Gotowa bibliografia na temat „Polycomb machinery”

Polycomb transcriptional silencing machinery is implicated in the maintenance of precursor fates, but how this repression is reversed to allow cell differentiation is unknown. Here we show that testis-specific TAF (TBP-associated factor) homologs required for terminal differentiation of male germ cells may activate target gene expression in part by counteracting repression by Polycomb. Chromatin immunoprecipitation revealed that testis TAFs bind to target promoters, reduce Polycomb binding, and promote local accumulation of H3K4me3, a mark of Trithorax action. Testis TAFs also promoted relocalization of Polycomb Repression Complex 1 components to the nucleolus in spermatocytes, implicating subnuclear architecture in the regulation of terminal differentiation.

Eukaryotic gene expression is activated by factors that interact within complex machinery to initiate transcription. An important component of this machinery is the DNA repair/transcription factor TFIIH. Mutations in TFIIH result in three human syndromes: xeroderma pigmentosum, Cockayne syndrome and trichothiodystrophy. Transcription and DNA repair defects have been linked to some clinical features of these syndromes. However, how mutations in TFIIH affect specific developmental programmes, allowing organisms to develop with particular phenotypes, is not well understood. Here, we show that mutations in the p52 and p8 subunits of TFIIH have a moderate effect on the gene expression programme in the Drosophila testis, causing germ cell differentiation arrest in meiosis, but no Polycomb enrichment at the promoter of the affected differentiation genes, supporting recent data that disagree with the current Polycomb-mediated repression model for regulating gene expression in the testis. Moreover, we found that TFIIH stability is not compromised in p8 subunit-depleted testes that show transcriptional defects, highlighting the role of p8 in transcription. Therefore, this study reveals how defects in TFIIH affect a specific cell differentiation programme and contributes to understanding the specific syndrome manifestations in TFIIH-afflicted patients.

Epigenetic mechanisms, including DNA and histone modifications, are pivotal for normal brain development and functions by modulating spatial and temporal gene expression. Dysregulation of the epigenetic machinery can serve as a causal role in numerous brain disorders. Proper mammalian brain development and functions depend on the precise expression of neuronal-specific genes, transcription factors and epigenetic modifications. Antagonistic polycomb and trithorax proteins form multimeric complexes and play important roles in these processes by epigenetically controlling gene repression or activation through various molecular mechanisms. Aberrant expression or disruption of either protein group can contribute to neurodegenerative diseases. This review focus on the current progress of Polycomb and Trithorax complexes in brain development and disease, and provides a future outlook of the field.

Populations of resident stem cells (SCs) are responsible for maintaining, repairing, and regenerating adult tissues. In addition to having the capacity to generate all the differentiated cell types of the tissue, adult SCs undergo long periods of quiescence within the niche to maintain themselves. The process of SC renewal and differentiation is tightly regulated for proper tissue regeneration throughout an organisms’ lifetime. Epigenetic regulators, such as the polycomb group (PcG) of proteins have been implicated in modulating gene expression in adult SCs to maintain homeostatic and regenerative balances in adult tissues. In this review, we summarize the recent findings that elucidate the composition and function of the polycomb repressive complex machinery and highlight their role in diverse adult stem cell compartments.

ABSTRACT The proteins of the Polycomb group (PcG) are required for maintaining regulator genes, such as the homeotic selectors, stably and heritably repressed in appropriate developmental domains. It has been suggested that PcG proteins silence genes by creating higher-order chromatin structures at their chromosomal targets, thus preventing the interaction of components of the transcriptional machinery with theircis-regulatory elements. An unresolved issue is how higher order-structures are anchored at the chromatin base, the nucleosomal fiber. Here we show a direct biochemical interaction of a PcG protein—the Polycomb (PC) protein—with nucleosomal core particles in vitro. The main nucleosome-binding domain coincides with a region in the C-terminal part of PC previously identified as the repression domain. Our results suggest that PC, by binding to the core particle, recruits other PcG proteins to chromatin. This interaction could provide a key step in the establishment or regulation of higher-order chromatin structures.

Abstract The Polycomb group (PcG) gene RNF2 (RING2) encodes a catalytic subunit of the Polycomb repressive complex 1 (PRC1), an evolutionarily conserved machinery that post-translationally modifies chromatin to maintain epigenetic transcriptional repressive states of target genes including Hox genes. Here, we describe two individuals, each with rare de novo missense variants in RNF2. Their phenotypes include intrauterine growth retardation, severe intellectual disabilities, behavioral problems, seizures, feeding difficulties and dysmorphic features. Population genomics data suggest that RNF2 is highly constrained for loss-of-function (LoF) and missense variants, and both p.R70H and p.S82R variants have not been reported to date. Structural analyses of the two alleles indicate that these changes likely impact the interaction between RNF2 and BMI1, another PRC1 subunit or its substrate Histone H2A, respectively. Finally, we provide functional data in Drosophila that these two missense variants behave as LoF alleles in vivo. The evidence provide support for deleterious alleles in RNF2 being associated with a new and recognizable genetic disorder. This tentative gene-disease association in addition to the 12 previously identified disorders caused by PcG genes attests to the importance of these chromatin regulators in Mendelian disorders.

Polycomb repressive complex 2 (PRC2) installs and spreads repressive histone methylation marks on eukaryotic chromosomes. Because of the key roles that PRC2 plays in development and disease, how this epigenetic machinery interacts with DNA and nucleosomes is of major interest. Nonetheless, the mechanism by which PRC2 engages with native-like chromatin remains incompletely understood. In this work, we employ single-molecule force spectroscopy and molecular dynamics simulations to dissect the behavior of PRC2 on polynucleosome arrays. Our results reveal an unexpectedly diverse repertoire of PRC2 binding configurations on chromatin. Besides reproducing known binding modes in which PRC2 interacts with bare DNA, mononucleosomes, and adjacent nucleosome pairs, our data also provide direct evidence that PRC2 can bridge pairs of distal nucleosomes. In particular, the “1–3” bridging mode, in which PRC2 engages two nucleosomes separated by one spacer nucleosome, is a preferred low-energy configuration. Moreover, we show that the distribution and stability of different PRC2–chromatin interaction modes are modulated by accessory subunits, oncogenic histone mutations, and the methylation state of chromatin. Overall, these findings have implications for the mechanism by which PRC2 spreads histone modifications and compacts chromatin. The experimental and computational platforms developed here provide a framework for understanding the molecular basis of epigenetic maintenance mediated by Polycomb-group proteins.

Abstract Loss of surface MHC I is an important mechanism by which cancer cells evade immune surveillance, and it can correlate with worse prognosis and resistance to immunotherapy. MHC I surface expression can be affected by genomic or transcriptional alterations to HLA-A/B/C, β2M, or the class I antigen processing machinery. In the case of transcriptional loss, we hypothesized that certain cancers might employ specific epigenetic programs to enforce downregulation of the MHC I pathway. Thus, we sought to identify novel MHC I regulators in MHC I-low cancers. In particular, we focused on Merkel cell carcinoma (MCC), a rare but aggressive neuroendocrine skin cancer that is caused by the Merkel cell polyomavirus in 80% of cases. Notably, MHC I downregulation is prevalent in MCC. We first generated and characterized a series of patient-derived MCC cell lines, in which MHC I surface expression was low but inducible with IFN-γ. We then conducted genome-scale CRISPR-KO and open reading frame (ORF) gain-of-function screens in one of our virus-positive MCC lines, using FACS to select for perturbations that upregulated surface MHC I. One of the top hits from these screens was PRC1.1, a non-canonical Polycomb repressive complex. Polycomb complexes are known to mediate gene silencing through chromatin modification and play an important role in development and cancer. These studies suggest a possible role for PRC1.1 in suppressing MHC I in MCC.

Chez les eucaryotes, la maintenance de l’identité cellulaire implique le contrôle précis de l’expression des gènes. Ceci résulte de l’action concertée des facteurs de transcription et des facteurs contrôlant la structure de la chromatine. Les complexes répresseurs Polycomb (PRC1 et PRC2) sont des modificateurs de la chromatine qui orchestrent la répression transcriptionnelle en catalysant respectivement l’ubiquitinylation de H2A (H2Aub) et la méthylation de H3K27. A l’inverse, BAP1 (BRCA1-Associated Protein 1) favorise la transcription en retirant H2Aub, agissant donc comme un antagoniste de PRC1. Toutefois, les détails du mécanisme par lequel BAP1 régule la transcription reste mal compris. L’interaction entre PRC1 et PRC2 est également un sujet encore débattu. Mon projet de thèse visait à étudier ces deux importantes questions.(1) La protéine BAP1 est localisée à une fraction des enhancers où elle stabilise le recrutement de BRD4.Dans ces études, nous avons montré que BAP1 favorise la transcription en s’opposant au complexe PRC1 et que BAP1 est inerte en son absence. Des analyses à l’échelle du génome entier ont révélé que la protéine BAP1 est recrutée à une fraction des enhancers. Par ailleurs, l’inactivation de BAP1 amène à l’accumulation de H2Aub et à l’altération du recrutement de BRD4. En accord avec ces résultats, des expériences de microscopie à super résolution indiquent une réduction des condensées de BRD4 et de MED1 dans les cellules knockout pour BAP1. Cela suggère que BAP1 a un rôle crucial pour l’intégrité de certains enhancers. De façon importante, en traitant des cellules isogèniques avec des inhibiteurs de BET, nous avons montré que les cellules mutantes pour BAP1 montrent une sensibilité particulière à l’inhibition de la prolifération. Ce résultat suggère que promouvoir les perturbations des enhancers pourrait constituer une stratégie thérapeutique dans les pathologies où le gène BAP1 est muté.(2) PRC2 réprime la transcription indépendamment de PRC1PRC1 et PRC2 ont été considérés depuis longtemps comme agissant de concert pour maintenir la répression. Toutefois, en analysant les profiles transcriptomiques de cellules où soit PRC1, soit PRC2, soit les deux sont inactivés, nous avons démontré que PRC1 et PRC2 peuvent agir de façon autonome pour réprimer la transcription. Au travers d’approches non-biaisées et d’approches basées sur une sélection de gènes candidats, nous essayons d’identifier les effecteurs de cette répression dépendant exclusivement de PRC2. Cela implique l’étude de protéines préalablement proposées comme interagissant avec H3K27me3. Cette étude est en cours mais il est probable qu’elle va révéler de nouveaux acteurs de la répression dépendant de PRC2
In eukaryotes, the maintenance of cell identity entails the precise control of gene expression, which results from the concerted actions of transcription factors and factors controlling chromatin structure. Polycomb repressive complex 1 and 2 (PRC1 and PRC2) are chromatin modifiers that orchestrate transcriptional repression by catalyzing H2Aub and H3K27me3, respectively. By contrast, BRCA1-associated protein 1 (BAP1) promotes transcription by removing H2Aub, acting as an antagonist of PRC1. However, the detailed mechanism of how BAP1 regulates transcription remains largely elusive. The interplay between PRC1 and PRC2 is also far from being fully understood. My PhD study aimed at investigating the underlying mechanisms for these two important questions.(1) BAP1 is recruited to a subset of active enhancers where it stabilizes BRD4 occupancy.In these studies, we showed that BAP1 promotes transcription by opposing PRC1 activity, and that BAP1 is mostly inert in its absence. Genome-wide analysis revealed that BAP1 is recruited to a subset of active enhancers. Besides, inactivation of BAP1 led to accumulation of H2Aub and impaired BRD4 recruitment. Consistently, super-resolution microscopy demonstrated reduced condensates of BRD4 and MED1 in BAP1-KO cells. This suggests that BAP1 has a crucial function for the integrity of a subset of enhancers. Importantly, by treating isogenic cells with BET inhibitors, we showed that cells mutant for BAP1 display a more pronounced proliferative response. This result suggests that further perturbation of enhancers function could be a therapeutic strategy for BAP1-null malignancies.(2) PRC2 represses transcription independently of PRC1PRC1 and PRC2 are long considered cooperating to maintain gene repression. However, analyzing transcriptomic profiles of PRC1-null, PRC2-null and PRC1/2-null cells, we demonstrated that both PRC1 and PRC2 can autonomously repress transcription. Through both unbiased and candidate-based approaches, we focus on identifying downstream effectors of PRC2-mediated silencing in the absence of PRC1. This includes investigating the roles of previously proposed H3K27me3 readers. While this study is still ongoing, it is likely that it will reveal new actor for PRC2-mediated repression