TIP48/Reptin and H2A.Z Requirement for Initiating Chromatin Remodeling in Estrogen-Activated Transcription

Abstract

Histone variants, including histone H2A.Z, are incorporated into specific genomic sites and participate in transcription regulation. The role of H2A.Z at these sites remains poorly characterized. Our study investigates changes in the chromatin environment at the Cyclin D1 gene (CCND1) during transcriptional initiation in response to estradiol in estrogen receptor positive mammary tumour cells. We show that H2A.Z is present at the transcription start-site and downstream enhancer sequences of CCND1 when the gene is poorly transcribed. Stimulation of CCND1 expression required release of H2A.Z concomitantly from both these DNA elements. The AAA+ family members TIP48/reptin and the histone variant H2A.Z are required to remodel the chromatin environment at CCND1 as a prerequisite for binding of the estrogen receptor (ERα) in the presence of hormone. TIP48 promotes acetylation and exchange of H2A.Z, which triggers a dissociation of the CCND1 3′ enhancer from the promoter, thereby releasing a repressive intragenic loop. This release then enables the estrogen receptor to bind to the CCND1 promoter. Our findings provide new insight into the priming of chromatin required for transcription factor access to their target sequence. Dynamic release of gene loops could be a rapid means to remodel chromatin and to stimulate transcription in response to hormones.

Monoubiquitination of Histone H2B Blocks Eviction of Histone Variant H2A.Z from Inducible Enhancers

“…In the nucleosome, H2B forms dimers with H2A or H2A variants, of which H2A.Z is preferentially found at TSS and enhancers (Jin et al, 2009;Dalvai et al, 2013;Brunelle et al, 2015). In agreement with this, we found that H2A.Z levels increase with the strength of the ERBS, independently of any induction ( Figures 4A and S4A).…”

Reptin and Pontin Oligomerization and Activity Are Modulated through Histone H3 N-terminal Tail Interaction

“…Relocation of Reptin has been observed on inducible promoters (this study and Ref. 30). 5 In the repressed state, Reptin is associated with promoter sequences.…”

“…Chromatin fragments were immunoprecipitated using antibodies against H3k9-me2 (ab1220, Abcam), pan-acetyl H3 (ab47915, Abcam), H3 (ab1791, ABCAM), or an irrelevant HA antibody (H6908, Sigma). Anti-Reptin was synthesized by the laboratory from the purified protein (30). The precipitated DNA was amplified by real time PCR, with primer sets designed to amplify the promoter (transcription start site) of the PGR gene (Fig.…”

“…We chose a promoter whose activity depends on estrogen receptor (ER)-␣ and the progesterone receptor promoter (PgR), in ER␣-positive human breast cancer cells (MCF-7). It was recently established that Reptin is required for initiating chromatin reorganization during 17␤-estradiol (E 2 ) induction of the ER␣ target oncogene, cyclin D1 (CCND1) (30). Pontin itself was not included in this study; nonetheless, it usually belongs to Reptin complexes, and in particular, silencing one protein led to depletion of both in MCF7 cells and other tumor cell lines (37).…”

H2B monoubiquitination: t'ub or not t'ub for inducible enhancers

“…12 Interestingly, a similar apparent contradiction between activation and repressive roles also exists for H2A.Z. 25,26 We demonstrated that H2Bub1 levels correlate with H2A.Z levels, 12 seemingly reinforcing the confusion. Our ChIP-seq data re-analysis highlighted the fact that the stronger an enhancer is, the higher the levels of H2Bub1 are across this enhancer, independently of whether it has already been induced or not.…”