Project description:Heterochromatin de-condensation in companion gametic cells is conserved in both plants and animals. In plants, microspore undergoes asymmetric pollen mitosis (PMI) to produce a vegetative cell (VC) and a generative cell (GC). Subsequently, the GC undergoes pollen mitosis (PMII) to produce two sperm cells (SC). Consistent with heterochromatin de-condensation in the VC, H3K9me2, a heterochromatin mark, is barely detected in VC. However, how H3K9me2 is differentially regulated during pollen mitosis remains unclear. Here, we show that H3K9me2 is gradually evicted from the VC since PMI but remain unchanged in the GC and SC. ARID1, a pollen-specific transcription factor that facilitates PMII, promotes H3K9me2 maintenance in the GC/SC but slows down its eviction in the VC. The genomic targets of ARID1 mostly overlaps with H3K9me2 loci, and ARID1 recruits H3K9 methyltransferase SUVH6. Our results uncover that differential pattern of H3K9me2 between two cell types is regulated by ARID1 during pollen mitosis.
Project description:In plants, sperm cell formation involves two rounds of pollen mitoses, in which the microspore initiates the first pollen mitosis (PMI) to produce a vegetative cell and a generative cell, then the generative cell continues the second mitosis (PMII) to produce two sperm cells. DUO1, a R2R3 Myb transcription factor, is activated in the generative cell to promote S-G2/M transition during PMII. Loss-of-function of DUO1 caused a complete arrest of PMII. Despite the importance of DUO1, how DUO1 is regulated is largely unexplored. We previously demonstrated that ARID1, an ARID transcription factor, stimulates DUO1 transcription. Here, we show that cell proliferation suppressor RBR1 interacts with ARID1 to stabilize DUO1. While the C-terminus of RBR1 is dispensable for vegetative growth, it plays a crucial role in reproductive development and facilitates interaction with ARID1. Moreover, DUO1 is a short-lived protein, ARID1 promotes the RBR1-DUO1 interaction, and RBR1 stabilizes DUO1 in a proteasome-dependent manner. Thus, RBR1 promotes DUO1-dependent PMII progression via antagonizing its repressive role in the cell cycle factors CDKA;1 and CYCB1;1. Collectively, we uncover that ARID1 and RBR1 act in concert to regulate DUO1 at both the transcriptional and post-transcriptional levels, balancing cell specification and cell division.
Project description:Seed development is dependent on a well-orchestrated interplay between different transcriptional programs operating in the embryo, the endosperm and the maternally derived seed coat. In angiosperms, the embryo and the endosperm are products of double fertilization during which the two pollen sperm cells fuse with the egg cell and the central cell of the female gametophyte. In Arabidopsis, mutation of the cell cycle regulator CYCLIN DEPENDENT KINASE A;1 (CKDA;1) results in pollen that only successfully fertilizes the egg cell. Seeds generated from crosses with cdka;1 pollen develop endosperm with solely maternal and no paternal contribution. Here we have exploited cdka;1 fertilization as a novel tool for genomic dissection of parental effects during seed development. We have generated genome-wide transcription profiles of cdka;1 fertilized seeds. By this approach, we identified 11 differentially expressed AGAMOUS-LIKE (AGL) genes encoding Type-I MADS-box transcription factors. Here, AGL36 was chosen for an in-depth study. We show that AGL36 is imprinted and only expressed from the maternal genome. In addition, we demonstrate that AGL36 imprinting is controlled by the activity of METHYLTRANSFERASE1 (MET1) maintenance DNA methyltransferase and DEMETER (DME) DNA glycosylase. Interestingly, our data also show that the active maternal allele of AGL36 is regulated throughout endosperm development by components of the FIS Polycomb Repressive Complex 2 (PRC2). These findings shed novel light on the interplay between maternal and paternal genomes in the seed and how imprinting is coordinated by different mechanisms.
Project description:Seed development is dependent on a well-orchestrated interplay between different transcriptional programs operating in the embryo, the endosperm and the maternally derived seed coat. In angiosperms, the embryo and the endosperm are products of double fertilization during which the two pollen sperm cells fuse with the egg cell and the central cell of the female gametophyte. In Arabidopsis, mutation of the cell cycle regulator CYCLIN DEPENDENT KINASE A;1 (CKDA;1) results in pollen that only successfully fertilizes the egg cell. Seeds generated from crosses with cdka;1 pollen develop endosperm with solely maternal and no paternal contribution. Here we have exploited cdka;1 fertilization as a novel tool for genomic dissection of parental effects during seed development. We have generated genome-wide transcription profiles of cdka;1 fertilized seeds. By this approach, we identified 11 differentially expressed AGAMOUS-LIKE (AGL) genes encoding Type-I MADS-box transcription factors. Here, AGL36 was chosen for an in-depth study. We show that AGL36 is imprinted and only expressed from the maternal genome. In addition, we demonstrate that AGL36 imprinting is controlled by the activity of METHYLTRANSFERASE1 (MET1) maintenance DNA methyltransferase and DEMETER (DME) DNA glycosylase. Interestingly, our data also show that the active maternal allele of AGL36 is regulated throughout endosperm development by components of the FIS Polycomb Repressive Complex 2 (PRC2). These findings shed novel light on the interplay between maternal and paternal genomes in the seed and how imprinting is coordinated by different mechanisms. 3 biological replicates, each consisting of 35 siliques from 10 individual plants, were used. Arabidopsis thaliana wild type (ecotype Colombia-0 and Ler-0) and cdka;1 mutant (At3g48750 /SALK_109806) were sown out on soil and grown under the following conditions: 18oC day and 18oC night, 16 hr day length with 30 min adjustment of light to on and off, and 85 μmol/m2/sek in light intensity. For each replica, 10 Ler plants were emasculated and pollinated after 48 hours, either with individual WT Col or with individual cdka;1 mutant pollen. Three days after pollination (3 DAP), the siliques of each biological replicate were dissected with hypodermic needles to remove carpel walls, the pistil with pollen attached to it and the pedicel with abcsission zone. The remaining seeds attached to the middle lamella were harvested in bulk in liquid nitrogen at the same time of the day. Biological replicas are dye-swapped between slides.
Project description:Pollen development from the microspore involves a series of coordinated cellular events, and the resultant mature pollen is specialized in function that it can quickly germinate and produces a polar-growth pollen tube derived from the vegetative cell to deliver two sperms for fertilization. Understanding the molecular program underlying pollen development and germination still remains a major challenge for plant biology. We used Affymetrix GeneChip Rice Genome Array to comprehensively analyzed the dynamic changes in the transcriptomes of rice pollen at five sequential developmental stages from microspores to germinated pollen. Among the 51,279 transcripts on the array, we found 25,062 pollen-preferential transcripts, among which 2,203 were development stage-enriched. The diversity of transcripts decreased greatly from microspores to mature and germinated pollen, whereas the number of stage-enriched transcripts displayed a U-type change, with the lowest at the bicellular pollen stage; and a transition of overrepresented stage-enriched transcript groups associated with different functional categories, which indicates a shift in gene expression program at the bicellular pollen stage. About 54% of the now-annotated rice F-box protein genes were expressed preferentially in pollen. The transcriptome profile of germinated pollen was significantly and positively correlated with that of mature pollen. Analysis of expression profiles and coexpressed features of the pollen-preferential transcripts related to cell cycle, transcription, the ubiquitin/26S proteasome system, phytohormone signalling, the kinase system and defense/stress response revealed five expression patterns, which are compatible with changes in major cellular events during pollen development and germination. A comparison of pollen transcriptomes between rice and Arabidopsis revealed that 56.6% of the rice pollen preferential genes had homologs in Arabidopsis genome, but 63.4% of these homologs were expressed, with a small proportion being expressed preferentially, in Arabidopsis pollen. Rice and Arabidopsis pollen had non-conservative transcription factors each. These results supply novel insights into the molecular program and key components of the regulatory network regulating pollen development and germination. KEYWORDS: rice (Oryza sativa L.), pollination and fertilization, stigma, molecular functions, signaling, microarray, stress response
Project description:Epigenetic marks are reprogrammed in the gametes to reset genomic potential in the next generation. In mammals, paternal chromatin is extensively reprogrammed through the global erasure of DNA methylation and the exchange of histones with protamines1,2. Precisely how the paternal epigenome is reprogrammed in flowering plants remains unclear since DNA is not demethylated in sperm and histones are retained3,4. Here, we describe a multi-layered mechanism by which H3K27me3 is globally lost from histone-based sperm chromatin in Arabidopsis. This mechanism involves silencing of H3K27me3 ‘writers’, the activity of H3K27me3 ‘erasers’ and deposition of a sperm-specific histone, H3.105, which we show is immune to lysine 27 methylation. The loss of H3K27me3 facilitates transcription of genes essential for spermatogenesis and pre-configures sperm with a chromatin state that forecasts gene expression in the next generation. Thus, plants have evolved a specific mechanism to simultaneously differentiate male gametes and reprogram the paternal epigenome.
Project description:Epigenetic marks are reprogrammed in the gametes to reset genomic potential in the next generation. In mammals, paternal chromatin is extensively reprogrammed through the global erasure of DNA methylation and the exchange of histones with protamines1,2. Precisely how the paternal epigenome is reprogrammed in flowering plants remains unclear since DNA is not demethylated in sperm and histones are retained3,4. Here, we describe a multi-layered mechanism by which H3K27me3 is globally lost from histone-based sperm chromatin in Arabidopsis. This mechanism involves silencing of H3K27me3 ‘writers’, the activity of H3K27me3 ‘erasers’ and deposition of a sperm-specific histone, H3.105, which we show is immune to lysine 27 methylation. The loss of H3K27me3 facilitates transcription of genes essential for spermatogenesis and pre-configures sperm with a chromatin state that forecasts gene expression in the next generation. Thus, plants have evolved a specific mechanism to simultaneously differentiate male gametes and reprogram the paternal epigenome.
Project description:Pollen is the male gametophyte of land plants. Proper development and maturation of pollen is necessary for the successful reproduction of seed plants. This process involves sophisticated coordination between sporophytic and gametophytic tissues in anthers. To advance the mechanistic studies of anther development, additional players need to be discovered for a comprehensive understanding of the underlying regulatory network. Here we show that the Arabidopsis dual specificity tyrosine phophorylated and regulated kinase (DRYK), AtYAK1, is essential for development of rosette leaves and the male but not female gametophyte in Arabidopsis. Arabidopsis mutant plants carrying a mutation in AtYAK1 produce developmentally stalled microspores, likely because of the defects in the two consecutive mitosis steps in the post-meiotic maturation process of pollen. The mutation of AtYAK1 has a significant effect on gene expression programs in developing pollen. Transcritpome analysis of atyak1 revealed downstream genes in families of protein kinases, transporters and transcription factors, which potentially contribute to pollen development. This study represents the first molecular characterization of DYRK in the plant kingdom. Our results also imply that the regulation of cytokinesis by DYRKs is evolutionally conserved in fungus, fruit fly, animals and plants.