Dynamic Replacement of Histone H3 Variants Reprograms Epigenetic Marks in Early Mouse Embryos

Upon fertilization, reprogramming of gene expression is required for embryo development. This step is marked by DNA demethylation and changes in histone variant composition. However, little is known about the molecular mechanisms causing these changes and their impact on histone modifications. We examined the global deposition of the DNA replication-dependent histone H3.1 and H3.2 variants and the DNA replication-independent H3.3 variant after fertilization in mice. We showed that H3.3, a euchromatic marker of gene activity, transiently disappears from the maternal genome, suggesting erasure of the oocyte-specific modifications carried by H3.3. After fertilization, H3.2 is incorporated into the transcriptionally silent heterochromatin, whereas H3.1 and H3.3 occupy unusual heterochromatic and euchromatin locations, respectively. After the two-cell stage, H3.1 and H3.3 variants resume their usual respective locations on heterochromatin and euchromatin. Preventing the incorporation of H3.1 and H3.2 by knockdown of the histone chaperone CAF-1 induces a reciprocal increase in H3.3 deposition and impairs heterochromatin formation. We propose that the deposition of different H3 variants influences the functional organization of chromatin. Taken together, these findings suggest that dynamic changes in the deposition of H3 variants are critical for chromatin reorganization during epigenetic reprogramming

Identification and characterization of the two isoforms of the vertebrate H2A.Z histone variant

Histone variants play important roles in the epigenetic regulation of genome function. The histone variant H2A.Z is evolutionarily conserved from yeast to vertebrates, and it has been reported to have multiple effects upon gene expression and insulation, and chromosome segregation. Recently two genes encoding H2A.Z were identified in the vertebrate genome. However, it is not yet clear whether the proteins transcribed from these genes are functionally distinct. To address this issue, we knocked out each gene individually in chicken DT40 cells. We found that two distinct proteins, H2A.Z-1 and H2A.Z-2, were produced from these genes, and that these proteins could be separated on a long SDS–PAGE gel. The two isoforms were deposited to a similar extent by the SRCAP chromatin-remodeling complex, suggesting redundancy to their function. However, cells lacking either one of the two isoforms exhibited distinct alterations in cell growth and gene expression, suggesting that the two isoforms have differential effects upon nucleosome stability and chromatin structure. These findings provide insight into the molecular basis of the multiple functions of the H2A.Z gene products

OsPIE1, the Rice Ortholog of Arabidopsis PHOTOPERIOD-INDEPENDENT EARLY FLOWERING1, Is Essential for Embryo Development

orthologs in monocots remains unknown., respectively).Taken together, our results suggest that OsPIE1 is the rice ortholog of Arabidopsis PIE1 and plays an essential role in rice embryo development

Heterochromatin Protein 1β (HP1β) has distinct functions and distinct nuclear distribution in pluripotent versus differentiated cells

Background: Pluripotent embryonic stem cells (ESCs) have the unique ability to differentiate into every cell type and to self-renew. These characteristics correlate with a distinct nuclear architecture, epigenetic signatures enriched for active chromatin marks and hyperdynamic binding of structural chromatin proteins. Recently, several chromatin-related proteins have been shown to regulate ESC pluripotency and/or differentiation, yet the role of the major heterochromatin proteins in pluripotency is unknown. Results: Here we identify Heterochromatin Protein 1β (HP1β) as an essential protein for proper differentiation, and, unexpectedly, for the maintenance of pluripotency in ESCs. In pluripotent and differentiated cells HP1β is differentially localized and differentially associated with chromatin. Deletion of HP1β, but not HP1aα, in ESCs provokes a loss of the morphological and proliferative characteristics of embryonic pluripotent cells, reduces expression of pluripotency factors and causes aberrant differentiation. However, in differentiated cells, loss of HP1β has the opposite effect, perturbing maintenance of the differentiation state and facilitating reprogramming to an induced pluripotent state. Microscopy, biochemical fractionation and chromatin immunoprecipitation reveal a diffuse nucleoplasmic distribution, weak association with chromatin and high expression levels for HP1β in ESCs. The minor fraction of HP1β that is chromatin-bound in ESCs is enriched within exons, unlike the situation in differentiated cells, where it binds heterochromatic satellite repeats and chromocenters. Conclusions: We demonstrate an unexpected duality in the role of HP1β: it is essential in ESCs for maintaining pluripotency, while it is required for proper differentiation in differentiated cells. Thus, HP1β function both depends on, and regulates, the pluripotent state

Heterochromatin and the molecular mechanisms of 'parent-of-origin' effects in animals.

Twenty five years ago it was proposed that conserved components of constitutive heterochromatin assemble heterochromatinlike complexes in euchromatin and this could provide a general mechanism for regulating heritable (cell-to-cell) changes in gene expressibility. As a special case, differences in the assembly of heterochromatin-like complexes on homologous chromosomes might also regulate the parent-of-origin-dependent gene expression observed in placental mammals. Here, the progress made in the intervening period with emphasis on the role of heterochromatin and heterochromatin-like complexes in parent-of-origin effects in animals is reviewed

Regulation of transcription and chromatin organisation in the oocyte and the early mouse embryo

La fécondation d'un ovocyte en métaphase II par un spermatozoïde, deux cellules différenciées, induit la formation d'un zygote, une cellule totipotente à l'origine des tissus embryonnaires et extraembryonnaires d'un organisme. Dans une première partie deFertilisation of a metaphase II oocyte by the sperm, two differentiated cells, leads to the formation of a zygote, a totipotent cell capable of generating a new organism. In the first part of my thesis, I showed that TBP2 (TATA binding protein 2), an ooc

Regulation of transcription and chromatin organisation in the oocyte and the early mouse embryo

La fécondation d'un ovocyte en métaphase II par un spermatozoïde, deux cellules différenciées, induit la formation d'un zygote, une cellule totipotente à l'origine des tissus embryonnaires et extraembryonnaires d'un organisme. Dans une première partie de ma thèse, j ai montré que la protéine TBP2 (TATA-binding protein 2), un facteur de transcription général spécifique des ovocytes chez la souris, est nécessaire au développement et à la maturation des ovocytes. TBP2 intervient dans l'établissement d'un programme d'expression génique spécifique des ovocytes et lie le promoteur de certains gènes activement transcrits. Dans la deuxième partie de ma thèse, j ai abordé le phénomène d'établissement de l'hétérochromatine péricentrique dans le pronoyau paternel au sein du zygote. J ai montré que l'hétérochromatinisation des régions péricentriques nécessite l'intervention d'au moins trois acteurs : des modifications post-traductionnelles de H3.3K27, le recrutement de HP1B et d'un ARN double brin transcrit à partir des major satellites. Des résultats préliminaires suggèrent qu une localisation correcte de ces régions après fécondation est nécessaire au développement préimplantatoire de la souris. Les facteurs de transcription de base et les modifications chromatiniennes contribuent donc à l'établissement de programmes d'expression génique spécifiques et à la formation de structures chromatiniennes nécessaires au maintien de l'intégrité du génome. Dans l'ovocyte et le zygote de souris, l'évolution des programmes d'expression génique et des modifications chromatiniennes sont le reflet des reprogrammations qui caractérisent le passage de cellules différenciées à une cellule totipotente.Fertilisation of a metaphase II oocyte by the sperm, two differentiated cells, leads to the formation of a zygote, a totipotent cell capable of generating a new organism. In the first part of my thesis, I showed that TBP2 (TATA binding protein 2), an oocyte specific general transcription factor in the mouse, is necessary for oocyte development and maturation during folliculogenesis. TBP2 is necessary for the establishment of an oocyte specific expression pattern, and functions at least in part, through its binding to promoters of actively transcribed genes. In the second part of my thesis, I focused on the establishment of pericentric heterochromatin in the paternal pronucleus in zygotes. I showed that three main actors are responsible for heterochromatinisation of pericentric regions : post-translational modifications of H3.3K27, recruitment of HP1B and of a double stranded RNA transcribed from the major satellites repeats. Some preliminary results also show that the intranuclear localization of pericentric regions after fertilization is crucial for early embryonic development. Basal transcription factors and chromatin modifications are involved in the establishment of specific gene expression pattern and in the formation of chromatin structures necessary to maintain genome integrity. In the mouse oocyte, sperm and zygote, the evolution of specific gene expression pattern and of chromatin modifications are directed by both genetic information and epigenetic reprogramming in order to generate a totipotent cell