that these \u003cem\u003eMST\u003c/em\u003e genes have undergone purifying selection and have been evolutionarily conserved.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAnalysis of \u003cem\u003ecis\u003c/em\u003e-acting elements in the TaMST family of wheat\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo further elucidate the functional characteristics of the \u003cem\u003eTaMST\u003c/em\u003e gene family, \u003cem\u003ecis\u003c/em\u003e-acting elements within the 2 kb upstream promoter regions of these genes were analyzed. Eighteen representative elements were selected for visualization (Fig. 6). The results indicated the presence of three major categories of \u003cem\u003ecis\u003c/em\u003e-acting elements: those associated with growth and development, hormone responsiveness, and abiotic stress responses. Within the growth and development-related category, several photosynthesis-related elements, such as G-Box, C-Box, MRE, and Sp1, were commonly identified across all members. Additionally, root-specific regulatory elements, meristem tissue-specific elements, and palisade mesophyll cell differentiation-related elements were also detected. Regarding abiotic stress response elements, drought-inducible elements, defense/stress-responsive elements, cold stress-responsive elements, and hypoxia-inducible elements were identified. Notably, 77 and 66 genes contained drought- and cold-responsive elements, respectively. With respect to hormone-responsive elements, 60.5%, 27.5%, and 90.0% of the genes contained auxin-, gibberellin-, and abscisic acid-responsive elements, respectively. The gene \u003cem\u003eTaERD25\u003c/em\u003e exhibited the lowest number of regulatory elements, with only seven, and lacked any hormone-responsive motifs. In contrast, \u003cem\u003eTaSTP28\u003c/em\u003e possessed the highest number of regulatory elements (55), including motifs related to abscisic acid, auxin, gibberellin, light regulation, and hypoxia. Statistical analysis of all \u003cem\u003eTaMST\u003c/em\u003e genes revealed that 97.5%, 99.5%, and 100% of the members contained at least one hormone-responsive element, abiotic stress-related element, and growth/development-related regulatory element, respectively. These findings suggest that the \u003cem\u003eTaMST\u003c/em\u003e gene family plays a pivotal role in regulating wheat growth and development, hormone response, and in mediating responses to abiotic stress.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eExpression analysis of the \u003cem\u003eTaMST\u003c/em\u003e families in different tissues\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo study the expression levels of the \u003cem\u003eTaMST\u0026nbsp;\u003c/em\u003egenes in different tissues and organs, the RNA-Seq data of wheat in roots, leaves, buds, grains and spikes were analyzed using the public wheat database (Fig. 7). The results showed that the expression levels of \u003cem\u003eTaMST\u003c/em\u003e family members were higher in leaves, buds, roots and spike types, accounting for 34.5%, 33.0%, 33.5% and 32.0% respectively (log\u003csub\u003e2\u003c/sub\u003e\u003csup\u003eTPM+1\u003c/sup\u003e＞2). The expression levels of genes in grains were overall lower, with 87.5% of genes having low expression levels (log\u003csub\u003e2\u003c/sub\u003e\u003csup\u003eTPM+1\u003c/sup\u003e＜2), and 70.5% of them were not expressed (log\u003csub\u003e2\u003c/sub\u003e\u003csup\u003eTPM+1\u003c/sup\u003e＝0). The distribution of gene expression levels in the \u003cem\u003eTaMST\u003c/em\u003e subfamilies was uneven. For example, in the \u003cem\u003eTaINT\u003c/em\u003e subfamily, 4 genes did not express in these tissues, while other genes had higher expression levels. The overall expression level of the \u003cem\u003eTaSTP\u003c/em\u003e subfamily was very low, with 49.54% of \u003cem\u003eTaSTP\u003c/em\u003e genes having almost no expression in these organs. \u003cem\u003eTaPMT\u003c/em\u003e had one-third of its genes expressed only in roots, while \u003cem\u003eTapGlcT\u003c/em\u003e had half of its genes not expressed in roots. The overall expression levels of \u003cem\u003eTaERD\u003c/em\u003e and \u003cem\u003eTaVGT\u003c/em\u003e subfamilies were relatively high. The uneven distribution of gene expression levels in different tissues indicated the functional differentiation of the \u003cem\u003eTaMST\u003c/em\u003e gene family.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eExpression pattern analysis of wheat \u003cem\u003eTaMST\u003c/em\u003e family in response to exogenous sugars\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn order to explore the expression of \u003cem\u003eTaMST\u003c/em\u003e genes at the RNA level in response to different exogenous sugars, 8 \u003cem\u003eTaMST\u0026nbsp;\u003c/em\u003egenes were selected for RT-qPCR detection based on their expression levels and classification in roots and leaves (Fig. 8 and Fig. 9). It was found that the expression patterns of \u003cem\u003eTaMST\u003c/em\u003e genes in roots and leaves were significantly different.\u003c/p\u003e\n\u003cp\u003eIn leaves, \u003cem\u003eTaERD3\u003c/em\u003e and \u003cem\u003eTaTMT9\u003c/em\u003e had decreased expression levels under the treatment of three sugars, but the decrease was not significant under fructose treatment. \u003cem\u003eTaINT2\u003c/em\u003e, \u003cem\u003eTaPMT27\u003c/em\u003e, \u003cem\u003eTaSTP5\u003c/em\u003e and \u003cem\u003eTaSTP60\u003c/em\u003e had increased expression levels under fructose treatment, with \u003cem\u003eTaSTP59\u003c/em\u003e and \u003cem\u003eTaPMT27\u003c/em\u003e having the highest increase, rising by 2.6 times and 1.6 times respectively. The expression levels of \u003cem\u003eTaINT2\u003c/em\u003e and \u003cem\u003eTapGlcT1\u003c/em\u003e had increased under glucose treatment, with \u003cem\u003eTaINT2\u003c/em\u003e having the most significant increase, rising by 1.2 times. \u003cem\u003eTaPMT27\u003c/em\u003e and \u003cem\u003eTaSTP59\u003c/em\u003e had significantly decreased under glucose treatment, while the expression levels of the other 4 genes did not change dramatically. \u003cem\u003eTaPMT27\u003c/em\u003e, \u003cem\u003eTaSTP59\u003c/em\u003e and \u003cem\u003eTaVGT2\u003c/em\u003e had up-regulated expression under sucrose treatment, with \u003cem\u003eTaPMT27\u003c/em\u003e having the most obvious increase, up by 7.2 times compared to CK. \u003cem\u003eTaVGT2\u003c/em\u003e only had increased under sucrose treatment, and there was no evidence change under the other two exogenous sugar treatments. In roots, all genes had noticeably decreased expression levels after treatment with three exogenous sugars, with 6 genes expressing a lower degree of down-regulation compared to other two sugars, and 6 genes having the highest degree after glucose treatment. The down-regulation trend of \u003cem\u003eTaERD3\u003c/em\u003e, \u003cem\u003eTapGlcT\u003c/em\u003e, \u003cem\u003eTaSTP59\u003c/em\u003e and \u003cem\u003eTaSTP60\u003c/em\u003e was similar under the treatment of three exogenous sugars, with expression levels from high to low being fructose treatment, sucrose treatment and glucose treatment. \u003cem\u003eTaPMT9\u003c/em\u003e had a 4.0-fold and 19.6-fold higher expression level than other two exogenous sugars under exogenous sucrose treatment, consistent with the expression trend in leaves.\u003c/p\u003e\n\u003cp\u003eAnalyzing gene expression profiles in conjunction with evolutionary relationships facilitates the understanding of gene function(Sun et al., 2011). The first plant \u003cem\u003eMST\u003c/em\u003e gene identified was \u003cem\u003eAtSTP1\u0026nbsp;\u003c/em\u003ein \u003cem\u003eArabidopsis thaliana\u003c/em\u003e, which exhibited high expression levels in guard cells of (Sauer et al., 1990; Stadler et al., 2003). \u003cem\u003eOsSTP3\u003c/em\u003e was the first \u003cem\u003eMST\u003c/em\u003e gene identified in rice and was found to be highly expressed in leaves, leaf sheaths, roots, and callus tissues, playing a role in the transport of monosaccharides essential for cell wall synthesis(Kyoko et al., 2000). \u003cem\u003eTaSTP13\u003c/em\u003e was the first \u003cem\u003eMST\u003c/em\u003e gene cloned and characterized in wheat. Expression analysis revealed that \u003cem\u003eTaSTP13\u003c/em\u003e was predominantly expressed in source tissues such as leaves and exhibited significant responses to mechanical injury, low temperature, exogenous MeJA, and stripe rust infection, suggesting its potential involvement in the MeJA signaling pathway(Huai et al., 2020). With the exception of \u003cem\u003eSTP\u003c/em\u003e and \u003cem\u003eTMT\u003c/em\u003e subfamilies, most members of the other five wheat \u003cem\u003eMST\u003c/em\u003e subfamilies exhibited tissue-specific expression patterns in organs such as buds, leaves, spikes, grains, and roots, indicating their potential roles in wheat growth and development. These findings were consistent with previous studies in other species(B\u0026uuml;ttner, 2010; Mamun et al., 2006; Qin et al., 2018). \u003cem\u003eTaPMT27\u0026nbsp;\u003c/em\u003eexhibited relatively high expression across all analyzed tissues and organs, suggesting its potentially broad functional role in wheat growth and development, possibly as a broad-spectrum monosaccharide transporter. \u003cem\u003eTaSTP18\u003c/em\u003e was highly expressed in both aerial parts and roots, indicating its involvement in both vegetative growth and carbohydrate allocation between leaf \u0026quot;source\u0026quot; and root \u0026quot;sink\u0026quot; tissues. Significant functional divergence existed among \u003cem\u003eMST\u003c/em\u003e family members in terms of sugar transport specificity. For instance, exogenous glucose, fructose, and sucrose can remarkably induce the expression of \u003cem\u003eOsSTP1\u003c/em\u003e in rice, suggesting its role in the transport of these sugars. \u003cem\u003eOsSTP4\u0026nbsp;\u003c/em\u003ewas a constitutively expressed gene in rice, showing high expression in leaves, leaf sheaths, and embryos, and was capable of transporting fructose, mannose, glucose, and galactose. \u003cem\u003eOsSTP13\u0026nbsp;\u003c/em\u003eand \u003cem\u003eOsSTP27\u003c/em\u003e were predominantly expressed in roots and aerial parts, respectively, and responded to exogenous hormones and sugars(Johnson and Thomas, 2007; Mamun et al., 2006). Notably, compared with Arabidopsis, the \u003cem\u003eSTP\u003c/em\u003e subfamily in rice has undergone significant expansion, and interspecies differences in expression patterns suggest distinct functional roles in plant development(Johnson et al., 2006). Research on other \u003cem\u003eMST\u003c/em\u003e subfamilies remains limited. Exogenous fructose and glucose have been shown to regulate the expression of certain \u003cem\u003eMST\u003c/em\u003e genes, indicating their potential roles in sugar sensing and transport.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSubcellular localization analysis of wheat TaMST proteins\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo verify the subcellular localization of wheat TaMST proteins, three genes were selected for localization analysis. The results illustrated that the empty vector GFP was widely expressed in cells, and the green fluorescence signal was mainly distributed on the cell nucleus and cell membrane. While TaERD3, TaPMT29 and TaSTP18 were all located on the cell membrane, which was consistent with the results predicted through bioinformatics analysis previously (Fig. 10).\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThis study identified 200 members of the monosaccharide transporter (MST) gene family across the entire wheat genome. Based on gene structure and the topological arrangement of the phylogenetic tree, these members were classified into seven subfamilies: \u003cem\u003eTaERD\u003c/em\u003e, \u003cem\u003eTaINT\u003c/em\u003e, \u003cem\u003eTapGlcT\u003c/em\u003e, \u003cem\u003eTaPMT\u003c/em\u003e, \u003cem\u003eTaSTP\u003c/em\u003e, \u003cem\u003eTaTMT\u003c/em\u003e, and \u003cem\u003eTaVGT\u003c/em\u003e. The \u003cem\u003eTaSTP\u003c/em\u003e subfamily contained the largest number of members, with a total of 74, whereas the \u003cem\u003eTaVGT\u003c/em\u003e subfamily had the fewest, with only 6. The 200 \u003cem\u003eMST\u003c/em\u003e gene family members were evenly distributed across the 22 chromosomes of wheat. Motifs 2, 5, 8, and 13 were identified as conserved motifs shared among \u003cem\u003eMST\u003c/em\u003e family members. Gene structure analysis revealed significant variation in the number of exons and introns among \u003cem\u003eMST\u003c/em\u003e genes, ranging from 1 to 18 exons and 0 to 17 introns, respectively. Gene duplication events, including fragment duplication and tandem duplication, were found to play crucial roles in the expansion of the \u003cem\u003eMST\u003c/em\u003e gene family. All \u003cem\u003eTaMST\u003c/em\u003e genes were found to contain cis-regulatory elements associated with growth and development regulation. Expression analysis indicated that \u003cem\u003eTaMST\u003c/em\u003e genes were most highly expressed in leaves and least expressed in grains. Furthermore, different \u003cem\u003eMST\u003c/em\u003e family members exhibited complex response patterns to exogenous fructose, glucose, and sucrose, suggesting that \u003cem\u003eMST\u003c/em\u003e genes primarily function in \"source\" organs and individual members may exhibit diverse sugar transport mechanisms.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAuthor contributions\u0026nbsp;\u003c/strong\u003eJun Zhang performed the bioinformatics analysis of the MSTs gene family in wheat (\u003cem\u003eTriticum aestivum\u003c/em\u003e L.), and was a major contributor in writing the manuscript. Bo Zhang wrote, reviewed and edited. Shuang Zhou analyzed the transcriptome data. Wenzhong Tian constructed the phylogenetic tree and analyzed the protein structural domains. Yiren Chen completed qPCR. \u0026nbsp;Binbin Guo contributed to subcellular localization. Chao Ma provided conceptualization, project administration, and funding acquisition. All authors read and approved the final manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u0026nbsp;\u003c/strong\u003eThis work is supported by the National Natural Science Foundation of China (32372227), the Natural Science Foundation of Henan Province (242300421319), Henan Provincial Science and Technology Key Research Project (252102111075). Undergraduate Innovation and Entrepreneurship Training Program of Henan University of Science and Technology (2025424, 2025426, 2025435)\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003e\u0026nbsp;\u0026nbsp;\u003c/em\u003e\u003c/strong\u003eThe datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003e\u0026nbsp;\u0026nbsp;\u003c/em\u003e\u003c/strong\u003eThe authors declare that they have no competing interests.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eAfoufa-Bastien D, Medici A, Jeauffre J, Coutos-Th\u0026eacute;venot P, Lemoine R, Atanassova R, Laloi M (2010) The Vitis vinifera sugar transporter gene family: phylogenetic overview and macroarray expression profiling. 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BMC Plant Biol 24(1): 496. https://doi.org/10.1186/s12870-024-05186-2.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"genetica","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"gene","sideBox":"Learn more about [Genetica](http://link.springer.com/journal/10709)","snPcode":"10709","submissionUrl":"https://submission.nature.com/new-submission/10709/3","title":"Genetica","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Wheat, Monosaccharide transporter, Functional divergence, Expression analysis, Subcellular localization","lastPublishedDoi":"10.21203/rs.3.rs-7561375/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7561375/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eCarbohydrates function as both energy sources and signaling molecules in various critical physiological processes. Monosaccharide transporters (MSTs) are a class of membrane-bound carrier proteins in crops that mediate the transmembrane transport of monosaccharides, thereby playing a central role in crop growth and development, resource allocation, and responses to environmental stimuli. In this study, a total of 200 \u003cem\u003eMST\u003c/em\u003e family genes were identified in wheat and categorized into seven subfamilies. Twenty conserved motifs were detected within the TaMST family, with each subfamily exhibiting similar conserved motif patterns. The \u003cem\u003eTaMST\u003c/em\u003e gene family was evenly distributed across the three wheat subgenomes, with both segmental and tandem duplications contributing to gene family expansion. The TaMST gene family was found to contain numerous cis-regulatory elements associated with growth and development, hormone signaling, and abiotic stress responses. Expression analysis revealed that most \u003cem\u003eTaMSTs\u003c/em\u003e were expressed at low levels in wheat grains, whereas 69, 66, 67, and 64 genes exhibited high expression levels in leaves, buds, roots, and spikes, respectively. Following exogenous sugar treatments, the expression of all \u003cem\u003eTaMSTs\u003c/em\u003e in roots was down-regulated, while 4, 2, and 3 genes showed up-regulated expression in leaves after treatment with fructose, glucose, and sucrose, respectively. Subcellular localization displayed TaERD3, TaPMT29 and TaSTP18 were all located on the cell membrane. These findings suggest that MSTs play essential roles not only in wheat organ development but also in the perception and response to sugar signaling. This study provides valuable insights for future investigations into the functional divergence of the MST gene family.\u003c/p\u003e","manuscriptTitle":"Genome-wide identification and expression analysis of the monosaccharide transporter (MST) gene family in wheat (Triticum aestivum L.)","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-10-30 13:14:25","doi":"10.21203/rs.3.rs-7561375/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-09-18T08:05:53+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-09-18T07:49:09+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-09-09T05:16:25+00:00","index":"","fulltext":""},{"type":"submitted","content":"Genetica","date":"2025-09-08T07:39:28+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"genetica","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"gene","sideBox":"Learn more about [Genetica](http://link.springer.com/journal/10709)","snPcode":"10709","submissionUrl":"https://submission.nature.com/new-submission/10709/3","title":"Genetica","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"25c5144a-8ec4-451c-a341-2befcb1454c0","owner":[],"postedDate":"October 30th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2026-01-26T15:59:46+00:00","versionOfRecord":{"articleIdentity":"rs-7561375","link":"https://doi.org/10.1007/s10709-026-00257-8","journal":{"identity":"genetica","isVorOnly":false,"title":"Genetica"},"publishedOn":"2026-01-21 15:57:07","publishedOnDateReadable":"January 21st, 2026"},"versionCreatedAt":"2025-10-30 13:14:25","video":"","vorDoi":"10.1007/s10709-026-00257-8","vorDoiUrl":"https://doi.org/10.1007/s10709-026-00257-8","workflowStages":[]},"version":"v1","identity":"rs-7561375","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7561375","identity":"rs-7561375","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}