Poly(ADP-Ribosyl) Glycohydrolase Prevents the Accumulation of Unusual Replication Structures during Unperturbed S Phase

Poly(ADP-ribosyl)ation (PAR) has been implicated in various aspects of the cellular response to DNA damage and genome stability. Although 17 human poly(ADP-ribose) polymerase (PARP) genes have been identified, a single poly(ADP-ribosyl) glycohydrolase (PARG) mediates PAR degradation. Here we investi...

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Veröffentlicht in:Molecular and cellular biology Jg. 35; H. 5; S. 856 - 865
Hauptverfasser: Ray Chaudhuri, Arnab, Ahuja, Akshay Kumar, Herrador, Raquel, Lopes, Massimo
Format: Journal Article
Sprache:Englisch
Veröffentlicht: United States Taylor & Francis 01.03.2015
American Society for Microbiology
Schlagworte:
Ataxia Telangiectasia Mutated Proteins - metabolism
Cell Proliferation
chromatin
Chromatin - metabolism
DNA Damage
DNA Repair
DNA replication
DNA, Single-Stranded
drug therapy
electron microscopy
genome
Glycoside Hydrolases - metabolism
GTPase-Activating Proteins - metabolism
HeLa Cells
Humans
interphase
Intracellular Signaling Peptides and Proteins - metabolism
Microscopy, Confocal
Microscopy, Electron
Phenotype
phosphorylation
Rad51 Recombinase - metabolism
S Phase
single-stranded DNA
Tumor Suppressor p53-Binding Protein 1
ISSN:1098-5549, 0270-7306, 1098-5549
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Zusammenfassung:Poly(ADP-ribosyl)ation (PAR) has been implicated in various aspects of the cellular response to DNA damage and genome stability. Although 17 human poly(ADP-ribose) polymerase (PARP) genes have been identified, a single poly(ADP-ribosyl) glycohydrolase (PARG) mediates PAR degradation. Here we investigated the role of PARG in the replication of human chromosomes. We show that PARG depletion affects cell proliferation and DNA synthesis, leading to replication-coupled H2AX phosphorylation. Furthermore, PARG depletion or inhibition per se slows down individual replication forks similarly to mild chemotherapeutic treatment. Electron microscopic analysis of replication intermediates reveals marked accumulation of reversed forks and single-stranded DNA (ssDNA) gaps in unperturbed PARG-defective cells. Intriguingly, while we found no physical evidence for chromosomal breakage, PARG-defective cells displayed both ataxia-telangiectasia-mutated (ATM) and ataxia-Rad3-related (ATR) activation, as well as chromatin recruitment of standard double-strand-break-repair factors, such as 53BP1 and RAD51. Overall, these data prove PAR degradation to be essential to promote resumption of replication at endogenous and exogenous lesions, preventing idle recruitment of repair factors to remodeled replication forks. Furthermore, they suggest that fork remodeling and restarting are surprisingly frequent in unperturbed cells and provide a molecular rationale to explore PARG inhibition in cancer chemotherapy.
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Citation Ray Chaudhuri A, Ahuja AK, Herrador R, Lopes M. 2015. Poly(ADP-ribosyl) glycohydrolase prevents the accumulation of unusual replication structures during unperturbed S phase. Mol Cell Biol 35:856–865. doi:10.1128/MCB.01077-14.
Present address: Arnab Ray Chaudhuri, Laboratory of Genome Integrity, Center for Cancer Research, National Cancer Institute, Bethesda, Maryland, USA.
A.R.C. and A.K.A. contributed equally to this article.
ISSN:1098-5549
0270-7306
1098-5549
DOI:10.1128/MCB.01077-14