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Drouineau, E.

Publications and source records attributed to Drouineau, E..

3 recordsLinked to original sources

How cBAF and PBAF regulate the nucleosomal and subnucleosomal landscape of promoters, enhancers and REST repressor binding sites

BAF (SWI/SNF) chromatin remodeling complexes belong to three subfamilies, cBAF, PBAF, and ncBAF, that regulate the nucleosomal organization and accessibility of transcription factors (TFs) DNA binding sites at cis-regulatory elements (CREs). However, the functional specificities of these complexes at promoters, enhancers, and other categories of CREs are poorly understood. Here, we examined how the BAF complexes regulate the nucleosomal and subnucleosomal landscape in which transcription initiation occurs in vivo. We first described how these complexes regulate the earliest step of transcription preinitiation complex (PIC) assembly, which is defined by the binding of the general transcription factors TBP and TFIIA. We show that in the presence of a TATA box, PIC formation occurs onto a core promoter or enhancer region still wrapped into a fragile nucleosome or a subnucleosomal particle. Conversely, without a TATA box, PIC assembles onto genomic templates mostly stripped of histones. We establish that cBAF, but not PBAF or ncBAF, regulates this chromatin organization and the first step of PIC formation at promoters and enhancers. We observed that individual subunits of cBAF have specific functions at promoters: the BRG1 ATPase and SMARCB1 regulate the production of fragile nucleosomes and subnucleosomal particles, while ARID1A stimulates the binding of TBP/TFIIA independently of the ATPase subunit. In contrast, all three subunits are systematically required for both chromatin organization and TBP/TFIIA binding at enhancers. Finally, we revealed that a subset of PBAF subunits controls the nucleosomal and subnucleosomal landscape at REST repressor binding sites. Our integrative study identified specific functions of cBAF and PBAF subunits in controlling the binding of TFs and the nucleosomal and subnucleosomal organization of CREs.

genomics↗

Oct4 interacts with subnucleosomal particles generated by Brg1 at enhancers

BRG1, the catalytic subunit of the mammalian SWI/SNF complexes, is essential for chromatin opening at enhancers. However, the nature of the open chromatin remains unclear. Here we show that in addition to producing histone-free DNA, BRG1 generates hemisome-like subnucleosomal particles containing the four core histones associated with 50-80 base pairs of DNA. Our genome-wide analysis indicates that BRG1 makes these particles by targeting and splitting fragile nucleosomes. In mouse embryonic stem cells, these subnucleosomes become an in vivo binding substrate for the master transcription factor OCT4 independently of the presence of OCT4 DNA motifs. At enhancers, the OCT4-subnucleosome interaction increases OCT4 occupancy and amplifies the genomic interval bound by OCT4 by up to one order of magnitude, compared to the region occupied on histone-free DNA. We suggest that BRG1-dependent subnucleosomes orchestrate an epigenetic mechanism that projects OCT4 function in chromatin opening beyond its DNA motifs.

genomics↗

Identification of RNAs bound by Hfq reveals widespread RNA partners and a sporulation regulator in the human pathogen Clostridioides difficile

Noncoding RNAs (ncRNA) have emerged as important components of regulatory networks governing bacterial physiology and virulence. Previous deep-sequencing analysis identified a large diversity of ncRNAs in the human enteropathogen Clostridioides (Clostridium) difficile. Some of them are trans-encoded RNAs that could require the RNA chaperone protein Hfq for their action. Recent analysis suggested a pleiotropic role of Hfq in C. difficile with the most pronounced effect on sporulation, a key process during the infectious cycle of this pathogen. However, a global view of RNAs interacting with C. difficile Hfq is missing. In the present study, we performed RNA immunoprecipitation high-throughput sequencing (RIP-Seq) to identify Hfq-associated RNAs in C. difficile. Our work revealed a large set of Hfq-interacting mRNAs and ncRNAs, including mRNA leaders and coding regions, known and potential new ncRNAs. In addition to trans-encoded RNAs, new categories of Hfq ligands were found including cis-antisense RNAs, riboswitches and CRISPR RNAs. ncRNA-mRNA and ncRNA-ncRNA pairings were postulated through computational predictions. Investigation of one of the Hfq-associated ncRNAs, RCd1, suggests that this RNA contributes to the control of late stages of sporulation in C. difficile. Altogether, these data provide essential molecular basis for further studies of post-transcriptional regulatory network in this enteropathogen.

microbiology↗