Plant Reproduction

The male germ unit (MGU) in Arabidopsis pollen is comprised of one vegetative nucleus (VN) and two sperm nuclei (SN). It is evolutionarily specialized to deliver immotile sperm nuclei to an ovule for fertilization. Despite some progress in research on MGU, its organization and transport remain only partially understood. Here, we identified Nucleoporin1/136 as a new player in the structural organization and positioning of MGU in pollen tubes. We and others have previously reported the reduced fertility of nup1-1 plants; however, the mechanism remains unknown. In this work, we further examined the role of NUP1 in fertility using two mutant alleles, nup1-1 and nup1-2-/+. The reciprocal crosses between the nup1 mutants and the Col-0 wild type indicate that the nup1 mutant pollen is defective. To study the effect of a complete NUP1 knockout on pollen, we generated a transgenic line that produces pollen with and without NUP1 expression. This work led to the surprising discovery that the NUP1 protein is inherited from the pollen mother cell to the daughter cell during microgametophyte development. Subsequent in vitro experiments showed that NUP1 is required for pollen germination and pollen tube elongation. Further microscopic studies demonstrated that NUP1 is highly expressed in VN and essential for maintaining nuclear shape and size. We also demonstrated that NUP1 is required for proper MGU organization and transport, likely through maintaining VN morphology. Notably, our finding of nuclear morphology-mediated regulation of MGU may also explain the mechanistic details underlying the defective MGU movement in previously reported mutants such as kaku4, wit, and wip, which have abnormal nuclear morphology.

Background and AimsPaspalum notatum reproduces through either sexuality or apomixis, two pathways that may coexist within the same individual and are regulated by interconnected molecular networks responsive to environmental cues. Here, we characterized the transcript structure and expression of BUD SITE SELECTION PROTEIN 13 (BUD13), a component of the RES spliceosomal complex previously reported as differentially expressed in florets of sexual and apomictic plants, as a first step toward testing its involvement in the molecular regulation of the apomixis-sexuality switch. MethodsPreviously generated floral and leaf transcriptomes from sexual and apomictic Paspalum notatum plants, including Oxford Nanopore long-read data, were mined to characterize BUD13 transcript structure and expression. Phylogenetic analyses and in silico mapping were conducted to infer evolutionary relationships and determine the origin of the transcripts. Differential expression was validated by RT-qPCR, while in situ hybridization was used to reveal cell-specific ovule expression patterns. Key resultsBUD13 is expressed in Paspalum notatum florets as a truncated isoform (SHORT) encoding a small protein lacking part of the herpes simplex virus regulatory protein (ICP4) domain. Two SHORT transcripts, SHORT1 and SHORT2, with different 5' untranslated region (UTR) regions, were identified in flowers. SHORT1 was consistently upregulated in apomictic ovules from premeiosis to anthesis. Both transcripts originated from a single genomic locus located in the subtelomeric region of the short arm of chromosome 6. SHORT isoforms with variable structures were detected in other monocots. In situ hybridization showed that, whereas BUD13 was expressed throughout sexual ovules, expression was absent from the female germline of apomictic ovules. A consistent expression was observed in somatic proembryos of aposporous embryo sacs. ConclusionsOur findings reveal structural, spatial and temporal divergence in BUD13 expression between sexual and apomictic reproductive programs, providing new insights into the molecular regulation of asexual seed formation.

The number of pollen grains, which carry male gametes in seed plants, has attracted interest in genetics, evolution, and breeding. Rapid evolutionary reductions in pollen number and anther length were reported in selfing species as well as domesticated species, although this poses a challenge for hybrid breeding. Here, we studied the variation of pollen number and anther length of the hexaploid bread wheat (Triticum aestivum) by employing a quick pollen counting method. Pollen numbers in cultivars were lower than those in landraces among 54 lines of diverse geographic origins. Using the year of registration of traditional and modern cultivars, we found a reduction in pollen number over the past 150 years. We detected high heritability and variation among Asian landraces and cultivars. Thus, we conducted QTL mapping of pollen number as well as of anther length using nested association mapping lines in which Norin 61 was the common parent. Genomic loci encompassing Green Revolution genes (Rht-B1, Rht-D1, and Ppd-D1) showed significant effects on pollen number and anther length, but their contributions were relatively minor. Although anther length has often been used as a proxy for pollen number in bread wheat, our data showed that their correlations are not necessarily high. Interestingly, we identified a new QTL of pollen number that was not detected by measuring anther length, and, vice versa, a new QTL specific to anther length. These data suggest that pollen number has reduced rapidly in bread wheat but can be modified using the genetic diversity of landraces. Significance statementWe found that modern cultivars of bread wheat have reduced pollen number and shorter anther length, which are common in domesticated species but can be a challenge for hybrid breeding. Using underutilized Asian landraces and cultivars, we reported that new quantitative trait loci as well as loci used in the Green Revolution, are responsible for the traits, which can be employed to increase pollen numbers.

Starch synthesis in wheat endosperm involves the initiation of large A-type starch granules during early grain development, followed by small B-type granules in later grain development. It is established that MAR-BINDING FILAMENT-LIKE PROTEIN 1 (MFP1) plays an important role in granule initiation in Arabidopsis chloroplasts, but how it influences A- and B-type initiations in wheat amyloplasts is not known. We discovered that due to a gene duplication in cereals, wheat contains two MFP1 paralogs, MFP1.1 and MFP1.2, which are both expressed in the developing endosperm. We generated a series of durum wheat mutants defective in all homoeologs of either MFP1.1 or MFP1.2, or both. While starch granule size distributions and granule morphology of mfp1.1 and mfp1.2 mutants were identical to those of the wild-type, the mfp1.1 mfp1.2 mutants had fewer, but larger B-type granules - suggesting that the two paralogs play redundant roles in B-type granule initiation. Consistent with this, both paralogs interacted with B-GRANULE CONTENT 1 (BGC1), a key protein required for proper B-type granule initiation in wheat, and both paralogs could partially complement defects in starch initiation in the Arabidopsis mfp1 mutant. Our work demonstrates that MFP1 is required for establishing correct starch granule number in non-photosynthetic amyloplasts, but its role in wheat is limited to B-type granule initiation. One-sentence summaryWheat has two MFP1 paralogs that interact with the granule initiation protein, BGC1 and influence B-type granule initiation in non-photosynthetic amyloplasts of endosperm.

Quantitative pollen viability analysis is a critical but labor-intensive step in plant reproductive biology. Existing deep-learning Segment Anything Models (SAM) fail to reliably segment viable pollen in Alexander-stained anthers. To address this, we fine-tuned an existing Cellpose-SAM model for pollen segmentation. We integrated it into PAT (Pollen Analysis Tool), a cross-platform desktop application. PAT features instance segmentation with interactive quality control, an in-app model retraining module, and publication-ready statistical outputs. We deployed PAT in an EMS suppressor screen of semi-sterile Arabidopsis smg7-6 mutants, enabling efficient candidate prioritization for whole genome sequencing and mapping candidate mutation. This screen led to the identification of a point mutation in CAP-D2 (capd2-2), a Condensin I subunit, that rescues the smg7-6 meiotic phenotype. Notably, mutation in a Condensin II subunits (CAP-D3 and CAP-H2) does not confer rescue. Further characterization suggests the capd2-2 allele is hypomorphic, showing no defects in vegetative growth, chromocenter compaction, or transposable element silencing. Collectively, we demonstrate that accessible AI tools have the potential to bridge gaps in plant phenotyping and accelerate the pace of biological discovery. HighlightWe combined AI-powered image analysis with an easy-to-use desktop app to automate plant pollen counting, then used it to identify a new genetic suppressor of meiotic defects.

Grain legumes such as pea (Pisum sativum L.) accumulate large amounts of seed storage proteins without nitrogen fertilization due to their symbiosis with nitrogen-fixing bacteria, making them a key source of plant-based proteins. Seed growth and the accumulation of seed storage proteins are tightly regulated by complex gene networks; however, the mechanisms governing these processes in pea remain poorly understood. In this study, we generated a comprehensive seed expression atlas covering six developmental stages in pea (cv Cameor), including the key transition stage from embryogenesis to early seed filling, providing a detailed temporal resolution of transcriptional dynamics throughout seed development in this species. Co-expression network analysis highlighted several candidate transcription factors potentially involved in the transition towards seed filling. Among them, we characterized the seed-specific NF-YB transcription factor PsLEC1-like (PsL1L), the major LEC1-type factor expressed during early pea seed development. Functional analyses using TILLING mutants demonstrated that loss of PsL1L function reduces seed size and seed nitrogen content and impairs early embryo growth from the end of embryogenesis. Finally, we show that the expression of the B3-domain transcription factor PsFUS3, but not that of PsLEC2 or PsABI3, is reduced in the loss-of-function l1l mutant, suggesting that PsL1L acts upstream of PsFUS3 to control seed size.

Awns of wild barley (Hordeum vulgare ssp. spontaneum L.) are rough by default due to silicified upward-oriented trichomes on the awns epidermis, forming a ratcheted surface, which is advantageous for seed dispersal and burial. Cultivated barley, however, may carry smooth awns covered by smaller barbs or lacking barbs completely. The gene Raw1 on chromosome 5H is a major factor controlling barley awn roughness and was shown to encode a LONG AND BARBED AWN1 (LABA1) homolog. Here we report, by using quantitative analysis of the barb trait, map-based cloning and Cas9-mediated gene knock-out, a second gene Raw7, located on barley chromosome 7H, encoding a putative two-component response regulator. We propose that Raw7 acts downstream of Raw1 in a cytokinin signaling pathway underlying cell cycle control in epidermal barb primordia cells. Raw1 and Raw7 show epistatic interaction, suggesting that Raw1 acts as the primary driver of barb initiation, while Raw7 modulates barb size and frequency. Our findings provide the foundation to study the selection and domestication history of the awn roughness trait in barley, and thus to dissect if awn roughness is providing an advantage in cultivated barley or if the trait persisted after domestication due to linkage drag. SummaryO_LIThe presence of silicified upward-oriented trichomes or barbs arising from the epidermis of barley awns is a prominent trait. They form a ratcheted surface which is advantageous for seed dispersal and burial and defense against herbivory. C_LIO_LIPrevious work identified Raw1 on chromosome 5H as a major determinant of awn roughness in barley. Here, we identify a second awn roughness gene, Raw7 on chromosome 7H, combining quantitative phenotyping of barb traits, map-based cloning and Cas9-mediated targeted mutagenesis for functional analyses. C_LIO_LIGenetic and functional evidence suggests a complex epistatic interaction in which Raw1 primarily drives barb formation and Raw7 fine-tunes endoreduplication-dependent epidermal cell expansion and patterning in barb primordia cells. Raw1 and Raw7 likely act in a cytokinin-dependent two-component signaling pathway, where Raw1 promotes local cytokinin activation and Raw7, a type-B response regulator, mediates downstream transcriptional responses. C_LIO_LIThe proposed pathway suggests additional undetected loci may contribute to awn roughness, and emerging barley pangenome and pan-transcriptome resources provide a framework to identify and functionally validate new candidates. C_LI

Legumes include some of the worlds most significant crop species, such as cowpea (Vigna unguiculata), a subsistence crop widely grown in sub-Saharan Africa. Despite their importance, legume crop improvement is hindered by a lack of high-resolution expression data, particularly for reproductive tissues and cell types. Here, we report on VigExp, a tool for visualising cowpea gene expression datasets. We demonstrate its utility across a range of vegetative and reproductive cell types of varieties IT97K-499-35 and IT86D-1010, which exhibit 93.75% protein sequence conservation and are amenable to stable transformation. This includes previously published transcriptomes of vegetative, floral and seed tissues, combined with developmentally staged male and female reproductive tissues. Also integrated are novel transcriptomes of laser-captured cell types covering reproductive development from meiosis to early embryo formation post-fertilisation. Spatial expression patterns and transcript levels can be visualised through an electronic fluorescent pictograph (eFP) browser. Validated by RT-qPCR, in situ hybridisation, transgenic, and CRISPR gene editing analyses, the predictive accuracy of VigExp matches prior cowpea functional study observations. Critical genes for nodule development and regulation were also identified and their expression patterns established in cowpea. Novel reference genes, constitutively expressed gene promoters for visualization makers/gene-editing, and tissue and cell specific gene promoters for targeting these regions, are identified. The A-type cyclin, VuTAM2, was also identified, with a critical role in male meiosis established. Collectively, VigExp represents an adaptable and updatable resource to support crop improvement in cowpea and other legumes, which are often highly syntenic with respect to genome composition.

The transformation of the free nuclear syncytium into cellular endosperm tissue with starch and protein accumulation is a well-established phenomenon, at least in the fruits of cereals of the Triticeae tribe. The present article demonstrates that there is considerable diversity inherent in this type of caryopsis morphogenesis. By examining various taxa (species, varieties, and cultivars) of wheat, oats, and some wild grasses, this research reveals significant deviations in endosperm morphogenesis from the typical state. A new developmental pattern of endosperm was identified, characterized by several distinctive features such as incomplete cellularization of the syncytium and starch accumulation within the acellular endosperm domains and the endosperm cavity. A large number of plastids were observed in the syncytium stage, which served as the basis for the later amyloplast stage. The acellular endosperm domains and the cavity domain exhibited connections at specific discontinuities in the modified aleurone layer surrounding the cavity. The peripheral parts of the caryopsis received fewer assimilates necessary for starch synthesis, which was attributed to their increased distance from the transfer system and a likely reduction in the efficiency of assimilate transport through the apoplast in these areas. The starch cavity volume constituted a few percent of the overall caryopsis volume, which could serve as a foundation for potential breeding improvements to enhance starch yields across different varieties.

BackgroundCell geometry plays a central role in determining division orientation and body axis formation during early embryogenesis in Arabidopsis thaliana. However, quantitative analysis of dynamic three-dimensional (3D) morphology remains challenging because live-imaging studies often rely on two-dimensional (2D) projections, while existing 3D reconstruction approaches, including mesh-based methods, often lose the original orientation information relative to the ovule and require labor-intensive mesh correction. In addition, embryo positional fluctuation caused by floating in liquid medium and continuous growth makes it difficult to analyze temporal morphological changes within a common coordinate system. ResultsWe developed a robust framework for quantitative 3D and four-dimensional (4D; 3D + time) analysis of embryo initial cell (apical cell) morphology. The method first establishes a standardized 3D coordinate system by normalizing cell orientation based on the bottom plane and the optical axis of the observation. Cell morphology is then reconstructed through ellipse-based approximation of serial cross-sections extracted from stacked imaging data, enabling accurate geometric characterization without the need for complex surface mesh reconstruction. To evaluate shape anisotropy, we quantified the apical cell shape in 3D. The framework further supports the characterization of volumetric features of subsequent division, providing a basis for quantifying 3D embryogenesis. ConclusionOur framework provides a simple and noise-reduced approach for quantitative analysis of living cell morphology in 3D. We named the integrated method of combining coordinate normalization with elliptical cross-section-based reconstruction Apical3DTip. This method enables consistent comparison of cell shapes without extensive manual corrections. The method overcomes key limitations of 2D projection-based and mesh-dependent analyses and offers a practical platform for quantifying cell shape and daughter cell shapes in 3D. More broadly, it provides a quantitative foundation for exploring the relationship between cell geometry, morphodynamics, and developmental patterning in living plant embryos.

Haploid induction is a key component of doubled haploid technology and an increasingly valuable tool for plant breeding, genome editing, and clonal seed production. While in planta haploid induction through haploid inducer lines offers significant advantages over in vitro approaches, its application remains limited in many crop species. Previously, disruption of the sperm cell-expressed KOKOPELLI (KPL) gene was shown to induce maternal haploids in Arabidopsis thaliana. Here, we report the creation of novel haploid inducer lines in two globally important crops, maize (Zea mays), a major staple food crop, and tomato (Solanum lycopersicum), a widely cultivated vegetable crop. Using targeted genome editing, we generated mutations in KPL orthologs and demonstrated that loss of KPL function confers haploid induction capacity, enabling the production of haploid seedlings in both species. These findings establish KPL as a conserved target to trigger haploid induction and expand the genetic toolbox available for haploid inducer development in crop species.

Pollen development and fertilization are considered the most heat-sensitive stages of plant reproduction. While heat stress severely impairs pollen germination and tube growth, the physiological diversity within a single flowers pollen load suggests that subpopulations may exhibit differential climate resilience. In this study, we tested the hypothesis that this heterogeneity reflects a dormancy-based reserve mechanism that preserves fertilization under heat stress. Using flow cytometry and fluorescence-activated cell sorting in Arabidopsis thaliana and Solanum lycopersicum (MicroTom), we resolved pollen subpopulations by reactive oxygen species (ROS) status and examined their behavior under increasing heat stress. In both species, ROS-defined metabolic state was tightly associated with pollen size: high-ROS pollen was larger and readily germination-competent, whereas low-ROS pollen was smaller and showed low basal germination, consistent with dormancy. Heat stress preferentially depleted the high-ROS fraction, whereas the low-ROS fraction persisted and, under heat stress, increased metabolic activity and size. By isolating low-ROS and high-ROS pollen, we further show that a brief heat treatment suppresses germination of active high-ROS pollen but promotes germination of dormant low-ROS pollen. These findings provide direct evidence that heat can release dormancy in low-ROS pollen and support a conserved model in which dormant pollen serves as a heat-resilient reproductive reserve.

PremiseSeed size and morphology are critical traits in agriculture, ecology, and genetics, but high-throughput quantification of these traits is often limited by labor-intensive manual measurements or expensive, platform-specific imaging software. Methods and ResultsWe developed SeedMeasure, a lightweight, open-source, and cross-platform command-line tool written in Python that automates the measurement of seed area, length, and width from images. Using a simple imaging setup, the program processes images by correcting for perspective skew, filtering debris, and exports quantitative data alongside quality-check images. We validated SeedMeasure across nine diverse species, ranging from small Arabidopsis thaliana seeds to large Zea mays kernels. The tool quickly handles images using multithreading and demonstrates high reproducibility, yielding low coefficients of variation across repeated runs. ConclusionsCompared to existing software, SeedMeasure is free, offers faster processing through parallel computing, and provides standalone executables that require no programming dependencies. SeedMeasure offers an accessible, cost-effective, and high-throughput approach for rapid phenotypic profiling, making advanced seed morphological analysis available to researchers without specialized laboratory hardware.

Insect herbivory triggers cytosolic proteome reprogramming by activating defense pathways and modulating key metabolic processes. We found that simulated herbivory in pigeon pea (Cajanus cajan) induced reactive oxygen species (ROS) production and molecular alterations within 12 hours (h) of post treatment. We compared the leaf proteome profiles of two cultivated genotypes, ICPL 332 (moderately resistant) and ICPL 87 (susceptible), using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) coupled with mass spectrometry (MS). More than 220 protein spots were detected in ICPL 332 and over 200 in ICPL 87. Comparative analysis revealed 75 differentially accumulated proteins (DAPs), of which 40 were consistently reproducible across biological replicates. These included 11 unique to ICPL 87, 9 unique to ICPL 332, and 10 common to both genotypes. Among the shared DAPs, ICPL 332 showed five upregulated and five downregulated, whereas ICPL 87 exhibited only two upregulated and eight downregulated. Functional categorization grouped DAPs into primary metabolism, stress response, and growth and development. Proteins related to primary metabolism were largely downregulated in both genotypes, while stress-associated proteins exhibited substantial downregulation in ICPL 87 compared to ICPL 332. Overall, the results demonstrate proteomic adjustments underlying defense responses in pigeon pea genotypes.

In plant breeding and genetics, recent advances in high-throughput phenotyping are beginning to meet the growing demand for large-scale, high-quality phenotypic data that emerged after the development of next-generation sequencing technologies. Recent developments in phenomics have been incorporated into almond breeding programs, facilitating the large-scale acquisition of quantitative phenotypes and the dissection of the genetic architecture underlying morphological and quality-related traits. The implementation of a high-throughput phenotyping platform integrating RGB and hyperspectral imaging with genotyping using the 60K almond SNP array enabled the large-scale characterization of almond populations and the identification of 567 robust marker-trait associations across 66 traits. These analyses revealed two major genomic hotspots on chromosomes 2 and 5 associated with morphological and quality-related traits. These regions harbored biologically relevant candidate genes, including genes associated with OVATE family proteins, brassinosteroid signaling, protein ubiquitination, and acyl-CoA metabolism, as well as other regulators of organ growth, cell proliferation, hormone signaling, and seed development. Furthermore, a novel candidate gene encoding a COMT-like O-methyltransferase involved in lignin biosynthesis was identified and proposed to contribute to shell hardness, a major genetically controlled trait in almond. Together, these findings demonstrate the potential of integrating high-throughput phenomics and genomics to dissect complex traits, identify candidate genes, and accelerate genomics-informed breeding in almond.

Long non-coding RNAs (lncRNAs) have emerged as critical players in plant development and stress responses, yet their involvement in pollination responses is largely unknown. To address this gap, we identified and characterized lncRNAs and their cis-acting, trans-acting, and miRNA-mediated regulatory interactions during both compatible and incompatible pollination in Arabidopsis thaliana. Leveraging publicly available datasets, we analyzed expression profiles at 10 and 60 minutes post-pollination. We identified 1,073 novel and 3,422 annotated lncRNAs, with 1,002 novel and 985 annotated, respectively, showing detectable expression after filtering. Differential expression analysis identified 12 lncRNAs at 10 min and 32 lncRNAs at 60 min post-pollination. Further investigation revealed 9 cis-targets, 112 trans-targets, and 144 miRNA-mediated regulatory interactions, many of which were enriched in pathways related to stress, defense, and self-incompatibility. Notably, the regulatory landscape is more active at 60 minutes than at 10 minutes post-pollination. These findings provide a robust framework and resource to facilitate future functional studies of lncRNAs during pollination.

Understanding how gene families evolve can offer great insight into adaptation at the phenotypic and ecological levels. This is particularly true in plants, where transcription factor gene families are often targeted for breeding programs to improve the agronomic traits of economically important crops. While recent advances in next generation sequencing have accelerated the wealth of genomics data, there remains a lack of accessible and reproducible genome mining pipelines tailored for gene family characterisation. Here, we address this gap by developing GENE-FAM, an automated, scalable and open-source pipeline designed to mine and predict gene families based on conserved domains and motifs. To illustrate its application, we apply GENE-FAM to annotate MADS-box transcription factor genes across multiple Cannabis sativa genomes. A comprehensive set of MADS-box genes was identified across three C. sativa cultivars, including both previously annotated and newly predicted genes. Through phylogenetic analyses, we confirm that all type II MADS-box gene subfamilies represented in flowering plants are present in C. sativa. Comparing our annotations with those of Arabidopsis thaliana and Solanum lycopersicum revealed that while most MADS type II families are highly conserved, SEPALLATA-like genes have undergone diversification in C. sativa. Together, these results demonstrate the application of GENE-FAM for genome-wide identification and characterisation of gene families in non-model species, revealing novel insights into MADS-box gene family evolution in C. sativa.

In Asteraceae species, the capitulum is a compact inflorescence, featuring a characteristic reproductive structure. Despite the identification of a few key regulatory factors, the transcriptome-level information on the developing capitulum remains limited. Here, we applied single-cell and spatial transcriptome sequencing to investigate the developing Gerbera hybridas capitulum during floret differentiation. We obtained a transcriptomics atlas encompassing different stages of the Gerbera capitulum and analyzed the cellular and spatial dynamics of gene expression. Using marker gene expression and GO enrichment of cluster-specific DEGs, we annotated putative cell types and described changes in gene expression across sampled stages, potentially associated with ongoing developmental processes. We detected activity of previously undescribed MADS-box genes and defined their spatial expression patterns. Notably, the MADS-box gene GAGL12 was found to be enriched in the putative capitulum phloem cells. The GAGL12 protein was shown in yeast two-hybrid assays to interact with several other MADS-domain proteins with hypothesized functions in vasculature development, and further detailed in silico analyses supported a candidate role in the development of capitulum vasculature. Altogether, we provide integrative and dynamic transcriptomic insight into capitulum and floret development and lay a basis for future functional studies of the control and development of this intriguing reproductive structure.

BackgroundPlant genomes harbour a substantial proportion of dispensable genes - present only in a subset of individuals - that differ from ubiquitously shared core genes in multiple genomic and expression features. While these differences have been repeatedly documented, the factors shaping gene dispensability remain poorly understood. ResultsWe assembled a pan-gene set from eight maize inbred lines from American and European germplasms, together with their transcriptomic profile across 22 tissues/conditions, revealing the genomic and transcriptomic determinants of maize gene dispensability. Multivariate analysis demonstrates that gene expression level and purifying selection - rather than gene size - are the primary factors distinguishing core from dispensable genes. Dispensable genes overlap Helitrons at 4.6 times the rate of core genes, implicating Helitron-mediated gene capture as a major mechanism of dispensable gene formation. Classifying genes into stably expressed, variably expressed, and on-off categories shows that all three classes contain dispensable genes, though in different proportions than for core genes. Contrary to previous assumptions, we show that dispensable genes can participate in basal biological functions just as core genes, and that gene duplication likely provides only a partial mechanism for functional complementation of accessory genes absence. ConclusionsOur results provide novel insights into the molecular and evolutionary factors distinguishing core from dispensable genes and into the biological mechanisms shaping gene dispensability in maize, and demonstrate that classifying genes by transcriptional patterns provides a powerful framework for understanding the biological functions and evolutionary dynamics of both core and dispensable genes.

The transition to flowering in Arabidopsis thaliana is a complex process governed by many biological and environmental stimuli. Although many of the genes which regulate this process have been identified over the past 30 years, it remains unclear how these networks are integrated. In this study, we used the transcriptional responses of Col-0, Ler-1, and three mutant lines, to build a genome wide regulatory network of Arabidopsis thaliana during the flowering transition. The expression profiles of 22,810 genes across five genotypes were collected from the GEO database Series GSE57 from which we assigned flowering-time genes to different interacting modules by an adapted form of Hierarchical Complete Linkage Clustering (HCLC) after reconstruction of regulatory networks according to the Position Weight Matrix (PWM)-based method. Within these modules, we identified 77 core genes and 31 controller or driver genes. We identify two genes, LHY and, less expectedly, the transcription factor TCP16, to be topographically positioned at the regulatory hubs a nine-gene transcriptional control unit, implying they have the capacity to integrate information from across the flowering time pathways which interpret different environmental or endogenous cues during the vegetative-reproductive transition. Interrogating their behaviour across transcriptional datasets, we show that both LHY and TCP16 show transcriptional oscillations during the flowering transition, with a wavelength that varies depending on environmental conditions. We suggest that the transcriptional responses of LHY and TCP16 allow them to regulate the flow of information through the genetic networks which integrates different floral transition cues, and that genetic modelling approaches can provide new insights into the regulation of well-studied biological processes such as the flowering transition. Author summaryHow plants decide when to flower is a critical stage for completing their life cycles. It is also of key agricultural importance, as crops need to flower at the right time of year to allow efficient pollination and harvesting. Many genes are known to affect flowering time control in plants. Here, we use computational approaches to estimate how different genes interact in flowering time control in Arabidopsis, a small plant in the mustard family which is widely used for molecular studies. We use large-scale studies of how gene expression changes in different plant lines which have disrupted or adjusted flowering time to group the many genes involved in flowering into different interacting pathway, which we visualise as sets of coloured nodes controlling one another in a network. We show that two genes may have new rols in integrating information from different pathways, and discuss how their behaviour might help them to function as intregrators of biological information - including the daily oscaillations in their expression.