Tables S1 to S3, S5, S6, and S9 as zipped Excel files

The development of multicellular organisms is controlled by differential gene expression whereby cells adopt distinct fates. A spatially resolved view of gene expression allows the elucidation of transcriptional networks that are linked to cellular identity and function. The haploid female gametophyte of flowering plants is a highly reduced organism: at maturity, it often consists of as few as three cell types derived from a common precursor [1, 2]. However, because of its inaccessibility and small size, we know little about the molecular basis of cell specification and differentiation in the female gametophyte. Here we report expression profiles of all cell types in the mature Arabidopsis female gametophyte. Differentially expressed posttranscriptional regulatory modules and metabolic pathways characterize the distinct cell types. Several transcription factor families are overrepresented in the female gametophyte in comparison to other plant tissues, e.g., type I MADS domain, RWP-RK, and reproductive meristem transcription factors. PAZ/Piwi-domain encoding genes are upregulated in the egg, indicating a role of epigenetic regulation through small RNA pathways-a feature paralleled in the germline of animals [3]. A comparison of human and Arabidopsis egg cells for enrichment of functional groups identified several similarities that may represent a consequence of coevolution or ancestral gametic features.

The data presented in this study indicate that closely related miRNA family members that are predicted to target the same set of genes can have different functions during development, possibly because of nonoverlapping expression patterns.

In plants, members of microRNA (miRNA) families are often predicted to target the same or overlapping sets of genes. It has thus been hypothesized that these miRNAs may act in a functionally redundant manner. This hypothesis is tested here by studying the effects of elimination of all three members of the MIR164 family from Arabidopsis. It was found that a loss of miR164 activity leads to a severe disruption of shoot development, in contrast to the effect of mutation in any single MIR164 gene. This indicates that these miRNAs are indeed functionally redundant. Differences in the expression patterns of the individual MIR164 genes imply, however, that redundancy among them is not complete, and that these miRNAs show functional specialization. Furthermore, the results of molecular and genetic analyses of miR164-mediated target regulation indicate that miR164 miRNAs function to control the transcript levels, as well as the expression patterns, of their targets, suggesting that they might contribute to developmental robustness. For two of the miR164 targets, namely CUP-SHAPED COTYLEDON1 (CUC1) and CUC2, we provide evidence for their involvement in the regulation of growth and show that their derepression in miR164 loss-of-function mutants is likely to account for most of the mutant phenotype.

The Arabidopsis homeotic gene AGAMOUS (AG) is necessary for the specification of reproductive organs (stamens and carpels) during the early steps of flower development. AG encodes a transcription factor of the MADS-box family that is expressed in stamen and carpel primordia. At later stages of development, AG is expressed in distinct regions of the reproductive organs. This suggests that AG might function during the maturation of stamens and carpels, as well as in their early development. However, the developmental processes that AG might control during organogenesis and the genes that are regulated by this factor are largely unknown. Here we show that microsporogenesis, the process leading to pollen formation, is induced by AG through activation of the SPOROCYTELESS gene (SPL, also known as NOZZLE,NZZ), a regulator of sporogenesis. Furthermore, we demonstrate that SPL can induce microsporogenesis in the absence of AG function, suggesting that AG controls a specific process during organogenesis by activating another regulator that performs a subset of its functions.

How different organs are formed from small sets of undifferentiated precursor cells is a key question in developmental biology. To understand the molecular mechanisms underlying organ specification in plants, we studied the function of the homeotic selector genes APETALA3 (AP3) and PISTILLATA (PI), which control the formation of petals and stamens during Arabidopsis flower development. To this end, we characterized the activities of the transcription factors that AP3 and PI encode throughout flower development by using perturbation assays as well as transcript profiling and genomewide localization studies, in combination with a floral induction system that allows a stage-specific analysis of flower development by genomic technologies. We discovered considerable spatial and temporal differences in the requirement for AP3/PI activity during flower formation and show that they control different sets of genes at distinct phases of flower development. The genomewide identification of target genes revealed that AP3/PI act as bifunctional transcription factors: they activate genes involved in the control of numerous developmental processes required for organogenesis and repress key regulators of carpel formation. Our results imply considerable changes in the composition and topology of the gene network controlled by AP3/PI during the course of flower development. We discuss our results in light of a model for the mechanism underlying sex-determination in seed plants, in which AP3/PI orthologues might act as a switch between the activation of male and the repression of female development.F lowers are typically composed of four organ types, which are disposed in four floral whorls. From the outside of the flower to the center, they are sepals, petals, stamens, and carpels (the subunits of the gynoecium). The developmental fate of these different types of organs is specified by a small number of floral organ identity genes. The pivotal role of these genes was uncovered through the analysis of mutants that form flowers with homeotic transformations, i.e., the replacement of one type of organ with another (1-4). Based on the morphological defects of the individual mutants and their genetic interactions, it was proposed that the floral organ identity genes act in a combinatorial manner and have distinct functions during flower development, with the so-called A function genes being required for the formation of sepals and petals, B function genes for petal and stamen development, and C function genes for the formation of stamens and carpels. This well-established ABC model of floral organ identity specification (5) has provided, since its introduction more than 20 y ago, an invaluable framework for the analysis of the genetic mechanisms underlying the formation and evolution of flowers.Molecular characterization of the floral organ identity genes in different species revealed that they encode transcription factors and belong, with few exceptions, to the family of MADS domain proteins (1-3). The floral organ identity factors w...

BackgroundDevelopment of eukaryotic organisms is controlled by transcription factors that trigger specific and global changes in gene expression programs. In plants, MADS-domain transcription factors act as master regulators of developmental switches and organ specification. However, the mechanisms by which these factors dynamically regulate the expression of their target genes at different developmental stages are still poorly understood.ResultsWe characterized the relationship of chromatin accessibility, gene expression, and DNA binding of two MADS-domain proteins at different stages of Arabidopsis flower development. Dynamic changes in APETALA1 and SEPALLATA3 DNA binding correlated with changes in gene expression, and many of the target genes could be associated with the developmental stage in which they are transcriptionally controlled. We also observe dynamic changes in chromatin accessibility during flower development. Remarkably, DNA binding of APETALA1 and SEPALLATA3 is largely independent of the accessibility status of their binding regions and it can precede increases in DNA accessibility. These results suggest that APETALA1 and SEPALLATA3 may modulate chromatin accessibility, thereby facilitating access of other transcriptional regulators to their target genes.ConclusionsOur findings indicate that different homeotic factors regulate partly overlapping, yet also distinctive sets of target genes in a partly stage-specific fashion. By combining the information from DNA-binding and gene expression data, we are able to propose models of stage-specific regulatory interactions, thereby addressing dynamics of regulatory networks throughout flower development. Furthermore, MADS-domain TFs may regulate gene expression by alternative strategies, one of which is modulation of chromatin accessibility.

No abstract