TRAUCO, a Trithorax-group gene homologue, is required for early embryogenesis in Arabidopsis thaliana.
ABSTRACT: Embryogenesis is a critical stage during the plant life cycle in which a unicellular zygote develops into a multicellular organism. Co-ordinated gene expression is thus necessary for proper embryo development. Polycomb and Trithorax group genes are members of evolutionarily conserved machinery that maintains the correct expression patterns of key developmental regulators by repressing and activating gene transcription. TRAUCO (TRO), a gene homologous to the Trithorax group of genes that can functionally complement a BRE2P yeast mutant, has been identified in Arabidopsis thaliana. It is demonstrated that TRO is a nuclear gene product expressed during embryogenesis, and loss of TRO function leads to impaired early embryo development. Embryos that arrested at the globular stage in the tro-1 mutant allele were fully rescued by a TRO expression clone, a demonstration that the tro-1 mutation is a true loss-of-function in TRO. Our data have established that TRO is the first trithorax-group gene homologue in plants that is required for early embryogenesis.
Project description:In Drosophila, the maintenance of developmentally important transcription patterns is controlled at the level of chromatin structure. The Polycomb group (PcG) and trithorax group (trxG) genes encode proteins involved in chromatin remodelling. PcG genes have been proposed to act by packaging transcriptional repressed chromosomal domains into condensed heterochromatin-like structures. Some of the trxG proteins characterized so far are members of chromatin opening complexes (e.g. SWI/SNF and GAGA/NURF) which facilitate binding of transcription factors and components of the basal transcriptional machinery. Genetic and biochemical data suggest that these two groups of regulatory factors may act through a common set of DNA elements. In the present study, we have investigated the binding of Trithorax (TRX) and Polycomb (PC) protein in the bithorax complex (BX-C) during embryogenesis. In addition, we have identified the minimal fragments from the Ultrabithorax (Ubx) regulatory region that are capable of recruiting TRX to chromosomal sites containing them. Comparative analysis of the binding of the two proteins shows that TRX and PC bind target sequences (PcG-regulated elements, PREs) by cellular blastoderm, when BX-C transcription begins. At the same stage, TRX but not PC is strongly associated with core promoters. Later, at germ band extension, the time of derepression in Polycomb mutants, PC binding is also detected outside core PREs and additionally binds to the fragments containing promoters.
Project description:WUSCHEL-related homeobox (WOX) gene is a plant-specific clade of homeobox transcription factors. Increasing evidences reveal that WOXs play critical roles in early embryogenesis, which involves zygote development, initiation of zygote division, and apical or basal cell lineage establishment. However, how WOXs regulate these developmental events remains largely unknown, and even detailed expression pattern in gametes and early proembryos is not yet available. Here, 13 WOX family genes were identified in Nicotiana tabacum genome. Comparative analysis of 13 WOX family genes with their homologs in Arabidopsis thaliana reveals relatively conserved expression pattern of WUS and WOX5 in shoot/root apical meristem. Whereas variations were also found, e.g., lacking homolog of WOX8 (a marker for suspensor cell) in tobacco genome and the expression of WOX2/WOX9 in both apical cell and basal cell. Transient transcriptional activity analysis revealed that WOXs in WUS clade have repressive activities for their target's transcription, whereas WOXs in ancient and intermediate clade have activation activities, giving a molecular basis for the phylogenetic classification of tobacco WOXs into three major clades. Expression pattern analysis revealed that some WOXs (e.g., WOX 13a) expressed in both male and female gametes and some WOXs (e.g., WOX 11 and WOX 13b) displayed the characteristics of parent-of-origin genes. Interestingly, some WOXs (e.g., WOX2 and WOX9), which are essential for early embryo patterning, were de novo transcribed in zygote, indicating relevant mechanism for embryo pattern formation is only established in zygote right after fertilization and not carried in by gametes. We also found that most WOXs displayed a stage-specific and cell type-specific expression pattern. Taken together, this work provides a detailed landscape of WOXs in tobacco during fertilization and early embryogenesis, which will facilitate the understanding of their specific roles in these critical developmental processes of embryogenesis.
Project description:The fusogenic lipid diacylglycerol is essential for remodeling gamete and zygote nuclear envelopes (NE) during early embryogenesis. It is unclear whether upstream signaling molecules are likewise conserved. Here we demonstrate PLC? and its activator SFK1, which co-operate during male pronuclear envelope formation, also promote the subsequent male and female pronuclear fusion. PLC? and SFK1 interact directly at the fusion site leading to PLC? activation. This is accompanied by a spatially restricted reduction of PtdIns(4,5)P2. Consequently, pronuclear fusion is blocked by PLC? or SFK1 inhibition. These findings identify new regulators of events in the early embryo and suggest a conserved "toolkit" of fusion machinery drives successive NE fusion events during embryogenesis.
Project description:BACKGROUND: In animals, early embryonic development is largely dependent on maternal transcripts synthesized during gametogenesis. However, in higher plants, the extent of maternal control over zygote development and early embryogenesis is not fully understood yet. Nothing is known about the activity of the parental genomes during seed formation of interspecies hybrids. METHODOLOGY/PRINCIPAL FINDINGS: Here, we report that an interspecies hybridization system between SR1 (Nicotiana tabacum) and Hamayan (N. rustica) has been successfully established. Based on the system we selected 58 genes that have polymorphic sites between SR1 and Hamayan, and analyzed the allele-specific expression of 28 genes in their hybrid zygotes (Hamayan x SR1). Finally the allele-specific expressions of 8 genes in hybrid zygotes were repeatedly confirmed. Among them, 4 genes were of paternal origin, 1 gene was of maternal origin and 3 genes were of biparental origin. These results revealed obvious biparental involvement and differentially contribution of parental-origin genes to zygote development in the interspecies hybrid. We further detected the expression pattern of the genes at 8-celled embryo stage found that the involvement of the parental-origin genes may change at different stages of embryogenesis. CONCLUSIONS/SIGNIFICANCE: We reveal that genes of both parental origins are differentially involved in early embryogenesis of a tobacco interspecies hybrid and functions in a developmental stage-dependent manner. This finding may open a window to seek for the possible molecular mechanism of hybrid vigor.
Project description:Trithorax (TRX) and ASH1 belong to the trithorax group (trxG) of transcriptional activator proteins, which maintains homeotic gene expression during Drosophila development. TRX and ASH1 are localized on chromosomes and share several homologous domains with other chromatin-associated proteins, including a highly conserved SET domain and PHD fingers. Based on genetic interactions between trx and ash1 and our previous observation that association of the TRX protein with polytene chromosomes is ash1 dependent, we investigated the possibility of a physical linkage between the two proteins. We found that the endogenous TRX and ASH1 proteins coimmunoprecipitate from embryonic extracts and colocalize on salivary gland polytene chromosomes. Furthermore, we demonstrated that TRX and ASH1 bind in vivo to a relatively small (4 kb) bxd subregion of the homeotic gene Ultrabithorax (Ubx), which contains several trx response elements. Analysis of the effects of ash1 mutations on the activity of this regulatory region indicates that it also contains ash1 response element(s). This suggests that ASH1 and TRX act on Ubx in relatively close proximity to each other. Finally, TRX and ASH1 appear to interact directly through their conserved SET domains, based on binding assays in vitro and in yeast and on coimmunoprecipitation assays with embryo extracts. Collectively, these results suggest that TRX and ASH1 are components that interact either within trxG protein complexes or between complexes that act in close proximity on regulatory DNA to maintain Ubx transcription.
Project description:The transcription factor absent, small, or homeotic discs 2 (ash2) gene is a member of the trithorax group of positive regulators of homeotic genes. Mutant alleles for ash2 are larvalpupal lethals and display imaginal disc and brain abnormalities. The allele used in this study is a true mutant for the trithorax function and lacks the longest transcript present in wild-type flies. In an attempt to identify gene targets of ash2, we have performed an expression analysis by using cDNA microarrays. Genes involved in cell cycle, cell proliferation, and cell adhesion are among these targets, and some of them are validated by functional and expression studies. Even though trithorax proteins act by modulating chromatin structure at particular chromosomal locations, evidence of physical aggregation of ash2-regulated genes has not been found. This work represents the first microarray analysis, to our knowledge, of a trithorax-group gene.
Project description:Small molecules inhibitors are powerful tools for studying multiple aspects of cell biology and stand at the forefront of drug discovery pipelines. However, in the early Caenorhabditis elegans (C. elegans) embryo, which is a powerful model system for cell and developmental biology, the use of small molecule inhibitors has been limited by the impermeability of the embryonic eggshell, the low-throughput manual embryo isolation methods, and the lack of well-controlled drug delivery protocols. This work reports a fully integrated microfluidic approach for studies of C. elegans early embryogenesis, including the possibility of testing small molecule inhibitors with increased throughput and versatility. The setup enables robust on-chip extraction of embryos from gravid adult worms in a dedicated pillar array chamber by mechanical compression, followed by rapid fluidic transfer of embryos into an adjacent microtrap array. Parallel analysis of ?100 embryos by high-resolution time-lapse imaging from the one-cell stage zygote until hatching can be performed with this device. The implementation of versatile microfluidic protocols, in particular time-controlled and reversible drug delivery to on-chip immobilized embryos, demonstrates the potential of the device for biochemical and pharmacological assays.
Project description:Seeds are essential for human civilization, so understanding the molecular events underpinning seed development and the zygotic embryo it contains is important. In addition, the approach of somatic embryogenesis is a critical propagation and regeneration strategy to increase desirable genotypes, to develop new genetically modified plants to meet agricultural challenges, and at a basic science level, to test gene function. We briefly review some of the transcription factors (TFs) involved in establishing primary and apical meristems during zygotic embryogenesis, as well as TFs necessary and/or sufficient to drive somatic embryo programs. We focus on the model plant Arabidopsis for which many tools are available, and review as well as speculate about comparisons and contrasts between zygotic and somatic embryo processes.
Project description:Polycomb group (PcG) and Trithorax group (TrxG) genes encode important regulators of development and differentiation in metazoans. These two groups of genes were discovered in Drosophila by their opposing effects on homeotic gene (Hox) expression. PcG genes collectively behave as genetic repressors of Hox genes, while the TrxG genes are necessary for HOX gene expression or function. Biochemical studies showed that many PcG proteins are present in two protein complexes, Polycomb repressive complexes 1 and 2, which repress transcription via chromatin modifications. TrxG proteins activate transcription via a variety of mechanisms. Here we summarize the large body of genetic and biochemical experiments in Drosophila on these two important groups of genes.
Project description:Asymmetric division of zygote is critical for pattern formation during early embryogenesis in plants and animals. It requires integration of the intrinsic and extrinsic cues prior to and/or after fertilization. How these cues are translated into developmental signals is poorly understood. Here through genetic screen for mutations affecting early embryogenesis, we identified an Arabidopsis mutant, zygotic arrest 1 (zar1), in which zygote asymmetric division and the cell fate of its daughter cells were impaired. ZAR1 encodes a member of the RLK/Pelle kinase family. We demonstrated that ZAR1 physically interacts with Calmodulin and the heterotrimeric G protein G?, and ZAR1 kinase is activated by their binding as well. ZAR1 is specifically expressed micropylarly in the embryo sac at eight-nucleate stage and then in central cell, egg cell and synergids in the mature embryo sac. After fertilization, ZAR1 is accumulated in zygote and endosperm. The disruption of ZAR1 and AGB1 results in short basal cell and an apical cell with basal cell fate. These data suggest that ZAR1 functions as a membrane integrator for extrinsic cues, Ca2+ signal and G protein signaling to regulate the division of zygote and the cell fate of its daughter cells in Arabidopsis.