Developmental rewiring of theNGAL/CUC/KLUnetwork associated with pleiotropic roles ofNGALgenes

Developmental rewiring of the NGAL/CUC/KLU network associated with pleiotropic roles of NGAL genes | bioRxiv /* */ /* */ <!-- <!-- /*! * yepnope1.5.4 * (c) WTFPL, GPLv2 */ (function(a,b,c){function d(a){return"[object Function]"==o.call(a)}function e(a){return"string"==typeof a}function f(){}function g(a){return!a||"loaded"==a||"complete"==a||"uninitialized"==a}function h(){var a=p.shift();q=1,a?a.t?m(function(){("c"==a.t?B.injectCss:B.injectJs)(a.s,0,a.a,a.x,a.e,1)},0):(a(),h()):q=0}function i(a,c,d,e,f,i,j){function k(b){if(!o&&g(l.readyState)&&(u.r=o=1,!q&&h(),l.onload=l.onreadystatechange=null,b)){"img"!=a&&m(function(){t.removeChild(l)},50);for(var d in y[c])y[c].hasOwnProperty(d)&&y[c][d].onload()}}var j=j||B.errorTimeout,l=b.createElement(a),o=0,r=0,u={t:d,s:c,e:f,a:i,x:j};1===y[c]&&(r=1,y[c]=[]),"object"==a?l.data=c:(l.src=c,l.type=a),l.width=l.height="0",l.onerror=l.onload=l.onreadystatechange=function(){k.call(this,r)},p.splice(e,0,u),"img"!=a&&(r||2===y[c]?(t.insertBefore(l,s?null:n),m(k,j)):y[c].push(l))}function j(a,b,c,d,f){return q=0,b=b||"j",e(a)?i("c"==b?v:u,a,b,this.i++,c,d,f):(p.splice(this.i++,0,a),1==p.length&&h()),this}function k(){var a=B;return a.loader={load:j,i:0},a}var l=b.documentElement,m=a.setTimeout,n=b.getElementsByTagName("script")[0],o={}.toString,p=[],q=0,r="MozAppearance"in l.style,s=r&&!!b.createRange().compareNode,t=s?l:n.parentNode,l=a.opera&&"[object Opera]"==o.call(a.opera),l=!!b.attachEvent&&!l,u=r?"object":l?"script":"img",v=l?"script":u,w=Array.isArray||function(a){return"[object Array]"==o.call(a)},x=[],y={},z={timeout:function(a,b){return b.length&&(a.timeout=b[0]),a}},A,B;B=function(a){function b(a){var a=a.split("!"),b=x.length,c=a.pop(),d=a.length,c={url:c,origUrl:c,prefixes:a},e,f,g;for(f=0;f<d;f++)g=a[f].split("="),(e=z[g.shift()])&&(c=e(c,g));for(f=0;f<b;f++)c=x[f](c);return c}function g(a,e,f,g,h){var i=b(a),j=i.autoCallback;i.url.split(".").pop().split("?").shift(),i.bypass||(e&&(e=d(e)?e:e[a]||e[g]||e[a.split("/").pop().split("?")[0]]),i.instead?i.instead(a,e,f,g,h):(y[i.url]?i.noexec=!0:y[i.url]=1,f.load(i.url,i.forceCSS||!i.forceJS&&"css"==i.url.split(".").pop().split("?").shift()?"c":c,i.noexec,i.attrs,i.timeout),(d(e)||d(j))&&f.load(function(){k(),e&&e(i.origUrl,h,g),j&&j(i.origUrl,h,g),y[i.url]=2})))}function h(a,b){function c(a,c){if(a){if(e(a))c||(j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}),g(a,j,b,0,h);else if(Object(a)===a)for(n in m=function(){var b=0,c;for(c in a)a.hasOwnProperty(c)&&b++;return b}(),a)a.hasOwnProperty(n)&&(!c&&!--m&&(d(j)?j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}:j[n]=function(a){return function(){var b=[].slice.call(arguments);a&&a.apply(this,b),l()}}(k[n])),g(a[n],j,b,n,h))}else!c&&l()}var h=!!a.test,i=a.load||a.both,j=a.callback||f,k=j,l=a.complete||f,m,n;c(h?a.yep:a.nope,!!i),i&&c(i)}var i,j,l=this.yepnope.loader;if(e(a))g(a,0,l,0);else if(w(a))for(i=0;i (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0];var j=d.createElement(s);var dl=l!='dataLayer'?'&l='+l:'';j.src='//www.googletagmanager.com/gtm.js?id='+i+dl;j.type='text/javascript';j.async=true;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-M677548'); Skip to main content Home About Submit ALERTS / RSS Search for this keyword Advanced Search New Results Developmental rewiring of the NGAL / CUC / KLU network associated with pleiotropic roles of NGAL genes View ORCID Profile Antoine Nicolas , Panagiotis Papadopoulos , View ORCID Profile Mattéo Caroulle , View ORCID Profile Bernard Adroher , Magali Goussot , View ORCID Profile Anne-Sophie Sarthou , View ORCID Profile Nicolas Arnaud , View ORCID Profile Aude Maugarny , View ORCID Profile Patrick Laufs doi: https://doi.org/10.1101/2024.12.12.627899 Antoine Nicolas 1 Université Paris-Saclay, INRAE, AgroParisTech, Institute Jean-Pierre Bourgin for Plant Sciences (IJPB) , 78000, Versailles, France 2 Université Paris-Saclay , 91405 Orsay, France Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Antoine Nicolas Panagiotis Papadopoulos 1 Université Paris-Saclay, INRAE, AgroParisTech, Institute Jean-Pierre Bourgin for Plant Sciences (IJPB) , 78000, Versailles, France Find this author on Google Scholar Find this author on PubMed Search for this author on this site Mattéo Caroulle 1 Université Paris-Saclay, INRAE, AgroParisTech, Institute Jean-Pierre Bourgin for Plant Sciences (IJPB) , 78000, Versailles, France Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Mattéo Caroulle Bernard Adroher 1 Université Paris-Saclay, INRAE, AgroParisTech, Institute Jean-Pierre Bourgin for Plant Sciences (IJPB) , 78000, Versailles, France Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Bernard Adroher Magali Goussot 1 Université Paris-Saclay, INRAE, AgroParisTech, Institute Jean-Pierre Bourgin for Plant Sciences (IJPB) , 78000, Versailles, France Find this author on Google Scholar Find this author on PubMed Search for this author on this site Anne-Sophie Sarthou 1 Université Paris-Saclay, INRAE, AgroParisTech, Institute Jean-Pierre Bourgin for Plant Sciences (IJPB) , 78000, Versailles, France Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Anne-Sophie Sarthou Nicolas Arnaud 1 Université Paris-Saclay, INRAE, AgroParisTech, Institute Jean-Pierre Bourgin for Plant Sciences (IJPB) , 78000, Versailles, France Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Nicolas Arnaud Aude Maugarny 1 Université Paris-Saclay, INRAE, AgroParisTech, Institute Jean-Pierre Bourgin for Plant Sciences (IJPB) , 78000, Versailles, France 2 Université Paris-Saclay , 91405 Orsay, France Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Aude Maugarny Patrick Laufs 1 Université Paris-Saclay, INRAE, AgroParisTech, Institute Jean-Pierre Bourgin for Plant Sciences (IJPB) , 78000, Versailles, France Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Patrick Laufs For correspondence: patrick.laufs{at}inrae.fr Abstract Full Text Info/History Metrics Supplementary material Preview PDF Abstract Gene Regulatory Networks (GRNs) play prominent roles in regulating developmental processes, and their modulation across species is a major source for evolutionary innovation. However, it remains poorly understood how GRNs are rewired between different organs within a single species. This question is particularly relevant for pleiotropic genes, which may exhibit organ-specific GRN modulations potentially reflecting their diverse functions. To address this, we investigated the NGATHA-like ( NGAL ) genes, as a model for pleiotropic genes that regulate growth or patterning in multiple Arabidopsis organs via two distinct pathways involving the CUP-SHAPED COTYLEDON ( CUC ) and KLUH ( KLU ) genes. By combining genetic analysis with gene expression characterization, we uncovered significant organ-specific rewiring of the NGAL / CUC / KLU regulatory module. Our findings highlight that changes in gene expression patterns, potentially arising from developmental constraints, play a pivotal role in the organ-specific modulation of GRNs. Furthermore, GRNs at the molecular and functional levels do not always align perfectly, potentially due to the influence of additional regulatory mechanisms. Altogether, our findings reveal significant modulation of the GRNs associated with pleiotropic genes. We propose that this flexibility in GRNs facilitates gene pleiotropy. Introduction A concept that has emerged during the two last decades is that biological processes are governed by gene regulatory networks (GRNs) ( Benfey and Weigel 2001 ; Davidson et al. 2003 ). While examining a GRN in a specific species or organ provides valuable insights into how it controls development and physiology, comparing GRNs across different species offers diverse insights into evolutionary processes. Thus, investigating the conservation of GRNs across species is a way to explore homology between structures in distantly related species, while GRN modulations can shed light on how innovations may have occurred during evolution ( Doebley and Lukens 1998 ; Hinman et al. 2003 ; Scotland 2010 ; Lynch and Roth 2011 ; Peter and Davidson 2011 ; Das Gupta and Tsiantis 2018; Cary et al. 2020 ; Srivastava 2021 ; Swift et al. 2024 ). In parallel, the more recent integration of single-cell transcriptomic data with other genome-wide omics has started to provide an unprecedent view of the GRN diversity at the cellular and tissular level ( Qian and Huang 2020 ; Longo et al. 2021 ; Badia-i-Mompel et al. 2023 ). However, understanding the functional significance of such GRN variations remains challenging. Here, we genetically addressed the question of inter-organ modulation using as a model a GRN that regulates multiple aspects of aerial organ development in Arabidopsis. A major mechanism driving the evolution of GRN structure and/or function is gene co-option ( Halfon 2017 ; Jiggins et al. 2017 ; Das Gupta and Tsiantis 2018). Gene co-option occurs when changes in the cis -regulatory sequences of a gene cause it to become active in a novel developmental context where it may acquire a novel function. Because genes do not function in isolation, a resulting question is to which extent the GRN associated with the co-opted gene is remodeled in the new biological environment ( Tyler et al. 2009 ; McQueen and Rebeiz 2020). Indeed, two opposite evolutionary patterns of GRN rearrangements have been observed. First, GRNs can keep their overall topology despite being redeployed in distantly related organisms. For instance, a similar GRN is promoting suberin deposition in the Arabidopsis endodermis and the tomato exodermis, two cell layers that share similar biological roles despite different positions in the root ( Cantó-Pastor et al. 2024 ). Redeployment of a conserved GRN can be also associated with different functions, as for instance a conserved GRN that controls eye development in the fly and cell migration in the sea urchin ( Martik and McClay 2015 ). Second, morphological innovations during evolution can be driven by co-opted GRNs that are partially modified such as those for the formation of trichomes or related unicellular projections in different organs or species of Drosophila ( Kittelmann et al. 2018 ; Rice et al. 2024 ), or that are more profoundly rewired, as shown for the development of simple and compound leaves ( Rast-Somssich et al. 2015 ) Another way to address GRN remodeling is to compare GRNs between different organs within a species. This is particularly relevant for GRNs centered around pleiotropic genes. Pleiotropy is defined as a phenomenon where a single gene or locus influences two or more distinct phenotypic traits ( Stearns 2010 ; Paaby and Rockman 2013 ). Remodeling of some GRNs between organs has been suggested in plants. For instance, the WUSCHEL/CLAVATA (WUS/CLV) network shows a regulation by AGAMOUS which is specific to the floral meristem and absent from the shoot apical meristem ( Lenhard et al. 2001 ). One can however argue that in this particular example the WUS/CLV function is not truly pleiotropic between floral and apical meristems, as in both organs it regulates stem cell homeostasis ( Schoof et al. 2000 ; Lenhard et al. 2001 ). The AINTEGUMENTA-LIKE/PLETHORA genes provide another example of organ-dependent rewiring of the GRN as these genes activate cell wall remodeling genes in the shoot while repressing them in the root ( Scheres and Krizek 2018 ), but yet again the pleiotropy of these genes could be discussed as they promote growth in both organs. Here, we investigate the remodeling of a small GRN centered on the pleiotropic NGATHA-like (NGAL) genes in different Arabidopsis organs. The NGATHA-like , NGATHA-like1 ( NGAL1 )/ ABNORMAL SHOOT 2 ( ABS2 ), NGAL2/SUPPRESSOR OF DA1 7 ( SOD7 ) and NGAL3/DEVELOPMENT-RELATED PCG TARGET IN THE APEX 4 ( DPA4 ) genes encode transcription factors of the B3 family ( Swaminathan et al. 2008 ; Romanel et al. 2009 ). Analysis of ngal simple and double mutants have revealed pleiotropic roles for these genes during the development of the aerial organs ( Engelhorn et al. 2012 ; Shao et al. 2012 , 2020 ; Chen et al. 2019 ; Nicolas et al. 2022 ; Zheng et al. 2023 ). For instance, DPA4 and SOD7 redundantly limit growth, particularly in seed, cotyledon and floral organs as these organs are larger in the dpa4 sod7 double mutant ( Zhang et al. 2015 ; Chen et al. 2019 ; Zheng et al. 2023 ). They also contribute to the establishment of phyllotaxis and to the development of axillary meristems. The dpa4 sod7 double mutant shows an altered phyllotaxis with the appearance of flower clusters on the main shoot and delayed axillary meristem initiation and outgrowth ( Chen et al. 2019 ; Nicolas et al. 2022 ). Finally, DPA4 , SOD7 and ABS2 limit the extent of leaf margin serration ( Shao et al. 2020 ). The pleiotropic role of NGAL genes during development has been associated with the function of two genetic pathways. First, there is strong evidence for genetic and molecular interactions linking NGAL and CUP-SHAPED COTYLEDON ( CUC ) genes, which code for NAC-family transcription factors ( Aida et al. 1997 ; Takada et al. 2001 ; Vroemen et al. 2003 ). Thus, interactions between NGAL and CUC genes shape leaf margins. The dpa4-1 sod7-1 abs2-2 triple mutant shows increased leaf margin serration compared to a wild-type plant, thus resembling plants with increased CUC activity ( Nikovics et al. 2006 ; Shao et al. 2020 ). On the contrary, lines overexpressing ABS2 / NGAL1 or DPA4 / NGAL3 have reduced leaf serration, similar to cuc2 or cuc3 mutants ( Nikovics et al. 2006 ; Engelhorn et al. 2012 ; Shao et al. 2012 , 2020 ). A similar interaction between NGAL and CUC genes occurs during axillary meristem development. CUC genes are required for axillary meristem initiation, likely by preventing cell differentiation and maintaining cells in a meristematic fate ( Raman et al. 2008 ). However, CUC genes need to be repressed by DPA4 and SOD7 for the initiating meristem to proceed to the establishment phase and allow stem cell formation and proper expression of WUS and CLV3 ( Nicolas et al. 2022 ). Several lines of evidence suggest the control of leaf and axillary meristem development by the NGAL genes mostly occurs via their repression of the CUC genes. First, NGAL proteins can bind to CUC promoters, and in ngal mutants the expression of the CUC genes is increased and/or ectopic. Second, transcriptomic data show that NGAL-mediated gene expression regulation is largely dependent on CUCs ( Shao et al. 2020 ). Third, leaf and axillary meristem defects of ngal mutants are mostly suppressed when CUC genes are additionally mutated ( Shao et al. 2020 ; Nicolas et al. 2022 ). The second pathway acting downstream the NGAL genes involves the KLUH ( KLU ) gene which codes for a cytochrome P450 of the CYP78A subfamily ( Anastasiou et al. 2007 ). KLU promotes growth in a non-cell-autonomous manner, possibly through a yet-to-identify mobile signal that maintains cell proliferation ( Anastasiou et al. 2007 ; Adamski et al. 2009 ; Eriksson et al. 2010 ; Kazama et al. 2010 ). Genetic analyses show that DPA4 and SOD7 act in a common pathway with KLU to regulate seed size ( Zhang et al. 2015 ). Thus, a lack of KLU repression by the DPA4 and SOD7 proteins lead to larger seeds as observed in the dpa4-2 sod7-3 mutant. In addition, SOD7 directly binds to the KLU promoter to repress its expression ( Zhang et al. 2015 ). Taken together, these data are demonstrating a strong genetic link between both CUC and NGAL genes and KLU and NGAL genes. Recent data also suggest a genetic link between the CUC and KLU genes. During embryo development and leaf margin morphogenesis, expression patterns of the CUC and KLU genes overlap ( Maugarny-Calès et al. 2019 ; Aida et al. 2020 ). Furthermore, KLU expression is absent at these two stages in cuc mutants, while its expression is induced following CUC activation. This suggests that KLU is acting downstream of the CUC genes. Interestingly, while expressing KLU under the control of CUC regulatory sequences is not sufficient to restore embryo defects of cuc mutants, it is to promote leaf serration ( Maugarny-Calès et al. 2019 ; Aida et al. 2020 ). This indicates that KLU has a more prominent role in the network acting downstream of CUC genes during leaf development than it has during embryo development. These elements indicate that the NGAL genes are important factors controlling plant development in a pleiotropic way and that this involves two possible pathways, involving either CUCs or KLU genes. However, we lack a systematic analysis of the contribution of these two pathways in mediating the function of the NGAL transcription factors in different organs, which could reveal possible organ-specific modulation of the NGAL GRN. That such modulation indeed exists is suggested by the observation that NGAL genes, despite being expressed during both embryonic and post-embryonnic meristem formation, are only required for the latter one ( Nicolas and Laufs 2022 ; Nicolas et al. 2022 ). Therefore, we explored here whether the NGAL / KLU / CUC module was modified between different organs. We first show that within the three NGAL genes, DPA4 and SOD7 have the most prominent role and redundantly control several aspects of aerial development, with, however, small variations in the contributions of the different genes. Next using a genetic approach, we reveal inter-organ modulation of the NGAL / KLU / CUC module. Hence while NGALs repress floral organ growth mostly via the KLU pathway, their repressive effect on cauline leaf growth occurs through both KLU and CUC pathways. Distinct from these, NGAL controls sepal numbers and phyllotaxis via CUC genes and mostly independently of KLU . Finally, NGAL and CUC independently repress seed growth. We further show that such network modulation can be due to different expression patterns of the acting genes. We also show that genetic regulations between components of the network are not necessarily translated into a functional effect, showing that molecular and functional networks may differ in an organ-specific way. Altogether, we provide evidence for the rewiring of the GRNs between different plant organs, and we suggest that such a flexibility of the GRN facilitates gene pleiotropy. Results DPA4 and SOD7 are the two major NGAL genes redundantly controlling plant development Previous reports indicate that dpa4 sod7 double mutants ( ds ) are affected in multiple aspects of aerial organ development as they show larger cotyledons, flowers and seeds, exhibit increased leaf serration and have phyllotaxy and branching defects ( Engelhorn et al. 2012 ; Shao et al. 2012 , 2020 ; Chen et al. 2019 ; Nicolas et al. 2022 ; Zheng et al. 2023 ). However, apart from shoot branching ( Nicolas et al. 2022 ), the contribution of individual NGAL genes to these phenotypes and their genetic redundancy was not systematically addressed. Therefore, we quantified floral organ numbers, the size of cauline leaves, petals, pistils and seeds and the phyllotaxis in the three single and three double ngal mutants ( Figure 1 , Figure S1). All these parameters were significantly different in ds compared to the wild type (WT) ( Figure 1 , Figure S1). Cauline leaves, petals, pistils and seeds were larger in ds compared to WT ( Figure 1A, G, I ). More flowers were located in clusters along the stem ( Figure 1C ). The ds flowers had a higher average number of sepals, petals, and carpels, but a lower average number of stamens ( Figure 1E , Figure S1A-H). In contrast, single mutants and the two other double mutant combinations ( abs2-1 dpa4-2 and abs2-1 sod7-2 ) showed only limited phenotypic abnormalities. Only the mean number of sepals was significantly increased in the abs2-1 sod7-2 double ( Figure 1E ). However, the number of sepals, petals and/or stamens were different from those in the WT in about 10% to 30% of ds flowers (Figure S1E-H), suggesting NGAL genes contribute to floral phenotype robustness. In addition, cauline leaves were slightly larger in dpa4-2 and abs2-1 sod7-2, as were pistils in abs2-1 dpa4-2 and abs2-1 sod7-2 ( Figure 1A ). In contrast, petals were smaller in abs2-1 (Figure S1I). Together, these observations indicated that each NGAL gene has only a very limited unique role. In contrast, DPA4 and SOD7 are mostly redundant in their roles to pleiotropically control plant development. This suggests that these two genes may have undergone limited subfunctionalization and neofunctionalization since their recent formation by duplication around 24 to 40 million years ago ( Romanel et al. 2009 ). Next, to genetically determine the contributions of the KLU and CUC pathways to the pleiotropic roles of DPA4 and SOD7 , we analyzed the phenotype of ds mutants in which KLU or CUC genes were inactivated. Download figure Open in new tab Figure 1. DPA4 and SOD7 have redundant, pleiotropic roles during plant development. A. Cauline leaf length in WT, single and double ngal mutants. B. Mature wild-type (WT) and dpa4-2 sod7-2 double mutant ( ds ) plants. C. Percentage of flowers grouped in clusters in WT, single and double ngal mutants. D. Part of the main inflorescence stem of wild-type (WT) and dpa4-2 sod7-2 double mutant ( ds ) plants. E. Sepal number in WT, single and double ngal mutants. F. Top view of mature wild-type (WT) and dpa4-2 and sod7-2 double mutant ( ds ) flower. G. Ovary width in WT, single and double ngal mutants. H. Scanning electron microscopy detail of wild-type (WT) and dpa4-2 sod7-2 double mutant ( ds ) pistil top part. I. Seed size in WT, single and double ngal mutants. J. Scanning electron microscopy view of wild-type (WT) and dpa4-2 sod7-2 double mutant ( ds ) seeds. In A , G and I data were normalized to the mean value of the WT. Rectangle represent the first and third quartiles, with a horizontal central line marking the median. Mean values are represented as red diamonds. In A , E and G, a Kruskal-Wallis test followed by a Dunn’s post-hoc test were performed to show significant differences between different mutants and in C and I, ANOVA followed by a Tukey post-hoc test were performed (p-value<0.05). Scale bars = 10 cm in A, 1 cm in D, 1 mm in F, 100 µm in H and J. DPA4 and SOD7 redundantly repress floral organ growth via KLU, independently of CUC genes The increase in petal and pistil size observed in the ds mutants was not affected by additional mutation of any of the CUC genes ( Figure 2A, B , Figure S2A, B). Furthermore, petals and pistils sizes were not modified in cuc single mutants, with the exception of a reduced size of cuc2-1 petals revealing a role of CUC2 in petal growth. In contrast, klu-4 mutants developed smaller petals ( Figure 2A, C , Anastasiou et al. 2007 ; Zhang et al. 2015 ) and pistils (Figure S2A, C). In dpa4-2 sod7-2 klu-4 triple mutants, petal size was fully restored ( Figure 2A, C , Zhang et al. 2015 ), while pistil size was only partially restored (Figure S2A, C). Thus, DPA4 and SOD7 repress floral organ growth via KLU independently of CUC genes ( Figure 6A ). However, the observation that the dpa4-2 sod7-2 klu-4 triple mutants has petals and pistils larger than klu-4 , suggest that DPA4 and SOD7 also repress growth via another pathway independent of KLU . Additionally, the partial restoration of pistil size in dpa4-2 sod7-2 klu-4 triple mutants also supports this hypothesis ( Figure 6A ). Download figure Open in new tab Figure 2. DPA4 / SOD7 redundantly repress petal growth via KLU and cauline leaf growth via KLU and CUC genes. A. Top view of mature wild type (WT), dpa4-2 sod7-2 ( ds ), cuc3-105 ( c3 ), dpa4-2 sod7-2 cuc3-105 ( dsc3 ), klu-4 ( k ) and dpa4-2 sod7-2 klu-4 ( dsk ) flower. B. Petal length in WT, single cuc , dpa4-2 sod7-2 double and dpa4-2 sod7-2 cuc triple mutants. C. Petal length in WT, single klu-4 , dpa4-2 sod7-2 double and dpa4-2 sod7-2 klu-4 triple mutants. D. Cauline leaf length in WT, single cuc , dpa4-2 sod7-2 double and dpa4-2 sod7-2 cuc triple mutants. E. Cauline leaf length in WT, single klu-4 , dpa4-2 sod7-2 double and dpa4-2 sod7-2 klu-4 triple mutants. F. Expression pattern in young developing petals of pDPA4:GFP ( DPA4 ), pSOD7:GFP ( SOD7 ), pCUC2:RFP ( CUC2 ), pCUC3:CFP ( CUC3 ) and pKLU:GFP ( KLU ) reporters. G. Detail of F , showing the expression of the reporters in the lower part of the petals. H. Expression pattern in young developing cauline leaves of pDPA4:GFP ( DPA4 ), pSOD7:GFP ( SOD7 ), pCUC2:RFP ( CUC2 ), pCUC3:CFP ( CUC3 ) and pKLU:GFP ( KLU ) reporters. For F to H , all reporters are represented in green regardless of the fluorescent reporter used. In F and H , chlorophyll fluorescence is shown in red. Note that the pictures of pCUC2:RFP ( CUC2 ), pCUC3:CFP ( CUC3 ) are from a same sample expressing both reporters simultaneously. In H , asterisks mark axillary meristems. In B to E , data were normalized to the mean value of the WT. Rectangle represent the first and third quartiles, with a horizontal central line marking the median. Mean values are represented as red diamonds. In B-D, a Kruskal-Wallis test followed by a Dunn’s post-hoc test were performed to show significant differences between different mutants (p- value<0.05). Scale bars = 100 µm in A, F-H DPA4 and SOD7 redundantly repress cauline leaf growth via KLU and CUC genes Cauline leaves of the ds mutants were larger than WT, and this phenotype was partially suppressed by cuc2-1 or cuc3-105 mutations and was totally suppressed by the klu-4 mutation ( Figure 2D, E ). Mutation of CUC1 had no effect on cauline leaf size in agreement with the absence of expression of this gene in developing leaves ( Hasson et al. 2011 ). These observations indicated that both KLU and CUC2 / CUC3 pathways are required for the increased cauline leaf size of the ds mutant, though with CUC2 / CUC3 having a less important contribution than KLU ( Figure 6B ). Since CUC2 promotes KLU expression during leaf development ( Maugarny-Calès et al. 2019 ), CUC2 / CUC3 may promote cauline leaf growth via KLU activation. Rewiring of the NGAL/CUC/KLU network between cauline leaves and petals is associated with modified gene expression patterns Our genetic analyses suggested that the networks controlling floral organ and cauline leaf growth shared a common DPA4/SOD7 – KLU edge but differed by a DPA4/SOD7 – CUC2/3 edge which is specific to cauline leaves ( Figure 6A,B ). To confirm this, we examined the expression of these genes during petal and cauline leaf development using transcriptional reporters ( Figure 2F-H ). DPA4 and SOD7 transcriptional reporters are strongly expressed in the region where the petals are attached to the flower receptacle and a weaker expression was observed in the proximal base of the petal. In contrast, CUC2 and CUC3 reporters where only expressed in the region where the petals are attached to the flower. The KLU transcriptional reporter is expressed along the petal margin and in the basal part of the petal ( Figure 2F-G ). Therefore, DPA4 , SOD7 and KLU , but not CUC2 and CUC3 have overlapping expression patterns in the basal part of the petal, a region in which KLU activity has been proposed to regulate organ growth ( Kazama et al. 2010 ). In developing leaves, DPA4 , SOD7 , KLU , CUC2 and CUC3 transcriptional reporters showed overlapping expression pattern along the proximal part of the leaf margin, the expression of the reporters being absent from the developing teeth ( Figure 2H ), in agreement with the proposed interactions of these genes in cauline leaves ( Figure 6B ). Therefore, the difference between the GRN regulating cauline leaf and petal growth ( Figure 6A,B ) may result from the CUC genes not being expressed in the latter organ. As strong epidermal auxin response foci have been associated with teeth formation and CUC gene expression at the leaf margin ( Hay et al. 2006 ; Kawamura et al. 2010 ; Bilsborough et al. 2011 ) we next compared DR5:VENUS reporter expression in petal and cauline leaves. This revealed the presence of epidermal foci with strong auxin response present at the margin of cauline leaves which were absent in petals (Figure S2D). Thus, different auxin response pattern may contribute to differential CUC expression, which in turn leads to variations in GRNs between petals and cauline leaves. Together, these data indicate that rewiring of the GRN may be the consequence of different expression patterns of the GRN actors. DPA4 and SOD7 control phyllotaxy via CUC gene repression independently of KLU To quantify more precisely the phyllotaxy defects observed in ds ( Figure 1C, D ), we analyzed the distribution of the internode lengths along the main stem. In the ds mutant the presence of numerous flower clusters resulting from short internodes separated by long internodes translated in a broader distribution of internode length and a reduction in the median internode length compared to WT ( Figure 3A, C ). Inactivating individual CUC genes in ds led to a partial restoration towards a WT phenotype with fewer visible flower clusters ( Figure 3C ). Quantification confirmed this, as a more compact distribution of the internode length and an increase in the median internode length was observed, with CUC3 inactivation leading to the best phenotypic restoration ( Figure 3A ). We could not analyze the phyllotaxy of ds with two simultaneous cuc mutations because these quadruple mutants showed fusion of the floral pedicel with the main stem. On the other hand, introducing a klu-4 mutation in ds increased its phyllotaxy defects ( Figure 3B,C ). These observations indicated that the DPA4 / SOD7 pathway controls phyllotaxy to a large extent via the CUC genes while this pathway is independent of the KLU gene ( Figure 6D ). Phyllotaxy defects of the ds mutant were already present early in development, as clusters of flowers were visible at the ds inflorescence apex, while in the WT flowers were inserted in a more regular manner ( Figure 3D ). CUC2 and CUC3 transcriptional reporters were ectopically expressed in the young ds inflorescence stem, either in large domains or small groups of cells ( Figure 3E ). Together, these observations indicate that DPA4 / SOD7 are required to establish a proper phyllotaxy by repressing ectopic CUC expression in developing stem, a similar role as the one proposed for microRNA miR164 that represses CUC2 ( Peaucelle et al. 2007 ; Sieber et al. 2007 ). Download figure Open in new tab Figure 3. DPA4 / SOD7 control phyllotaxis through the CUC genes. A. Internode length distribution in WT, single cuc , dpa4-2 sod7-2 double and dpa4-2 sod7-2 cuc triple mutants. The black horizontal segment represents the median. B. Internode length distribution in WT, single klu-4 , dpa4-2 sod7-2 double and dpa4-2 sod7-2 klu-4 triple mutants. The black horizontal segment represents the median. C. Part of the main inflorescence stem of wild-type (WT), dpa4-2 sod7-2 ( ds ), cuc1-13 ( c1 ), dpa4-2 sod7-2 cuc1-13 ( dsc1 ), cuc3-105 ( c3 ), dpa4-2 sod7-2 cuc3-105 ( dsc3 ), klu-4 ( k ) and dpa4-2 sod7-2 klu-4 ( dsk ) plants. D. Partially dissected inflorescence apex of wild type (WT), dpa4-2 sod7-2 ( ds ). The flowers were removed from the lower part (arrows). The squares are positioned on the type of zones which were imaged in E. E. Expression of pCUC2:RFP (in red) and pCUC3:CFP (in cyan) in young internodes of wild-type (WT) or dpa4-2 sod7-2 double mutant ( ds ) plants. In A,B , a two sample Kolmogorov-Smirnov test was performed to show significant differences between different mutants (p-value<0.01). Scale bars = 1 mm in D and 100 µm in E. DPA4 and SOD7 controls floral organ numbers via the CUC genes and independently of KLU We next investigated the contribution of the CUC and KLU genes to the increased sepal and petal numbers observed in ds ( Figure 1E, F , Figure S1A,B,E,F). The increase in sepal number in ds was not modified by any of the single mutation in the CUC genes, while mutating two CUC genes simultaneously significantly restored the phenotype ( Figure 4A, B , Figure S3D,E). Notably the dpa4-2 sod7-2 cuc1-13 cuc3-105 quadruple mutant had always 4 sepals like the WT ( Figure 4B , Figure S3E). Download figure Open in new tab Figure 4. DPA4 / SOD7 control sepal patterning through the CUC genes and independently of KLU. A. Sepal number in WT, single cuc , dpa4-2 sod7-2 double and dpa4-2 sod7-2 cuc triple mutants. B. Sepal number in WT, cuc double, dpa4-2 sod7 double and dpa4-2 sod7-2 cuc quadruple mutants C. Sepal number in WT, single klu-4 , dpa4-2 sod7-2 double and dpa4-2 sod7-2 klu-4 triple mutants. D-P. Scanning electron microscopy view of inflorescence apices or young developing floral meristems of wild-type (WT), dpa4-2 sod7-2 ( ds ), dpa4-2 sod7-2 cuc1-13 ( dsc1 ), dpa4-2 sod7-2 cuc3-105 ( dsc3 ), klu-4 ( k ) or dpa4-2 sod7-2 2 cuc1-13 cuc3-105 ( dsc1c3 ) plants. Q. Frequency of the flowers with different sepal primordia numbers in different genotypes R,S. Expression pattern in young developing flowers of pCUC3:CFP (CUC3 in cyan) and DR5:VENUS (DR5 in yellow) reporters in wild type (WT) and dpa4-2 sod7-2 ( ds ). The floral stage is indicated (S2 to S5). The adaxial part of the floral meristem is always on the top of the image. In A - C, a Kruskal-Wallis test followed by a Dunn’s post-hoc test were performed to show significant differences between our different mutants (p- value<0.05). Scale bars = 100 µm in all panels. The interaction between DPA4 / SOD7 and CUC genes for the control of petal number was more complex (Figure S3A,B,G,H), with for instance a reduction of the petal number below 4 in all three dpa4-2 sod7-2 cuc triple mutants. However, like for the sepals, the dpa4-2 sod7-2 cuc1-13 cuc3-105 quadruple mutant showed an almost fully restored number of petals, with a mean petal number close to 4 and strongly reduced variability of the petal number (Figure S3B,H). Together, this suggested that the regulation of sepal and petal number by DPA4 and SOD7 occurs through a pathway involving CUC1 and CUC3 . Like ds, the klu-4 mutant showed an increase in sepal number, and both effects were additive as the sepal number further increased in the dpa4-2 sod7-2 klu-4 triple mutant compared to ds and klu-4 ( Figure 4C , Figure S3C,F,I). Petal number was also higher in dpa4-2 sod7-2 klu-4 compared to ds and klu-4. This suggested that DPA4 / SOD7 and KLU act in independent pathways during floral organ specification ( Figure 6D ). To further support the genetic interactions between the NGAL / CUC / KLU genes during flower development, we precisely characterized their expression using lines expressing simultaneously combinations of two transcriptional reporters of these genes (Figure S4). The five genes showed overall overlapping expression patterns in the boundary domain of inflorescence and floral meristems, with KLU being additionally expressed within the meristems. At floral stages S1 and S2 (floral stages defined according to Smyth et al. 1990 ), DPA4 and SOD7 reporters showed a strong expression in the adaxial domain, which was also observed for CUC3 and to a lower extent KLU but not CUC2 . In addition, DPA4 and CUC3 reporter expressions transiently extended into the entire floral meristem during S1. At S3, when the sepal-meristem boundary became marked by CUC2 and CUC3 reporter expressions, DPA4 and SOD7 reporter expression were not observed in the floral meristem. This domain was marked by DPA4 and SOD7 reporters at later stage S4, as were more inner boundary domains at S5. No expression was however observed for DPA4 and SOD7 in the intra whorl sepal-sepal boundary domain in contrast to CUC2 . The outer abaxial epidermis of the sepal also strongly expressed the SOD7 reporter. At S2 and S3, KLU was expressed throughout the floral meristem, with a stronger expression at its boundary with the inflorescence meristem at S2 and the sepal boundaries at S3. A strong expression of KLU remained in the center of the flower meristem at S4 and S5 and the sepal primordia maintained KLU expression at these stages. Together, these observations suggested that the genetic interaction between DPA4 / SOD7 and CUC2 / CUC3 genes occurs at the organ boundaries while the KLU effect on organ number independent of DPA4 / SOD7 may result from the unique expression of KLU at the meristem center. Interaction of DPA4 and SOD7 with CUC genes controls sepal initiation patterns To further relate the changes in sepal numbers in mature flowers with early flower development we examined it by SEM ( Figure 4D-P ). While in the WT the stereotypical pattern of 4 initiating sepal primordia was observed ( Figure 4D-F,Q ), ds flowers showed highly variable patterns ( Figure 4G-M,Q ). Thus, we observed flowers with 3 equidistant and similar sepal primordia ( Figure 4H,I ) or flowers with 4 sepal primordia of inequal sizes and irregular positions ( Figure 4J,K ). Young flower primordia in which initiation of the adaxial sepal primordium appeared delayed could be observed ( Figure 4L ). Older flowers with 5 sepal primordia, including two small ones in adaxial position that could result from delayed initiation were present in the ds mutant ( Figure 4M ). Inactivating CUC1 or CUC3 alone in the ds background only very partially corrected sepal initiation defects, while inactivating CUC1 and CUC3 together almost totally suppressed ds sepal initiation defects ( Figure 4N- Q ). To follow sepal initiation at the molecular level and relate it to CUC gene expression patterns, we imaged WT and ds young flowers expressing the auxin response marker pDR5:VENUS and pCUC3:CFP ( Figure 4R,S ). The pDR5:VENUS marker is expressed in the incipient organ primordia ( Chandler et al. 2011 ; Kong et al. 2024 ). From floral stage S3 onwards, WT floral meristem had 4 separated DR5 expressing foci of cells, that correspond to the 4 sepal primordia initiation sites. By contrast, at S3 and S4, only 3 foci were often observed in ds flowers, one in the abaxial position, and two at an intermediate position between the median and adaxial positions. At later stages, five pDR5:VENUS - expressing cell foci were often observed, in agreement with the most frequent sepal primordium number observed in the ds mutants. At S2 stage, pCUC3:CFP expression was weak in WT floral meristems, while it appeared higher in ds floral meristems, in particular in its adaxial domain. This is also the region in which pDPA4:GFP and pSOD7:GFP are strongly expressed (Figure S4). Together, this suggest a model in which in the ds floral meristems, CUC3 is transiently locally derepressed in the adaxial domain, which in turn interferes with auxin signaling and sepal primordium formation. As our genetic data indicate that CUC1 is required with CUC3 to cause abnormal sepal numbers in ds, we can speculate that CUC1 may be similarly derepressed in ds and that it may also contribute to delayed sepal initiation as recently shown in the drmy1 mutant ( Kong et al. 2024 ). DPA4/SOD7 and CUC genes independently repress seed growth Mutation of the KLU gene leads to smaller seeds ( Adamski et al. 2009 ) and totally suppresses the increase in seed size of ds double mutants ( Zhang et al. 2015 ). Mutation of single CUC genes did not modify seed size in the ds mutant, but we observed a limited increase of seed size in single cuc mutants, which was however only statistically significant for cuc3-105 ( Figure 5A ). This prompted us to analyze the effects of multiple CUC gene mutations on seed size to overcome possible genetic redundancy. Indeed, seeds of both cuc1-13 cuc3-105 and cuc2-1 cuc3-105 double mutant were larger than WT ( Figure 5B, C ): seeds of the third double mutant, cuc1 cuc2, could not be analyzed as this double mutant does not produce any flowers ( Aida et al. 1997 ). A further increase in seed size was observed in the ds background when two CUC genes were simultaneously inactivated ( Figure 5B,C ). These observations indicate that CUC genes redundantly repress seed growth and do so independently of DPA4 and SOD7 . Again, as during cauline leaf development, our genetic analysis did not allow us to determine whether CUC genes act by repressing KLU or via a pathway independent of KLU . However, because CUC genes have been shown to activate KLU expression in embryos and leaves ( Maugarny-Calès et al. 2019 ; Aida et al. 2020 ), we rather favor the hypothesis of an action of CUC genes on seed growth independent of KLU ( Figure 6D ). This model for the genetic interactions directing seed size regulation contrasts with the ones identified in other organs ( Figure 6A-C ), since here DPA4 / SOD7 appears to act independently but with similar effects as the CUC genes, while in other organs DPA4 / SOD7 and CUC genes act antagonistically. This prompted us to question whether DPA4 / SOD7 represses CUC genes during seed development as it does in other organs ( Figure 3E , Engelhorn et al. 2012 ; Shao et al. 2020 ; Nicolas et al. 2022 ). To test this, we first questioned whether the expression of these genes overlap and, because DPA4 / SOD7 control seed size maternally by affecting ovule growth ( Zhang et al. 2015 ), we analyzed their expression patterns during ovule development. pDPA4:GFP is strongly expressed in the outer cells of the basal part of the nucellus and weakly in the inner integument primordia in stage 2-III ovules (stages according to Schneitz et al., 1995 ) (Figure S5A). At later stages, pDPA4:GFP expression remained high at the base of the inner integuments but extended into the inner integuments and became expressed at the base of the outer integuments too (Figure S5B-D). The pSOD7:GFP reporter showed an expression pattern very close to the one provided by the pDPA4:GFP reporter (Figure S5E-H) ( Zhang et al. 2015 ). pKLU:GFP reporter was expressed in domains that were also very similar to the ones of pDPA4:GFP and pSOD7:GFP (Figure S5M-P) ( Adamski et al. 2009 ). In contrast, pCUC2:RFP and pCUC3:CFP showed very different patterns (Figure S5I-L). At stage 2-III, pCUC3:CFP was not detectable and pCUC2:RFP was limited to a few cells at the very base of the nucellus (Figure S5I). At stage 3-I and later, pCUC3:CFP formed a ring-link expression pattern at the junction of the nucellus and the inner integument, while pCUC2:RFP was expressed below (Figure S5J-L). Therefore, despite having different patterns, the pCUC2:RFP and pCUC3:CFP domains were included in the domains expressing the pDPA4:GFP and pSOD7:GFP reporters, raising the possibility that DPA4 / SOD7 could regulate CUC gene expression during ovule development. To test this, we analyzed pCUC2:RFP and pCUC3:CFP expression during ds and WT ovule development. At stages when integuments were initiated, expression of pCUC2:RFP was extended in the nucellus of ds ovules compared to WT ( Figure 5D, E ). At later stages, a higher and enlarged pCUC2:RFP expression was observed in ds ovules ( Figure 5F, G ). The expression of pCUC3:CFP was not modified in ds ovules ( Figure 5D-G ). Together, these data suggested that DPA4 / SOD7 repressed CUC2 expression during ovule development, but that derepression of CUC2 in ds was not causal for the increase in seed size of the double mutant. Download figure Open in new tab Figure 5. DPA4 / SOD7 and CUC genes additively repress seed growth A. Seed size in WT, single cuc , dpa4-2 sod7-2 double and dpa4-2 sod7-2 cuc triple mutants. B. Seed size in WT, cuc double, dpa4-2 sod7 double and dpa4-2 sod7-2 cuc quadruple mutants C. Scanning electron microscopy view of wild-type (WT), cuc1-13 cuc3-105 ( c1c3 ), cuc2-1 cuc3-105 ( c2c3 ), dpa4-2 sod7-2 ( ds ), dpa4-2 sod7-2 cuc1-13 cuc3-105 ( dsc1c3 ) and dpa4-2 sod7-2 cuc2-1 cuc3-105 ( dsc2c3 ) seeds D. Expression of pCUC2:RFP (in red) and pCUC3:CFP (in cyan) in developing ovules at stage 2-IV ( Schneitz et al. 1995 ) of wild-type (WT) or dpa4-2 sod7-2 double mutant ( ds ) plants.ii: inner integument, io: outer integument, n: nucellus E. Quantification of nucellus cells expressing pCUC2:RFP. Ovules were at stages2-III or 2-IV. F. Expression of pCUC2:RFP (in red) and pCUC3:CFP (in cyan) in developing ovules at stage 3-III ( Schneitz et al. 1995 ) of wild-type (WT) or dpa4-2 sod7-2 double mutant ( ds ) G. Quantification of pCUC2:RFP and pCUC3:CFP expression levels in ovules at stages 3-III to 3-V. In A and B , data were normalized to the mean value of the WT. Rectangle represent the first and third quartiles, with a horizontal central line marking the median. Mean values are represented as red diamonds. In G , statistical significance is tested by Student’s tests: p-value is 0.001 for ****. In A and B, ANOVA followed by a Tukey post-hoc test was performed (p-value<0.05). In E, statistical significance is tested by Mann & Whitney tests: p-value is 0.001 for **** Scale bars = 100 µm in C, 10 µm in D and F. Discussion How gene regulatory networks (GRNs) centered around pleiotropic genes are remodeled remains a largely open question in developmental and evolutionary biology. In this study, we explored this issue by focusing on the NGAL family of transcription factors, which play pleiotropic roles in plant development. Hence, we determined the gene network linking NGALs with their downstream genes KLU and CUCs during the regulation of seven different biological processes (growth of cauline leaves, petals, pistils, seeds, initiation of sepals and petals, inflorescence phyllotaxis). We identified four distinct types of network organizations, each differing in how NGAL effects are mediated through either CUCs , KLU , or a combination of both ( Figure 6D ). The network regulating seed growth was notably distinct from the others. In this case, NGAL and CUC genes had additive effects on seed growth repression. In contrast, in the three other networks, CUC genes had either antagonistic roles to the NGAL genes or did not interact with the NGAL pathway. The network’s precise structure can be further refined due to genetic redundancy among the three CUC genes, with varying degrees of involvement for these different members. For instance, our data suggest that CUC2 and CUC3 had similar contribution to the NGAL -mediated regulation of cauline leaf growth while CUC1 had no apparent role in it. Importantly, the structure of the network was not conserved within a specific organ or in the regulation of the specific biological process across different organs. For instance, we observed different genetic networks governing petal initiation and their subsequent growth. Similarly, the structures of the network governing the growth of cauline leaves, petals or seeds were also different, illustrating inter-organ variation. Download figure Open in new tab Figure 6. Organ-wide rewiring of the NGAL / CUC / KLU regulatory network The GRNs were genetically determined by analyzing the phenotypes of single and multiple mutants. Gene expression patterns could be further used to support the GRNs. Repressing activities are shown by red lines ending by a point, while promoting activities are indicated by green arrows. Thickness of the red and green lines represent the importance of the regulatory step to the regulated biological process. A. The major pathway by which DPA4 / SOD7 repress petal and pistil growth acts through the repression of KLU pathway, which promotes growth of these organs. DPA4 / SOD7 also represses petal and pistil growth via a second, minor pathway independent of KLU . DPA4 / SOD7 act independently of the CUC genes. However, CUC2 has a minor role in the promotion of petal growth. B. DPA4 / SOD7 repress cauline leaf growth via two pathways. The major pathway mediating the growth-repressing effect of DPA4 / SOD7 involves the repression of the KLU pathway, which promotes growth. The second, more minor, pathway operates through CUC2 and CUC3 , which may act either via or independently of the KLU pathway. C. DPA4 / SOD7 determine the patterns of phyllotaxy and sepal and petal initiation via a pathway involving the CUC genes. Contribution of the individual CUC genes to this varies (not shown in the figure). For instance, CUC3 has an important contribution to phyllotaxy while CUC1 has a more minor role. In contrast, both CUC1 and CUC3 have an important contribution to sepal patterning. The KLU pathway has also a minor contribution in the determination of the sepal and petal initiation patterns and phyllotaxy, this role is however independent of DPA4 / SOD7 pathway. D. DPA4 / SOD7 repress seed growth via the KLU pathway ( Zhang et al. 2015 ). The CUC genes also repress seed growth. DPA4 / SOD7 -mediated seed growth repression is independent of the CUC genes, although DPA4 / SOD7 represses CUC2 expression during ovule development (not shown in the figure). To establish these gene networks, we used morphometric analyses across various genetic backgrounds, combined with gene expression characterization using imaging of reporter constructs. These methods are particularly well-suited for analyzing inter organ variations, as they allow linking functional roles with spatial resolution of gene expression domains. Alternative methods can be used to establish GRNs at the molecular level with different spatial resolutions ( O’Malley et al. 2016 ; Qian and Huang 2020 ; Tripathi and Wilkins 2021 ; Li et al. 2023 ; Yin et al. 2023 ; Yuan and Duren 2024 ). Combining spatial information provided by single cell analysis or other cell-type specific approaches helps resolving GRNs at the organ or cell type level ( Baumgart et al. ; Brady et al. 2011 ; Tian et al. 2014 ; de Luis Balaguer et al. 2017 ; Sonawane et al. 2017 ; Kajala et al. 2021 ; Robertson and Wilkins 2023 ). Such approaches have also been used to follow evolution of GRNs during cell differentiation or in response to pathogens ( Shahan et al. 2022 ; Kamimoto et al. 2023 ; Nobori et al. 2023 ; Zhang et al. 2023 ; Zhu et al. 2023 ) or to compare GRN evolution across organs or species ( Kajala et al. 2021 ; Guillotin et al. 2023 ). However, they do not necessarily establish the functional roles of different network elements. Indeed, our study provides an example where a regulatory node conserved at the molecular level is functionally relevant in some organs but not in all. For instance, NGAL repression of CUC2 contributes to the regulation of phyllotaxis, floral organ initiation, and cauline leaf growth, but not seed growth. More specifically, DPA4 / SOD7 repression of CUC2 transcription occurs during seed development but does not seem to contribute to larger ds seeds, as the dpa4 sod7 mutant does not show any restoration of seed size when CUC2 is mutated. These observations suggest that GRNs can be explored at both molecular and functional levels, and that these two levels may not always perfectly align. Several hypotheses may explain why a regulation occurring at the molecular level has no apparent functional effect as we found for CUC2 transcriptional repression by NGAL genes during ovule development. First, gene regulation occurs at multiple levels, and in the case of CUC2 , post-transcriptional regulation by the microRNA miR164 and protein-protein interactions are important factors regulating its activity ( Nikovics et al. 2006 ; Rubio-Somoza et al. 2014 ; Barro-Trastoy et al. 2022 ) and miR164 can even mask the effects of a transcriptional derepression ( Maugarny et al. 2024 ). A second, more specific hypothesis regarding CUC genes arises from their complex effects on growth. Indeed, CUC2 represses growth autonomously, in part by modifying the expression of cell wall modifying enzymes ( Peaucelle et al. 2007 ; Kawamura et al. 2010 ; Kierzkowski et al. 2019 ; Serra and Perrot-Rechenmann 2020 ; Bouré et al. 2022 ), while it promotes growth at a distance in part via the modulation of auxin response (Shankar et al.; Bilsborough et al. 2011 ; Maugarny-Calès et al. 2019 ; Hu et al. 2024 ). Hence, predicting the effects on growth of CUC modulation may be difficult, and for instance both larger or smaller leaves have been shown to result from CUC2 over expression, possibly as a result from different pattern and/or level of expression ( Nikovics et al. 2006 ; Larue et al. 2009 ; Li et al. 2020 ). Finally, and more specifically concerning seed size, both maternal and zygotic factors play a role in its regulation ( Li et al. 2024 ). Here we concentrated on the maternal control, as NGAL genes were shown to affect seed size through this pathway ( Zhang et al. 2015 ). However, the NGAL / KLU / CUC module which is also expressed during embryo development ( Aida et al. 1999 , 2020 ; Nicolas and Laufs 2022 ) may contribute to the regulation of seed size via the zygotic pathway, a hypothesis that remains to be tested. Our work identifies the modulation of gene expression patterns as an important mechanism leading to inter-organ changes in the NGAL / CUC / KLU GRN. This is supported by the comparison of growth regulation between petals and cauline leaves. Indeed, genetic analysis indicate that NGAL repression of growth is in part mediated by the CUC2 and CUC3 genes in cauline leaves, while it is independent from these genes in petals. We could relate this difference to the expression of the CUC2 and CUC3 genes at the cauline leaf margin where their expression overlaps with that of DPA4 and SOD7 , while the expression of the CUC genes is absent from the petal margin. Although leaves and petals are related organs, they also differ in their developmental and growth programs ( Pelaz et al. 2001 ; Sauret-Güeto et al. 2013 ; Le Gloanec et al. 2024 ). Indeed, we observed different patterns of auxin response distribution between petals and cauline leaves, which could contribute to the difference in CUC gene expression between these organs ( Hay et al. 2006 ; Kawamura et al. 2010 ; Bilsborough et al. 2011 ). Together, this suggests that developmental constraints, by influencing gene expression patterns, may contribute to inter-organ modulation of GRNs. However, variations in gene expression patterns can also influence GRNs through an alternative mechanism, as demonstrated in sepal initiation. In this second case, genetic and expression data converge to support a model in which sepal and petal primordia numbers are regulated by two different independent pathways: the first acting at the organ boundary involves DPA4 / SOD7 repressing CUC2 / CUC3 and the second acting more at the meristem center involves KLU . In this model, KLU activity at the meristem center would escape DPA4 / SOD7 regulation by being expressed in a domain where DPA4 / SOD7 expression is absent. These two examples illustrate that variations in the combinatorial expression of the GRN actors are a major driver of GRN rewiring. Here, we show that the pleiotropic effects of NGAL genes are linked to an organ-wide reconfiguration of their associated GRN, involving the KLU and CUC genes. The KLU and CUC pathways have distinct biological functions: KLU is primarily involved as a growth regulator ( Anastasiou et al. 2007 ; Adamski et al. 2009 ; Eriksson et al. 2010 ; Kazama et al. 2010 ; Chakrabarti et al. 2013 ; Zhang et al. 2015 ), whereas CUC genes not only influence growth but also play a crucial role in establishing developmental patterns ( Furutani et al. 2004 ; Bilsborough et al. 2011 ; Maugarny et al. 2016; Maugarny-Calès et al. 2019 ). Therefore, the rewiring of the GRN provides as a mechanism to balance the relative contributions of the KLU and CUC pathways across different organs, thereby determining the GRN’s impact on growth or patterning. In this view, GRN rewiring represents a means to shape diverse biological outcomes. More generally, we propose that organ-wide GRN plasticity facilitates the pleiotropic roles of genes. Materials and Methods Plant materials The ngal and cuc single and multiple mutants ( Nicolas et al. 2022 ) and klu-4 mutant ( Anastasiou et al. 2007 ) and the pKLU:GFP, pDR5:VENUS , ( Maugarny-Calès et al. 2019 ), pCUC2:RFP , pCUC3:CFP , pDPA4:GFP and pSOD7:GFP ( Nicolas et al. 2022 ) reporter lines have been described before. New combinations of mutants or mutants expressing reporter lines were generated by crossing. Lines homozygous for the desired mutation(s) were identified by PCR-based genotyping. The presence of the reporter line was verified through its expression and its homozygous state was determined by analyzing its segregation in the next generation. Only lines homozygous for the reporter(s) were analyzed except for the comparison between WT and dpa4-2 sod7-2 of pCUC2:RFP and pCUC3:CFP ( Figures 3E , 5D,F ) and the combinations of two reporters ( Figure 4R,S , Figure S4A-E, G,H), in which each reporter was at the heterozygous stage. Plant Growth and Phenotyping Plants were grown in the greenhouse in a mix peat/sand (TREF 1018201170, TREF, France). The length of the 3 first (most basal) cauline leaves was measured using a ruler on plants grown in the greenhouse. Floral organ numbers were determined under a binocular. Petal and pistils were dissected from mature flowers, mounted in water, imaged using a binocular and their size was measured using FIJI ( Schindelin et al. 2012 ). For internode length, fully grown inflorescences were taped on paper, scanned and the distances between successive nodes were measured using ImageJ. Flowers were defined as belonging to clusters when they were separated by internodes shorter than 3 mm. For seed size, seeds were sprayed on tape, imaged using a scanner and seed area was measured using a dedicated macro on ImageJ (Seed XXXXX). Images were threshold and the area of individual particles was measured. To remove possible non-seed particle, first size (> 0.05 mm 2 ) and circularity (0.75-1.00) filters were applied, the result of this filtering was then inspected manually. In particular, we removed data from particles which were fused, such fused seeds as described previously for cuc1 cuc3 and cuc2 cuc3 double mutants ( Gonçalves et al. 2015 ). Each point in the graph represents the mean area of at least 75 seeds of one plant. Data representation and statistical analysis All data were analysed using R ( R Core Team 2022 ). Boxplots (for cauline leaf, petal lengths, pistil width and seed size), were generated using the geom_boxplot function of the ggplot2 package (Wickham 2016). They compactly display the distribution of a continuous variable by allowing the visualization of the median (bold horizontal bar), the first and third quartile (respectively lower and upper hinges), the lowest and largest values no further than 1.5 * interquartile range from the hinge (respectively lower and upper whisker). In addition, all data are represented as individual points and the mean is represented. Values were normalized relative to the mean value of the WT. In bar plots (for floral organ numbers) error bars represent the standard errors. Violin plots (for internode length) were generated using the geom_violin function (ggplot2 package). Horizontal black bars represent the median of the values. Statistically significant differences were shown using an ANOVA followed by Tukey’s post-hoc test (p- value <0.05) ( Figure 1C, I ; 5A, B ), a nonparametric Kruskal-Wallis test followed by Dunn’s post-hoc test (p-value <0.05) ( Figure 1A, E, G ; S1A-D, I; 2B-E ; S2B, C; S3A-C; 4A-C ), a Student’s t test (p-value < 0.05) ( Figure 5G ), or using a non-parametric Mann & Whitney test (p-value < 0.05) ( Figure 5E ). The application conditions of ANOVA or Student’s t test were checked using the Shapiro-Wilk (distribution) and Levene (homogeneity of variances) tests respectively. Comparison of the internode length distributions ( Figure 4A, B ) were done by a two sample Kolmogorov-Smirnov test (p-value <0.01). The R packages car, dplyr, FSA and rcompanion were used for statistical analysis. Letters were used to highlight the significant phenotypic differences between different genotypes. Confocal imaging and quantification Confocal imaging was performed on a Leica SP5 inverted microscope (Leica Microsystems, Wetzlar, Germany). The lenses were Leica 20x or 40x HCX PL APO CS. Acquisition parameters are presented in Supplemental Table S1. Maximum intensity projections along the Z axis were made using ImageJ. Stitching of several cauline leaf pictures was performed using the Pairwise Stiching plugin ( Preibisch et al. 2009 ) and FigureJ ( Mutterer and Zinck 2013 ) was used to assemble the figures. For the comparison of pCUC2:RFP and pCUC3:CFP expression in ovules imaging was performed using the confocal same settings (laser power and signal gain) for WT and dpa4- 2 sod7-2 . For signal quantification, we first identified the median Z-section (with the two spots of pCUC3:CFP expression most distant), next we performed a max intensity projection along the Z axis including the 5 sections below and above the median section (Z step =2 µm). Fluorescent signal was then measured in the entire ovule (excluding the funiculus) and the background was subtracted from it. Scanning electron microscopy Freshly sampled tissues were cooled to −33C° by a Peltier cooling stage (Deben) and observed with a Hirox SH-1500 benchtop scanning electron microscope. Author contributions Conceptualization: AN, AM, PL Formal Analysis: AN, PP, MC Investigation: AN, PP, MC, BA, MG, NA, PL Writing – Original Draft: PL Writing – Review & Editing: AN, PP, MC, NA, AM, PL Visualization: AN, PL Supervision: PL, AM Supplemental Figure 1. Floral organ numbers and petal length in single and double ngal mutants A-D. Floral organ numbers in WT, single and double ngal mutants ( A , sepal; B , petal; C , stamen; D , carpel). in single and double ngal mutants E-H. Floral organ number frequency WT, in single and double ngal mutants ( E , sepal; F , petal; G , stamen; H , carpel). The color coding of the organ number is specific for each organ type and shown below the graphs. I. Petal length in WT, single and double ngal mutants. Data were normalized to the mean value of the WT. In A-D, a Kruskal-Wallis test followed by a Dunn’s post-hoc test were performed to show significant differences between different mutants (p-value<0.05). Supplemental Figure 2. DPA4 / SOD7 redundantly repress pistil growth via KLU and auxin response in petal and cauline leaf. A. Scanning electron microscopy detail of wild type (WT), dpa4-2 sod7-2 ( ds ), cuc3-105 ( c3 ), dpa4-2 sod7-2 cuc3-105 ( dsc3 ), klu-4 ( k ) and dpa4-2 sod7-2 klu-4 ( dsk ) pistil top part. B. Ovary width in WT, single cuc , dpa4-2 sod7-2 double and dpa4-2 sod7-2 cuc triple mutants. C. Ovary width in WT, single klu-4 , dpa4-2 sod7-2 double and dpa4-2 sod7-2 klu-4 triple mutants. D. DR5:VENUS ( DR5 ) expression pattern in yellow marking the auxin response in a young petal and cauline leaf. Chlorophyll fluorescence is shown in red. Scale bars = 100 µm in A and D. In B and C , data were normalized to the mean value of the WT. Rectangle represent the first and third quartiles, with a horizontal central line marking the median. Mean values are represented as red diamonds. In B and C, a Kruskal-Wallis test followed by a Dunn’s post-hoc test were performed to show significant differences between different mutants (p-value<0.05). Supplemental Figure 3. DPA4 / SOD7 control sepal and petal patterning via the CUC genes. A. Petal number in WT, single cuc , dpa4-2 sod7-2 double and dpa4-2 sod7-2 cuc triple mutants. B. Petal number in WT, cuc double, dpa4-2 sod7 double and dpa4-2 sod7-2 cuc quadruple mutants C. Petal number in WT, single klu-4 , dpa4-2 sod7-2 double and dpa4-2 sod7-2 klu-4 triple mutants. D. Sepal frequency in WT, single cuc , dpa4-2 sod7-2 double and dpa4-2 sod7-2 cuc triple mutants. E. Sepal frequency in WT, cuc double, dpa4-2 sod7 double and dpa4-2 sod7-2 cuc quadruple mutants F. Sepal frequency in WT, single klu-4 , dpa4-2 sod7-2 double and dpa4-2 sod7-2 klu-4 triple mutants. G. Petal frequency in WT, single cuc , dpa4-2 sod7-2 double and dpa4-2 sod7-2 cuc triple mutants. H. Petal frequency in WT, cuc double, dpa4-2 sod7 double and dpa4-2 sod7-2 cuc quadruple mutants I. Petal frequency in WT, single klu-4 , dpa4-2 sod7-2 double and dpa4-2 sod7-2 klu-4 triple mutants. In A - C, a Kruskal-Wallis test followed by a Dunn’s post-hoc test was performed to show significant differences between our different mutants (p-value<0.05). Supplemental Figure 4. DPA4, SOD7, CUC2, CUC2 and KLU reporter expression during flower development. A. Inflorescence apex of a plant expressing simultaneously pSOD7:GFP (SOD7, green) and pCUC2:RFP (CUC2, red) reporters. The two first panels show separated channels while the last one shows the overlay. B. Inflorescence apex of a plant expressing simultaneously pDPA4:GFP (DPA4, green) and pCUC2:RFP (CUC2, red) reporters. The two first panels show separated channels while the last one shows the overlay. C. Inflorescence apex of a plant expressing simultaneously pKLU:GFP (KLU, green) and pCUC3:CFP (CUC3, cyan) reporters. The two first panels show separated channels while the last one shows the overlay. D to H . Details of reporter expression at different floral stages indicated on the right (S1 to S5). The column D shows pSOD7:GFP (SOD7, green) expression. The column E shows pCUC2:RFP (CUC2, red) expression. The column F shows pDPA4:GFP (DPA4, green) expression. The column G shows pKLU:GFP (KLU, green) expression. The column H shows pCUC3:CFP (CUC3, cyan) expression. Note that pSOD7:GFP and pCUC2:RFP are expressed simultaneously in the same plants, as are pKLU:GFP and pCUC3:CFP . Scale bars = 50 µm in all panels. Supplemental Figure 5. DPA4, SOD7, CUC2, CUC2 and KLU reporter expression during ovule development. A-D. Developing ovules expressing a pDPA4:GFP ( DPA4 , green) reporter. E-H. Developing ovules expressing a pSOD7:GFP ( SOD7 , green) reporter. E-H. Developing ovules expressing simultaneously a pCUC2:RFP ( CUC2, red) and pCUC3:CFP ( CUC3, cyan) reporter. E-H. Developing ovules expressing a pKLU:GFP ( KLU , green) reporter. Bars = 50 µm in all panels Acknowledgments The IJPB benefits from the support of Saclay Plant Sciences-SPS (ANR-17-EUR-0007). This work has benefited from the support of IJPB’s Plant Observatory platforms PO-Plants and PO-Cyto. PP was partly supported by a scholarship from the Erasmus+ program of the European Union. We thank Michael Lenhard for helpful comments on the manuscript. References ↵ Adamski NM , Anastasiou E , Eriksson S , O’Neill CM , and Lenhard M . Local maternal control of seed size by KLUH/CYP78A5-dependent growth signaling . Proc Natl Acad Sci U S A . 2009 : 106 ( 47 ): 20115 – 20 . doi: 10.1073/pnas.0907024106 OpenUrl Abstract / FREE Full Text ↵ Aida M , Ishida T , Fukaki H , Fujisawa H , and Tasaka M . Genes involved in organ separation in Arabidopsis: an analysis of the cup-shaped cotyledon mutant . Plant Cell . 1997 : 9 ( 6 ): 841 – 857 . doi: 10.1105/tpc.9.6.841 OpenUrl Abstract / FREE Full Text ↵ Aida M , Ishida T , and Tasaka M . Shoot apical meristem and cotyledon formation during Arabidopsis embryogenesis: interaction among the CUP-SHAPED COTYLEDON and SHOOT MERISTEMLESS genes . Development . 1999 : 126 ( 8 ): 1563 – 70 . doi: 10.1242/dev.126.8.1563 OpenUrl Abstract ↵ Aida M , Tsubakimoto Y , Shimizu S , Ogisu H , Kamiya M , Iwamoto R , Takeda S , Karim M , Mizutani M , Lenhard M , et al. Establishment of the Embryonic Shoot Meristem Involves Activation of Two Classes of Genes with Opposing Functions for Meristem Activities . Int J Mol Sci . 2020 : 21 ( 16 ): 5864 . doi: 10.3390/ijms21165864 OpenUrl CrossRef PubMed ↵ Anastasiou E , Kenz S , Gerstung M , MacLean D , Timmer J , Fleck C , and Lenhard M . Control of plant organ size by KLUH/CYP78A5-dependent intercellular signaling . Dev Cell . 2007 : 13 ( 6 ): 843 – 56 . doi: 10.1016/j.devcel.2007.10.001 OpenUrl CrossRef PubMed Web of Science ↵ Badia-i-Mompel P , Wessels L , Müller-Dott S , Trimbour R , Ramirez Flores RO , Argelaguet R , and Saez-Rodriguez J . Gene regulatory network inference in the era of single-cell multi-omics . Nat Rev Genet . 2023 : 24 ( 11 ): 739 – 754 . doi: 10.1038/s41576-023-00618-5 OpenUrl CrossRef PubMed ↵ Barro-Trastoy D , Gomez MD , Blanco-Touriñán N , Tornero P , and Perez-Amador MA . Gibberellins regulate ovule number through a DELLA–CUC2 complex in Arabidopsis . The Plant Journal . 2022 : 1 – 15 . doi: 10.1111/tpj.15607 OpenUrl CrossRef ↵ Baumgart LA , Morales-Cruz A , Greenblum SI , Wang P , Zhang Y , Yang L , Chen C , Dilworth DJ , Garretson AC , Grosjean N , et al. An atlas of conserved transcription factor binding sites reveals the cell type-resolved gene regulatory landscape of flowering plants . doi: 10.1101/2024.10.08.617089 OpenUrl Abstract / FREE Full Text ↵ Benfey PN and Weigel D . Transcriptional Networks Controlling Plant Development . Plant Physiol . 2001 : 125 ( 1 ): 109 – 111 . doi: 10.1104/pp.125.1.109 OpenUrl FREE Full Text ↵ Bilsborough GD , Runions A , Barkoulas M , Jenkins HW , Hasson A , Galinha C , Laufs P , Hay A , Prusinkiewicz P , and Tsiantis M . Model for the regulation of Arabidopsis thaliana leaf margin development . Proceedings of the National Academy of Sciences . 2011 : 108 ( 8 ): 3424 – 3429 . doi: 10.1073/pnas.1015162108 OpenUrl Abstract / FREE Full Text ↵ Bouré N , Peaucelle A , Goussot M , Adroher B , Soubigou-Taconnat L , Borrega N , Biot E , Tariq Z , Martin-Magniette M-L , Pautot V , et al. A cell wall-associated gene network shapes leaf boundary domains . Development . 2022 : 149 ( 11 ). doi: 10.1242/dev.200359 OpenUrl CrossRef ↵ Brady SM , Zhang L , Megraw M , Martinez NJ , Jiang E , Yi CS , Liu W , Zeng A , Taylor-Teeples M , Kim D , et al. A stele-enriched gene regulatory network in the Arabidopsis root . Mol Syst Biol . 2011 : 7 : 459 . doi: 10.1038/msb.2010.114 OpenUrl Abstract / FREE Full Text ↵ Cantó-Pastor A , Kajala K , Shaar-Moshe L , Manzano C , Timilsena P , De Bellis D , Gray S , Holbein J , Yang H , Mohammad S , et al. A suberized exodermis is required for tomato drought tolerance . Nature Plants . 2024 : 10 ( 1 ): 118 – 130 . doi: 10.1038/s41477-023-01567-x OpenUrl CrossRef PubMed ↵ Cary GA , McCauley BS , Zueva O , Pattinato J , Longabaugh W , and Hinman VF . Systematic comparison of sea urchin and sea star developmental gene regulatory networks explains how novelty is incorporated in early development . Nat Commun . 2020 : 11 ( 1 ): 6235 . doi: 10.1038/s41467-020-20023-4 OpenUrl CrossRef PubMed ↵ Chakrabarti M , Zhang N , Sauvage C , Muños S , Blanca J , Cañizares J , Diez MJ , Schneider R , Mazourek M , McClead J , et al. A cytochrome P450 regulates a domestication trait in cultivated tomato . Proc Natl Acad Sci U S A . 2013 : 110 ( 42 ): 17125 – 17130 . doi: 10.1073/pnas.1307313110 OpenUrl Abstract / FREE Full Text ↵ Chandler JW , Jacobs B , Cole M , Comelli P , and Werr W . DORNRÖSCHEN-LIKE expression marks Arabidopsis floral organ founder cells and precedes auxin response maxima . Plant Mol Biol . 2011 : 76 ( 1–2 ): 171 – 185 . doi: 10.1007/s11103-011-9779-8 OpenUrl CrossRef PubMed Web of Science ↵ Chen S , Zhang N , Zhang Q , Zhou G , Tian H , Hussain S , Ahmed S , Wang T , and Wang S . Genome editing to integrate seed size and abiotic stress tolerance traits in arabidopsis reveals a role for DPA4 and SOD7 in the regulation of inflorescence architecture . Int J Mol Sci . 2019 : 20 ( 11 ): 1 – 15 . doi: 10.3390/ijms20112695 OpenUrl CrossRef ↵ Davidson EH , McClay DR , and Hood L . Regulatory gene networks and the properties of the developmental process . Proceedings of the National Academy of Sciences . 2003 : 100 ( 4 ): 1475 – 1480 . doi: 10.1073/pnas.0437746100 OpenUrl Abstract / FREE Full Text ↵ Doebley J and Lukens L . Transcriptional regulators and the evolution of plant form . Plant Cell . 1998 : 10 ( 7 ): 1075 – 1082 . doi: 10.1105/tpc.10.7.1075 OpenUrl FREE Full Text ↵ Engelhorn J , Reimer JJ , Leuz I , Göbel U , Huettel B , Farrona S , and Turck F . DEVELOPMENT-RELATED PcG TARGET IN THE APEX 4 controls leaf margin architecture in Arabidopsis thaliana . Development . 2012 : 139 ( 14 ): 2566 – 2575 . doi: 10.1242/dev.078618 OpenUrl Abstract / FREE Full Text ↵ Eriksson S , Stransfeld L , Adamski NM , Breuninger H , and Lenhard M . KLUH/CYP78A5-dependent growth signaling coordinates floral organ growth in Arabidopsis . Curr Biol . 2010 : 20 ( 6 ): 527 – 32 . doi: 10.1016/j.cub.2010.01.039 OpenUrl CrossRef PubMed ↵ Furutani M , Vernoux T , Traas J , Kato T , Tasaka M , and Aida M . PIN-FORMED1 and PINOID regulate boundary formation and cotyledon development in Arabidopsis embryogenesis . Development . 2004 : 131 ( 20 ): 5021 – 30 . doi: 10.1242/dev.01388 OpenUrl Abstract / FREE Full Text ↵ Le Gloanec C , Gómez-Felipe A , Alimchandani V , Branchini E , Bauer A , Routier-Kierzkowska A-L , and Kierzkowski D . Modulation of cell differentiation and growth underlies the shift from bud protection to light capture in cauline leaves . Plant Physiol . 2024 . doi: 10.1093/plphys/kiae408 OpenUrl CrossRef ↵ Gonçalves B , Hasson A , Belcram K , Cortizo M , Morin H , Nikovics K , Vialette-Guiraud A , Takeda S , Aida M , Laufs P , et al. A conserved role for CUP-SHAPED COTYLEDON genes during ovule development . The Plant Journal . 2015 : 83 ( 4 ): 732 – 742 . doi: 10.1111/tpj.12923 OpenUrl CrossRef PubMed ↵ Guillotin B , Rahni R , Passalacqua M , Mohammed MA , Xu X , Raju SK , Ramírez CO , Jackson D , Groen SC , Gillis J , et al. A pan-grass transcriptome reveals patterns of cellular divergence in crops . Nature . 2023 : 617 ( 7962 ):785–791. doi: 10.1038/s41586-023-06053-0 OpenUrl CrossRef Das Gupta M and Tsiantis M . Gene networks and the evolution of plant morphology . Curr Opin Plant Biol . 2018 : 45 : 82 – 87 . doi: 10.1016/j.pbi.2018.05.011 OpenUrl CrossRef PubMed ↵ Halfon MS . Perspectives on Gene Regulatory Network Evolution . Trends in Genetics . 2017 : 33 ( 7 ): 436 – 447 . doi: 10.1016/j.tig.2017.04.005 OpenUrl CrossRef ↵ Hasson A , Plessis A , Blein T , Adroher B , Grigg S , Tsiantis M , Boudaoud A , Damerval C , and Laufs P . Evolution and Diverse Roles of the CUP-SHAPED COTYLEDON Genes in Arabidopsis Leaf Development . Plant Cell . 2011 : 23 ( 1 ): 54 – 68 . doi: 10.1105/tpc.110.081448 OpenUrl Abstract / FREE Full Text ↵ Hay A , Barkoulas M , and Tsiantis M . ASYMMETRIC LEAVES1 and auxin activities converge to repress BREVIPEDICELLUS expression and promote leaf development in Arabidopsis . Development . 2006 : 133 ( 20 ): 3955 – 61 . doi: 10.1242/dev.02545 OpenUrl Abstract / FREE Full Text ↵ Hinman VF , Nguyen AT , Cameron RA , and Davidson EH . Developmental gene regulatory network architecture across 500 million years of echinoderm evolution . Proceedings of the National Academy of Sciences . 2003 : 100 ( 23 ): 13356 – 13361 . doi: 10.1073/pnas.2235868100 OpenUrl Abstract / FREE Full Text ↵ Hu Z-L , Wilson-Sánchez D , Bhatia N , Rast-Somssich MI , Wu A , Vlad D , McGuire L , Nikolov LA , Laufs P , Gan X , et al. A CUC1/auxin genetic module links cell polarity to patterned tissue growth and leaf shape diversity in crucifer plants . Proceedings of the National Academy of Sciences . 2024 : 121 ( 26 ). doi: 10.1073/pnas.2321877121 OpenUrl CrossRef ↵ Jiggins CD , Wallbank RWR , and Hanly JJ . Waiting in the wings: what can we learn about gene co-option from the diversification of butterfly wing patterns? Philosophical Transactions of the Royal Society B: Biological Sciences . 2017 : 372 ( 1713 ):20150485. doi: 10.1098/rstb.2015.0485 OpenUrl CrossRef PubMed ↵ Kajala K , Gouran M , Shaar-Moshe L , Mason GA , Rodriguez-Medina J , Kawa D , Pauluzzi G , Reynoso M , Canto-Pastor A , Manzano C , et al. Innovation, conservation, and repurposing of gene function in root cell type development . Cell . 2021 : 184 ( 12 ): 3333 – 3348 .e19. doi: 10.1016/j.cell.2021.04.024 OpenUrl CrossRef PubMed ↵ Kamimoto K , Adil MT , Jindal K , Hoffmann CM , Kong W , Yang X , and Morris SA . Gene regulatory network reconfiguration in direct lineage reprogramming . Stem Cell Reports . 2023 : 18 ( 1 ): 97 – 112 . doi: 10.1016/j.stemcr.2022.11.010 OpenUrl CrossRef PubMed ↵ Kawamura E , Horiguchi G , and Tsukaya H . Mechanisms of leaf tooth formation in Arabidopsis . Plant J . 2010 : 62 ( 3 ): 429 – 41 . doi: 10.1111/j.1365-313X.2010.04156.x OpenUrl CrossRef PubMed Web of Science ↵ Kazama T , Ichihashi Y , Murata S , and Tsukaya H . The mechanism of cell cycle arrest front progression explained by a KLUH/CYP78A5-dependent mobile growth factor in developing leaves of Arabidopsis thaliana . Plant Cell Physiol . 2010 : 51 ( 6 ): 1046 – 1054 . doi: 10.1093/pcp/pcq051 OpenUrl CrossRef PubMed Web of Science ↵ Kierzkowski D , Runions A , Vuolo F , Strauss S , Lymbouridou R , Routier-Kierzkowska AL , Wilson-Sánchez D , Jenke H , Galinha C , Mosca G , et al. A Growth-Based Framework for Leaf Shape Development and Diversity . Cell . 2019 : 177 ( 6 ): 1405 – 1418 .e17. doi: 10.1016/j.cell.2019.05.011 OpenUrl CrossRef PubMed ↵ Kittelmann S , Buffry AD , Franke FA , Almudi I , Yoth M , Sabaris G , Couso JP , Nunes MDS , Frankel N , Gómez-Skarmeta JL , et al. Gene regulatory network architecture in different developmental contexts influences the genetic basis of morphological evolution . PLoS Genet . 2018 : 14 ( 5 ). doi: 10.1371/journal.pgen.1007375 OpenUrl CrossRef PubMed ↵ Kong S , Zhu M , Pan D , Lane B , Smith RS , and Roeder AHK . Tradeoff between speed and robustness in primordium initiation mediated by auxin-CUC1 interaction . Nat Commun . 2024 : 15 ( 1 ): 5911 . doi: 10.1038/s41467-024-50172-9 OpenUrl CrossRef PubMed ↵ Larue CT , Wen J , and Walker JC . A microRNA-transcription factor module regulates lateral organ size and patterning in Arabidopsis . Plant J . 2009 : 58 ( 3 ): 450 – 63 . doi: 10.1111/j.1365-313X.2009.03796.x OpenUrl CrossRef PubMed Web of Science ↵ Lenhard M , Bohnert A , Jürgens G , and Laux T . Termination of stem cell maintenance in Arabidopsis floral meristems by interactions between WUSCHEL and AGAMOUS . Cell . 2001 : 105 ( 6 ): 805 – 14 . doi: 10.1016/s0092-8674(01)00390-7 OpenUrl CrossRef PubMed Web of Science ↵ Li M , Yao T , Lin W , Hinckley WE , Galli M , Muchero W , Gallavotti A , Chen J-G , and Huang SC . Double DAP-seq uncovered synergistic DNA binding of interacting bZIP transcription factors . Nat Commun . 2023 : 14 ( 1 ): 2600 . doi: 10.1038/s41467-023-38096-2 OpenUrl CrossRef PubMed ↵ Li N , Xu R , and Li Y . Molecular Networks of Seed Size Control in Plants . Annual Review of Plant Biology Annu Rev Plant Biol . 2024 : 35 : 28 . doi: 10.1146/annurev-arplant-050718 OpenUrl CrossRef PubMed ↵ Li X , Zheng Y , Xing Q , Ardiansyah R , Zhou H , Ali S , Jing T , Tian J , Song XS , Li Y , et al. Ectopic expression of the transcription factor CUC2 restricts growth by cell cycle inhibition in Arabidopsis leaves . Plant Signal Behav . 2020 : 15 ( 1 ): 1706024 . doi: 10.1080/15592324.2019.1706024 OpenUrl CrossRef PubMed ↵ Longo SK , Guo MG , Ji AL , and Khavari PA . Integrating single-cell and spatial transcriptomics to elucidate intercellular tissue dynamics . Nat Rev Genet . 2021 : 22 ( 10 ): 627 – 644 . doi: 10.1038/s41576-021-00370-8 OpenUrl CrossRef PubMed ↵ de Luis Balaguer MA , Fisher AP , Clark NM , Fernandez-Espinosa MG , Möller BK , Weijers D , Lohmann JU , Williams C , Lorenzo O , and Sozzani R . Predicting gene regulatory networks by combining spatial and temporal gene expression data in Arabidopsis root stem cells . Proceedings of the National Academy of Sciences . 2017 : 114 ( 36 ): E7632 – E7640 . doi: 10.1073/pnas.1707566114 OpenUrl Abstract / FREE Full Text ↵ Lynch JA and Roth S . The evolution of dorsal–ventral patterning mechanisms in insects . Genes Dev . 2011 : 25 ( 2 ): 107 – 118 . doi: 10.1101/gad.2010711 OpenUrl Abstract / FREE Full Text ↵ Martik ML and McClay DR . Deployment of a retinal determination gene network drives directed cell migration in the sea urchin embryo . Elife . 2015 : 4 . doi: 10.7554/eLife.08827 OpenUrl CrossRef PubMed DH Gonzalez Maugarny A , Gonçalves B , Arnaud N , and Laufs P. CUC Transcription Factors: To the Meristem and Beyond . . In. Plant Transcription Factors , DH Gonzalez , ed. ( Elsevier ), pp. 229 – 247 . doi: 10.1016/B978-0-12-800854-6.00015-4 OpenUrl CrossRef ↵ Maugarny A , Vialette A , Adroher B , Sarthou A-S , Mathy-Franchet N , Azzopardi M , Nicolas A , Roudier F , and Laufs P . MIR164B ensures robust Arabidopsis leaf development by compensating for compromised POLYCOMB REPRESSIVE COMPLEX2 function . Plant Cell . 2024 . doi: 10.1093/plcell/koae260 OpenUrl CrossRef ↵ Maugarny-Calès A , Cortizo M , Adroher B , Borrega N , Gonçalves B , Brunoud G , Vernoux T , Arnaud N , and Laufs P . Dissecting the pathways coordinating patterning and growth by plant boundary domains . PLoS Genet . 2019 : 15 ( 1 ): e1007913 . doi: 10.1371/journal.pgen.1007913 OpenUrl CrossRef PubMed McQueen E and Rebeiz M. On the specificity of gene regulatory networks: How does network co-option affect subsequent evolution? . In. Current Topics in Developmental Biology . ( Academic Press Inc. ), pp. 375 – 405 . doi: 10.1016/bs.ctdb.2020.03.002 OpenUrl CrossRef PubMed ↵ Mutterer J and Zinck E . Quick-and-clean article figures with FigureJ . J Microsc . 2013 : 252 ( 1 ): 89 – 91 . doi: 10.1111/jmi.12069 OpenUrl CrossRef PubMed Web of Science ↵ Nicolas A and Laufs P . The NGATHA-like Genes DPA4 and SOD7 Are Not Required for Stem Cell Specification during Embryo Development in Arabidopsis thaliana . Int J Mol Sci . 2022 : 23 ( 19 ): 12007 . doi: 10.3390/ijms231912007 OpenUrl CrossRef PubMed ↵ Nicolas A , Maugarny-Calès A , Adroher B , Chelysheva L , Li Y , Burguet J , Bågman A-M , Smit ME , Brady SM , Li Y , et al. De novo stem cell establishment in meristems requires repression of organ boundary cell fate . Plant Cell . 2022 : 34 ( 12 ): 4738 – 4759 . doi: 10.1093/plcell/koac269 OpenUrl CrossRef PubMed ↵ Nikovics K , Blein T , Peaucelle A , Ishida T , Morin H , Aida M , and Laufs P . The Balance between the MIR164A and CUC2 Genes Controls Leaf Margin Serration in Arabidopsis . Plant Cell . 2006 : 18 ( 11 ): 2929 – 2945 . doi: 10.1105/tpc.106.045617 OpenUrl Abstract / FREE Full Text ↵ Nobori T , Monell A , Lee TA , Zhou J , Nery J , and Ecker JR . Time-resolved single-cell and spatial gene regulatory atlas of plants under pathogen attack . 2023 . doi: 10.1101/2023.04.10.536170 OpenUrl Abstract / FREE Full Text ↵ O’Malley RC , Huang SC , Song L , Lewsey MG , Bartlett A , Nery JR , Galli M , Gallavotti A , and Ecker JR . Cistrome and Epicistrome Features Shape the Regulatory DNA Landscape . Cell . 2016 : 165 ( 5 ): 1280 – 1292 . doi: 10.1016/j.cell.2016.04.038 OpenUrl CrossRef PubMed ↵ Paaby AB and Rockman M V . The many faces of pleiotropy . Trends in Genetics . 2013 : 29 ( 2 ): 66 – 73 . doi: 10.1016/j.tig.2012.10.010 OpenUrl CrossRef PubMed ↵ Peaucelle A , Morin H , Traas J , and Laufs P . Plants expressing a miR164-resistant CUC2 gene reveal the importance of post-meristematic maintenance of phyllotaxy in Arabidopsis . Development . 2007 : 134 ( 6 ): 1045 – 50 . doi: 10.1242/dev.02774 OpenUrl Abstract / FREE Full Text ↵ Pelaz S , Tapia-López R , Alvarez-Buylla ER , and Yanofsky MF . Conversion of leaves into petals in Arabidopsis . Curr Biol . 2001 : 11 ( 3 ): 182 – 4 . doi: 10.1016/s0960-9822(01)00024-0 OpenUrl CrossRef PubMed Web of Science ↵ Peter IS and Davidson EH . Evolution of Gene Regulatory Networks Controlling Body Plan Development . Cell . 2011 : 144 ( 6 ): 970 – 985 . doi: 10.1016/j.cell.2011.02.017 OpenUrl CrossRef PubMed Web of Science ↵ Preibisch S , Saalfeld S , and Tomancak P . Globally optimal stitching of tiled 3D microscopic image acquisitions . Bioinformatics . 2009 : 25 ( 11 ): 1463 – 1465 . doi: 10.1093/bioinformatics/btp184 OpenUrl CrossRef PubMed Web of Science ↵ Qian Y and Huang SC . Improving plant gene regulatory network inference by integrative analysis of multi-omics and high resolution data sets . Curr Opin Syst Biol . 2020 : 22 : 8 – 15 . doi: 10.1016/j.coisb.2020.07.010 OpenUrl CrossRef ↵ R Core Team . A language and environment for statistical computing . R Foundation for Statistical Computing , Vienna, Austria. 2022 . https://www.r-project.org/ ↵ Raman S , Greb T , Peaucelle A , Blein T , Laufs P , and Theres K . Interplay of miR164, CUP-SHAPED COTYLEDON genes and LATERAL SUPPRESSOR controls axillary meristem formation in Arabidopsis thaliana . The Plant Journal . 2008 : 55 ( 1 ): 65 – 76 . doi: 10.1111/j.1365-313X.2008.03483.x OpenUrl CrossRef PubMed Web of Science ↵ Rast-Somssich MI , Broholm S , Jenkins H , Canales C , Vlad D , Kwantes M , Bilsborough G , Dello Ioio R , Ewing RM , Laufs P , et al. Alternate wiring of a KNOXI genetic network underlies differences in leaf development of A. thaliana and C. hirsuta . Genes Dev . 2015 : 29 ( 22 ): 2391 – 2404 . doi: 10.1101/gad.269050.115 OpenUrl Abstract / FREE Full Text ↵ Rice G , Gaitán-Escudero T , Charles-Obi K , Zeitlinger J , and Rebeiz M . Co-option of the trichome-forming network initiated the evolution of a morphological novelty in Drosophila eugracilis . Current Biology . 2024 : 34 ( 22 ): 5284 – 5294 .e3. doi: 10.1016/j.cub.2024.09.073 OpenUrl CrossRef PubMed ↵ Robertson SM and Wilkins O . Spatially resolved gene regulatory networks in Asian rice ( Oryza sativa cv . Nipponbare) leaves. The Plant Journal . 2023 : 116 ( 1 ): 269 – 281 . doi: 10.1111/tpj.16375 OpenUrl CrossRef PubMed ↵ Romanel EAC , Schrago CG , Counago RM , Russo CAM , Alves-Ferreira M , Couñago RM , Russo CAM , and Alves-Ferreira M . Evolution of the B3 DNA binding superfamily: New insights into REM family gene diversification . PLoS One . 2009 : 4 ( 6 ): e5791 . doi: 10.1371/journal.pone.0005791 OpenUrl CrossRef PubMed ↵ Rubio-Somoza I , Zhou C , Confraria A , Martinho C , von Born P , Baena-Gonzalez E , Wang J , and Weigel D . Temporal Control of Leaf Complexity by miRNA-Regulated Licensing of Protein Complexes . Current Biology . 2014 : 24 ( 22 ): 2714 – 2719 . doi: 10.1016/j.cub.2014.09.058 OpenUrl CrossRef PubMed ↵ Sauret-Güeto S , Schiessl K , Bangham A , Sablowski R , and Coen E . JAGGED Controls Arabidopsis Petal Growth and Shape by Interacting with a Divergent Polarity Field . PLoS Biol . 2013 : 11 ( 4 ). doi: 10.1371/journal.pbio.1001550 OpenUrl CrossRef PubMed ↵ Scheres B and Krizek BA . Coordination of growth in root and shoot apices by AIL/PLT transcription factors . Curr Opin Plant Biol . 2018 : 41 : 95 – 101 . doi: 10.1016/j.pbi.2017.10.002 OpenUrl CrossRef PubMed ↵ Schindelin J , Arganda-Carreras I , Frise E , Kaynig V , Longair M , Pietzsch T , Preibisch S , Rueden C , Saalfeld S , Schmid B , et al. Fiji: an open-source platform for biological-image analysis . Nat Methods . 2012 : 9 ( 7 ): 676 – 682 . doi: 10.1038/nmeth.2019 OpenUrl CrossRef PubMed Web of Science ↵ Schneitz K , Hülskamp M , and Pruitt RE . Wild-type ovule development in Arabidopsis thaliana fIJ: a light microscope study of cleared whole-mount tissue . The Plant Journal . 1995 : 7 ( 5 ): 731 – 749 . doi: 10.1046/j.1365-313X.1995.07050731.x OpenUrl CrossRef Web of Science ↵ Schoof H , Lenhard M , Haecker A , Mayer KF , Jürgens G , and Laux T . The stem cell population of Arabidopsis shoot meristems in maintained by a regulatory loop between the CLAVATA and WUSCHEL genes . Cell . 2000 : 100 ( 6 ): 635 – 44 . doi: 10.1016/s0092-8674(00)80700-x OpenUrl CrossRef PubMed Web of Science ↵ Scotland RW . Deep homology: A view from systematics . BioEssays . 2010 : 32 ( 5 ): 438 – 449 . doi: 10.1002/bies.200900175 OpenUrl CrossRef PubMed Web of Science ↵ Serra L and Perrot-Rechenmann C . Spatio temporal control of cell growth by CUC3 shapes leaf margins . Development . 2020 : 147 ( 6 ). doi: 10.1242/dev.183277 OpenUrl Abstract / FREE Full Text ↵ Shahan R , Hsu C-W , Nolan TM , Cole BJ , Taylor IW , Greenstreet L , Zhang S , Afanassiev A , Vlot AHC , Schiebinger G , et al. A single-cell Arabidopsis root atlas reveals developmental trajectories in wild-type and cell identity mutants . Dev Cell . 2022 : 57 ( 4 ): 543 – 560 .e9. doi: 10.1016/j.devcel.2022.01.008 OpenUrl CrossRef PubMed Shankar N , Gupta A , Roy S , Mane V , Hamant O , and Nath U . CUP-SHAPED COTYLEDON2 activates MIR319C transcription and 1 promotes cell proliferation in Arabidopsis leaf primordia 2 3 . doi: 10.1101/2024.09.07.611804 OpenUrl Abstract / FREE Full Text ↵ Shao J , Liu X , Wang R , Zhang G , and Yu F . The Over-Expression of an Arabidopsis B3 Transcription Factor, ABS2/NGAL1, Leads to the Loss of Flower Petals . PLoS One . 2012 : 7 ( 11 ): e49861 . doi: 10.1371/journal.pone.0049861 OpenUrl CrossRef PubMed ↵ Shao J , Meng J , Wang F , Shou B , Chen Y , Xue H , Zhao J , Qi Y , An L , Yu F , et al. NGATHA-LIKEs Control leaf margin development by repressing cup-shaped cotyledon2 transcription1 . Plant Physiol . 2020 : 184 ( 1 ): 345 – 358 . doi: 10.1104/PP.19.01598 OpenUrl Abstract / FREE Full Text ↵ Sieber P , Wellmer F , Gheyselinck J , Riechmann JL , and Meyerowitz EM . Redundancy and specialization among plant microRNAs: role of the MIR164 family in developmental robustness . Development . 2007 : 134 ( 6 ): 1051 – 60 . doi: 10.1242/dev.02817 OpenUrl Abstract / FREE Full Text ↵ Smyth DR , Bowman JL , and Meyerowitz EM . Early flower development in Arabidopsis . Plant Cell . 1990 : 2 ( 8 ): 755 – 67 . doi: 10.1105/tpc.2.8.755 OpenUrl Abstract / FREE Full Text ↵ Sonawane AR , Platig J , Fagny M , Chen CY , Paulson JN , Lopes-Ramos CM , DeMeo DL , Quackenbush J , Glass K , and Kuijjer ML . Understanding Tissue-Specific Gene Regulation . Cell Rep . 2017 : 21 ( 4 ): 1077 – 1088 . doi: 10.1016/j.celrep.2017.10.001 OpenUrl CrossRef PubMed ↵ Srivastava M . Beyond Casual Resemblance: Rigorous Frameworks for Comparing Regeneration Across Species . Annual Review of Cell and Developmental Biology Annu Rev Cell Dev Biol 2021 . 2021 : 37 : 415 – 455 . doi: 10.1146/annurev-cellbio-120319 OpenUrl CrossRef ↵ Stearns FW . One hundred years of pleiotropy: A retrospective . Genetics . 2010 : 186 ( 3 ): 767 – 773 . doi: 10.1534/genetics.110.122549 OpenUrl Abstract / FREE Full Text ↵ Swaminathan K , Peterson K , and Jack T . The plant B3 superfamily . Trends Plant Sci . 2008 : 13 ( 12 ): 647 – 655 . doi: 10.1016/j.tplants.2008.09.006 OpenUrl CrossRef PubMed Web of Science ↵ Swift J , Luginbuehl LH , Hua L , Schreier TB , Donald RM , Stanley S , Wang N , Lee TA , Nery JR , Ecker JR , et al. Exaptation of ancestral cell-identity networks enables C4 photosynthesis . Nature . 2024 . doi: 10.1038/s41586-024-08204-3 OpenUrl CrossRef ↵ Takada S , Hibara K , Ishida T , and Tasaka M . The CUP-SHAPED COTYLEDON1 gene of Arabidopsis regulates shoot apical meristem formation . Development . 2001 : 128 ( 7 ): 1127 – 35 . doi: 10.1242/dev.128.7.1127 OpenUrl Abstract ↵ Tian C , Zhang X , He J , Yu H , Wang Y , Shi B , Han Y , Wang G , Feng X , Zhang C , et al. An organ boundary-enriched gene regulatory network uncovers regulatory hierarchies underlying axillary meristem initiation . Mol Syst Biol . 2014 : 10 ( 10 ): 755 . doi: 10.15252/msb.20145470 OpenUrl Abstract / FREE Full Text ↵ Tripathi RK and Wilkins O . Single cell gene regulatory networks in plants: Opportunities for enhancing climate change stress resilience . Plant Cell Environ . 2021 : 44 ( 7 ): 2006 – 2017 . doi: 10.1111/pce.14012 OpenUrl CrossRef ↵ Tyler AL , Asselbergs FW , Williams SM , and Moore JH . Shadows of complexity: What biological networks reveal about epistasis and pleiotropy . BioEssays . 2009 : 31 ( 2 ): 220 – 227 . doi: 10.1002/bies.200800022 OpenUrl CrossRef PubMed Web of Science ↵ Vroemen CW , Mordhorst AP , Albrecht C , Kwaaitaal MACJ , and de Vries SC . The CUP-SHAPED COTYLEDON3 gene is required for boundary and shoot meristem formation in Arabidopsis . Plant Cell . 2003 : 15 ( 7 ): 1563 – 77 . doi: 10.1105/tpc.012203 OpenUrl Abstract / FREE Full Text Wickham H. ggplot2: Elegant Graphics for Data Analysis ( Springer-Verlag : New York ). ↵ Yin R , Xia K , and Xu X . Spatial transcriptomics drives a new era in plant research . The Plant Journal . 2023 : 116 ( 6 ): 1571 – 1581 . doi: 10.1111/tpj.16437 OpenUrl CrossRef PubMed ↵ Yuan Q and Duren Z . Inferring gene regulatory networks from single-cell multiome data using atlas-scale external data . Nat Biotechnol . 2024 . doi: 10.1038/s41587-024-02182-7 OpenUrl CrossRef ↵ Zhang S , Pyne S , Pietrzak S , Halberg S , McCalla SG , Siahpirani AF , Sridharan R , and Roy S . Inference of cell type-specific gene regulatory networks on cell lineages from single cell omic datasets . Nat Commun . 2023 : 14 ( 1 ). doi: 10.1038/s41467-023-38637-9 OpenUrl CrossRef PubMed ↵ Zhang Y , Du L , Xu R , Cui R , Hao J , Sun C , and Li Y . Transcription Factors SOD7/NGAL2 and DPA4/NGAL3 Act Redundantly to Regulate Seed Size by Directly Repressing KLU Expression in Arabidopsis thaliana . Plant Cell . 2015 : 27 ( 3 ): 620 – 632 . doi: 10.1105/tpc.114.135368 OpenUrl Abstract / FREE Full Text ↵ Zheng L , Wu H , Wang A , Zhang Y , Liu Z , Ling H-Q , Song X-J , and Li Y . The SOD7/DPA4–GIF1 module coordinates organ growth and iron uptake in Arabidopsis . Nat Plants . 2023 : 9 ( 8 ): 1318 – 1332 . doi: 10.1038/s41477-023-01475-0 OpenUrl CrossRef PubMed ↵ Zhu J , Lolle S , Tang A , Guel B , Kvitko B , Cole B , and Coaker G . Single-cell profiling of Arabidopsis leaves to Pseudomonas syringae infection . Cell Rep . 2023 : 42 ( 7 ). doi: 10.1016/j.celrep.2023.112676 OpenUrl CrossRef PubMed Back to top Previous Next Posted December 13, 2024. Download PDF Supplementary Material Email Thank you for your interest in spreading the word about bioRxiv. NOTE: Your email address is requested solely to identify you as the sender of this article. Your Email * Your Name * Send To * Enter multiple addresses on separate lines or separate them with commas. You are going to email the following Developmental rewiring of the NGAL/CUC/KLU network associated with pleiotropic roles of NGAL genes Message Subject (Your Name) has forwarded a page to you from bioRxiv Message Body (Your Name) thought you would like to see this page from the bioRxiv website. Your Personal Message CAPTCHA This question is for testing whether or not you are a human visitor and to prevent automated spam submissions. Share Developmental rewiring of the NGAL / CUC / KLU network associated with pleiotropic roles of NGAL genes Antoine Nicolas , Panagiotis Papadopoulos , Mattéo Caroulle , Bernard Adroher , Magali Goussot , Anne-Sophie Sarthou , Nicolas Arnaud , Aude Maugarny , Patrick Laufs bioRxiv 2024.12.12.627899; doi: https://doi.org/10.1101/2024.12.12.627899 Share This Article: Copy Citation Tools Developmental rewiring of the NGAL / CUC / KLU network associated with pleiotropic roles of NGAL genes Antoine Nicolas , Panagiotis Papadopoulos , Mattéo Caroulle , Bernard Adroher , Magali Goussot , Anne-Sophie Sarthou , Nicolas Arnaud , Aude Maugarny , Patrick Laufs bioRxiv 2024.12.12.627899; doi: https://doi.org/10.1101/2024.12.12.627899 Citation Manager Formats BibTeX Bookends EasyBib EndNote (tagged) EndNote 8 (xml) Medlars Mendeley Papers RefWorks Tagged Ref Manager RIS Zotero Tweet Widget Facebook Like Google Plus One Subject Area Plant Biology Subject Areas All Articles Animal Behavior and Cognition (7807) Biochemistry (18226) Bioengineering (14392) Bioinformatics (43131) Biophysics (22001) Cancer Biology (19111) Cell Biology (26194) Clinical Trials (138) Developmental Biology (13651) Ecology (20406) Epidemiology (2067) Evolutionary Biology (24867) Genetics (15861) Genomics (22996) Immunology (18215) Microbiology (41382) Molecular Biology (17552) Neuroscience (90901) Paleontology (679) Pathology (2907) Pharmacology and Toxicology (4955) Physiology (7881) Plant Biology (15518) Scientific Communication and Education (2068) Synthetic Biology (4433) Systems Biology (10014) Zoology (2319) window.__CF$cv$params={r:'a1deec761dc97414',t:'MTc4NDUxODYwOA==',u:'019f7d990e527361869b5845872eb9bf',ut:'ckip2s147OAJTeSmcSdUAC6EbeJsA3pvL4j.7EoTq3E-1784518610-1.2.1.1-88xn2ZwjtYzdEeOiS3jYaOPccrJbzdnbnKISA.BUd3deKPQ2JRkmU3eAEOvNm_C6AG2liIuv0PaoMVRvFfkPVGosnOwuGOonGNycu6XqW0w',i:60};(function(){if(!document.body)return;var s=document.createElement('script');s.src='/cdn-cgi/challenge-platform/scripts/precursor/main.js';document.head.appendChild(s);})();