Uncoupling of the transactivation and transrepression functions of PARP1 protein

Abstract

Poly(ADP ribose) polymerase 1 (PARP1) is a nuclear protein that regulates chromatin remodeling and transcription as well as DNA repair and genome stability pathways. Recent studies have revealed a paradoxical dual role of PARP1 protein in transcription. Specifically, although PARP1 controls transcriptional activation of a subset of genes that are heat shock-or hormone-dependent, it also directly inactivates transcription, establishes heterochromatin domains, and silences retrotransposable elements. However, the domains required for these disparate functions are currently unknown. In this paper, we report the discovery of a previously undescribed mutation in the Drosophila Parp locus. We show that the mutants express a deletion mutant of PARP1 protein with an altered DNA binding domain that carries only the second Znfinger. We demonstrate that this alteration specifically excludes PARP1 protein from heterochromatin and makes PARP1 unable to maintain repression of retrotransposable elements. By characterizing the biological activity of this unique PARP1 mutant protein isoform, we have uncoupled the transactivation and transrepression functions of this protein.chromatin | poly(ADP ribose) polymerase | transcription P oly(ADP ribose) polymerase 1 (PARP1) protein has been known for decades as a nuclear protein that recognizes and binds nicks and ends of DNA and catalyses poly(ADP ribose) (pADPr) synthesis (1). The basic enzymatic reactions catalyzed by PARP1 involve transferring ADPr from nicotinamide-adenine dinucleotide to either a protein acceptor or an existing pADPr chain, the average length of which is 80 or more residues (2). PARP1 protein can modify numerous chromatin proteins in vivo and in vitro (3). A key role of PARP1 was shown in DNA repair and apoptosis (3), where PARP works as a trigger between the DNA repair (4) and apoptotic pathways (5). PARP1 enzymatic activity has also been shown to be required for normal assembly of higher order chromatin structures and for transcriptional activation (6). Moreover, it has been shown that PARP1 regulates the transcription of these genes by inducing chromatin loosening at targeted genetic loci (6, 7). Finally, PARP1 establishes silent chromatin domains and represses retrotransposable elements (8).The characterization of deletion mutants of PARP that distinguish among the varied functions of this protein is essential to establish a more complete understanding of PARP1 protein biology. At present, however, we have identified neither the mechanism of PARP protein targeting to specific chromatin domains nor the mechanism of local PARP activation. Closing these gaps in our current knowledge is complicated because the presence of 18 paralogous PARP proteins (9) in mammals most likely results in corresponding functional redundancies. The Drosophila genome (8, 10, 11) encodes only a single nuclear PARP (PARP1), making this animal an invaluable model system for the study of PARP functions.The PARP1 protein has three functionally defined domains conserved from human to Droso...

The zinc-finger domains of PARP1 cooperate to recognize DNA strand breaks

“…Activation must therefore be tightly regulated and restricted to sites of DNA damage, with minimal activation by its association with undamaged DNA, which nonetheless ensures a high concentration of PARP1 on chromatin and thereby enables its very rapid recruitment to sites of damage as a primary sensor. Data from Drosophila 40 suggest that ZnF1 and ZnF2 have differential roles in mediating chromatin association, but our data show that both zinc-finger domains are required for human PARP1 to be localized to and retained at sites of DNA damage in human cells.…”

Poly(ADP-Ribose) Polymerase 1 Promotes Transcriptional Repression of Integrated Retroviruses

“…These contradictory results may not be surprising if we take into consideration the PARP functional redundancy found in mammalian cells (17,18). Therefore, the study of PARP-1 functions in simpler organisms that lack this functional redundancy is advantageous, and this strategy has significantly contributed to the understanding of the cellular functions of the enzyme (21,22,25). In addition, PARP-1 functions identified in simpler organisms are relevant to more complex organisms due to the high evolutionary conservation of PARP-1 from Drosophila to humans.…”

“…Incorporation of PARP-1 into the chromatin causes chromatin compaction and transcriptional repression (20,21,(23)(24)(25). This repressive activity of PARP-1 on transcription mediates silencing of retrotransposable elements and the formation of heterochromatin in Drosophila (21,25). Activation of the enzymatic activity of nucleosome-incorporated PARP-1 leads to auto-PARylation and dissociation of PARP-1 from chromatin, causing chromatin decondensation and activation of transcription (20,22,24).…”

“…These structural domains interact dynamically and coordinate different catalytic-independent and -dependent PARP-1 functions. PARP-1 is incorporated into nucleosomes in a catalytic-independent manner (20)(21)(22) and requires the DNA binding domain (20,21,23,24) and the interaction of the C-terminal region (amino acids 214 to 1014) with the nucleosome core histone proteins (22). Incorporation of PARP-1 into the chromatin causes chromatin compaction and transcriptional repression (20,21,(23)(24)(25).…”

The expanding role ofPARPs in the establishment and maintenance of heterochromatin

“…Moreover, PARP1 binds to and regulates repressive heterochromatic structures such as telomeres [8,9], pericentromeric regions [10,11] and heterochromatic rRNA genes [12]. In Drosophila, genetic studies have implicated PARP1 in organizing the chromatin structure of nucleoli and heterochromatin domains and silencing retrotransposable elements [13,14]. The enzymatic activity of PARP1 has been proposed as the switch event that may distinguish between the co-repressor and co-activator functions of PARP1 [15].…”