The histone variant mH2A1.1 interferes with transcription by down-regulating PARP-1 enzymatic activity

Abstract

The histone variant mH2A is believed to be involved in transcriptional repression, but how it exerts its function remains elusive. By using chromatin immunoprecipitation and tandem affinity immunopurification of the mH2A1.1 nucleosome complex, we identified numerous genes with promoters containing mH2A1.1 nucleosomes. In particular, the promoters of the inducible Hsp70.1 and Hsp70.2 genes, but not that of the constitutively expressed Hsp70.8, were highly enriched in mH2A1.1. PARP-1 was identified as a part of the mH2A1.1 nucleosome complex and was found to be associated with the Hsp70.1 promoter. A specific interaction between mH2A1.1 and PARP-1 was demonstrated and found to be associated with inactivation of PARP-1 enzymatic activity. Heat shock released both mH2A1.1 and PARP-1 from the Hsp70.1 promoter and activated PARP-1 automodification activity. The data we present point to a novel mechanism for control of Hsp70.1 gene transcription. mH2A1.1 recruits PARP-1 to the promoter, thereby inactivating it. Upon heat shock, the Hsp70.1 promoter-bound PARP-1 is released to activate transcription through ADP-ribosylation of other Hsp70.1 promoter-bound proteins.[Keywords: Histone variant; mH2A; PARP-1; ADP-ribosylation; transcription] Supplemental material is available at http://www.genesdev.org.

Poly(ADP-ribose) Polymerase 1 Is Inhibited by a Histone H2A Variant, MacroH2A, and Contributes to Silencing of the Inactive X Chromosome

“…In this model, PARP-1 might function to compact chromatin in regions of the genome packaged with macroH2A-containing nucleosomes, contributing to heterochromatin formation and transcriptional repression. While this work was under review, Ouararhni et al (32) reported that macroH2A1.1 associates with and inhibits PARP-1 and that transcriptional repression of a heat shock gene is mediated by macroH2A1.1 and PARP-1, consistent with a role for the macroH2A-PARP-1 interaction in transcriptional silencing. (Fig.…”

HJURP binds CENP-A via a highly conserved N-terminal domain and mediates its deposition at centromeres

“…Cell extracts from the tagged cells were prepared and the e-H3.1 and e-CENP-A prenucleosomal complexes were purified by sequential immunoprecipitations with antiFLAG antibody followed by antiHA antibody (29). The proteins associated with e-CENP-A and e-H3.1 were separated in 4-12% gradient PAGE containing SDS and silver stained (Fig.…”

“…Prenucleosomal CENP-A and H3.1 complexes were purified from soluble nuclear extracts prepared from stable HeLa cell lines expressing either CENP-A or H3.1 proteins fused to C-terminal FLAG and HA epitope tags (e-CENP-A/e-H3.1). A tandem affinity purification protocol on antiFlag antibody-conjugated agarose followed by antiHA purification and peptide elution was used (29).…”

Transcription cofactors TRIM24, TRIM28, and TRIM33 associate to form regulatory complexes that suppress murine hepatocellular carcinoma

“…Generation of HeLa cell lines expressing the N-terminal FLAG-HA epitope-tagged (E)-TRIM24 protein was performed by retroviral infection, extract preparation and tandem affinity purification using the FLAG and HA tags were all performed essentially as previously described (34). Purified complexes were analyzed on 4-12% Tris-Bis NuPage gels (Invitrogen) and stained with Coomassie blue.…”