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

Abstract

Poly(ADP-ribose) polymerase 1 (PARP-1) is a nuclear enzyme that is involved in modulating chromatin structure, regulation of gene expression, and sensing DNA damage. Here, we report that PARP-1 enzymatic activity is inhibited by macroH2A, a vertebrate histone H2A variant that is enriched on facultative heterochromatin. MacroH2A family members have a large C-terminal non-histone domain (NHD) and H2A-like histone domain. MacroH2A1.2 and PARP-1 interact in vivo and in vitro via the NHD. The NHD of each macroH2A family member was sufficient to inhibit PARP-1 enzymatic activity in vitro. The NHD of macroH2A1.2 was a mixed inhibitor of PARP-1 catalytic activity, with affects on both catalytic activity and the substrate binding affinity of PARP-1. Depletion of PARP-1 by RNA interference caused reactivation of a reporter gene on the inactive X chromosome, demonstrating that PARP-1 participates in the maintenance of silencing. These results suggest that one function of macroH2A in gene silencing is to inhibit PARP-1 enzymatic activity, and this may affect PARP-1 association with chromatin.In eukaryotic cells DNA is packaged into chromatin, and this packaging impacts all DNA-templated processes, including transcription. Regulated changes in chromatin structure are crucial to establish and maintain the diverse expression profiles that characterize the hundreds of cell types in multicellular organisms (1). The nucleosome is the structural unit of chromatin and comprises 147 bp of DNA wrapped around a histone octamer, which is composed of two copies each of the four core histones H2A, H2B, H3, and H4.Chromatin structure can be affected through the action of ATP-dependent chromatin remodeling enzymes or by the covalent modification of histone proteins, creating binding sites for additional regulatory proteins (1, 2). In addition, chromatin structure can be modulated through the binding of effector proteins to nucleosomes. Poly(ADP-ribose) polymerase 1 (PARP-1) 5 is an example of a nucleosome binding protein that can affect chromatin structure (3, 4). PARP-1 is the prototypical member of a family of PARP proteins, which catalyze the transfer of ADP-ribose units from donor nicotinamide adenine dinucleotide (NAD ϩ ) molecules to target proteins (5). PARP-1 functions as a structural component of chromatin, modulator of chromatin structure, and a sensor of DNA damage through its intrinsic enzymatic activity (4, 6, 7). In the absence of NAD ϩ , PARP-1 binds to nucleosomes, compacts chromatin, and inhibits transcription in vitro (3). Furthermore, catalytically inactive PARP-1 maintains silencing of heterochromatic retrotransposons in Drosophila (8). However, at physiological concentrations of NAD ϩ , PARP-1 is enzymatically active and does not bind nucleosomes (3). Despite this, PARP-1 binds chromatin in vivo and is implicated in transcriptional silencing, suggesting that modulation of PARP-1 activity in vivo may be a mechanism that is employed to direct changes in chromatin structure.Chromatin structure can also be regulated by rep...

Weak but Uniform Enrichment of the Histone Variant macroH2A1 along the Inactive X Chromosome

“…A potential candidate is PARP-1. Indeed, PARP-1 was found to be associated with mH2A nucleosomes in vivo (24,25), and it was shown that the association of PARP-1 promotes its condensation in vitro (36).…”

Formation of nuclear heterochromatin

“…PARP1 localizes at telomeres 27 and centromeres, 28,29 is present within the nucleolus, 30,31 binds to mouse pericentric repeats and human α satellites 32 and it was implicated in the formation of facultative heterochromatin of the inactive X chromosome. 33 However, until recently, whether and how PARP1 functions in the formation of heterochromatin was not yet clear. Recent results identified PARP1 as an interaction partner of TIP5.…”

The macroPARP genes parp‐9 and parp‐14 are developmentally and differentially regulated in mouse tissues

“…A macro domain is present within the histone variant macroH2A that is associated to transcriptional repression and X chromosome inactivation (Angelov et al, 2003). Interestingly, this domain is able to negatively regulate PARP-1 activity on particular promoters that must remain inactive in the absence of appropriated stimulation or on the inactive X chromosome (Ouararhni et al, 2006;Nusinow et al, 2007). Macro domains were recently shown to bind ADP-ribose and its derivatives (Karras et al, 2005).…”