A Variant of the Breast Cancer Type 2 Susceptibility Protein (BRC) Repeat Is Essential for the RECQL5 Helicase to Interact with RAD51 Recombinase for Genome Stabilization

Islam, M. Nurul; Paquet, Nicolas; Fox, David; Dray, Eloïse; Zheng, X.F. Steven; Klein, Hannah L.; Sung, Patrick; Wang, Weidong

doi:10.1074/jbc.m112.375014

Cited by 45 publications

(71 citation statements)

References 40 publications

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“…DSBs are critical DNA lesions that require an immediate cellular response because failure to do so can result in chromosomal rearrangements, genomic instability, and even cell death. It has been shown that RECQL5 interacts with RAD51 to inhibit RAD51-induced D-loop formation and sister chromatid exchange (29). Consistent with that finding, RECQL5 prevents RAD51 single-stranded DNA (ssDNA) filament formation by promoting synthesis-dependent strand annealing to mediate DSB repair (30).…”

supporting

confidence: 64%

Differential and Concordant Roles for Poly(ADP-Ribose) Polymerase 1 and Poly(ADP-Ribose) in Regulating WRN and RECQL5 Activities

Khadka

Hsu

Veith

et al. 2015

Molecular and Cellular Biology

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b Poly(ADP-ribose) (PAR) polymerase 1 (PARP1) catalyzes the poly(ADP-ribosyl)ation (PARylation) of proteins, a posttranslational modification which forms the nucleic acid-like polymer PAR. PARP1 and PAR are integral players in the early DNA damage response, since PARylation orchestrates the recruitment of repair proteins to sites of damage. Human RecQ helicases are DNA unwinding proteins that are critical responders to DNA damage, but how their recruitment and activities are regulated by PARPs and PAR is poorly understood. Here we report that all human RecQ helicases interact with PAR noncovalently. Furthermore, we define the effects that PARP1, PARylated PARP1, and PAR have on RECQL5 and WRN, using both in vitro and in vivo assays. We show that PARylation is involved in the recruitment of RECQL5 and WRN to laser-induced DNA damage and that RECQL5 and WRN have differential responses to PARylated PARP1 and PAR. Furthermore, we show that the loss of RECQL5 or WRN resulted in increased sensitivity to PARP inhibition. In conclusion, our results demonstrate that PARP1 and PAR actively, and in some instances differentially, regulate the activities and cellular localization of RECQL5 and WRN, suggesting that PARylation acts as a fine-tuning mechanism to coordinate their functions in time and space during the genotoxic stress response. Mammalian cells are constantly exposed to various environmental genotoxic stresses that can hamper genomic stability. To maintain genomic integrity, cells have developed various complex DNA repair machineries that effectively identify DNA lesions and activate DNA damage responses (DDRs) and DNA repair (1, 2). One of the early DDR mechanisms is through the activation of poly(ADP-ribose) (PAR) polymerases (PARPs). PARP1 is the most ubiquitously expressed PARP family member and constitutes the majority of PAR synthesis in human cells (3,4). In response to DNA damage, such as DNA single-strand breaks or double-strand breaks (DSBs), PARP1 is activated and promotes PAR synthesis. It utilizes NAD (NAD ϩ ) to form PAR at the DNA lesions or breaks, thereby posttranslationally modifying itself and other proteins of interest. Importantly, PAR formation is highly dynamic, because shortly after being synthesized, it is rapidly hydrolyzed by PARP's catabolic counterparts, PAR glycohydrolase (PARG) and ADP-ribosylhydrolase 3 (ARH3) (5-8). Neither enzyme is able to remove the last ADP-ribose moiety from acceptor proteins; macrodomain-containing proteins, such as MacroD1 and MacroD2, fulfill this task, leaving behind an unmodified amino acid that is readily available for the next round of poly-(ADP-ribosyl)ation (PARylation) (9, 10). The site of formation of PAR can act as a docking site to promote protein recruitment and protein-protein interactions. This docking site serves to integrate diverse cellular signaling pathways, including DNA damage detection and repair, transcription, chromatin remodeling, and cell death (11). PAR formation plays a critical role in the rapid recruitment of DNA repair protein...

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supporting

confidence: 64%

Differential and Concordant Roles for Poly(ADP-Ribose) Polymerase 1 and Poly(ADP-Ribose) in Regulating WRN and RECQL5 Activities

Khadka

Hsu

Veith

et al. 2015

Molecular and Cellular Biology

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“…Srs2‐BRCv structurally resembles the BRC repeat of the human tumour suppressor BRCA2 (Islam et al , 2012) which mediates BRCA2–RAD51 interaction, thereby promoting recruitment of RAD51 to DSBs and regulation of RAD51 recombinase activity (Jensen et al , 2010; Liu et al , 2010; Thorslund et al , 2010). Srs2‐BRCv also promotes Srs2–Rad51 interaction in vitro (Islam et al , 2012). Notably, in our genetic assays, srs2 mutants lacking BRCv, Srs2(1–836), had an intermediate phenotype: they resembled wild type in SSA and BIR but were partially deficient in de novo telomere addition (Fig 8B–D).…”

Section: Discussionmentioning

confidence: 99%

Unloading of homologous recombination factors is required for restoring double‐stranded DNA at damage repair loci

et al. 2017

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Cells use homology‐dependent DNA repair to mend chromosome breaks and restore broken replication forks, thereby ensuring genome stability and cell survival. DNA break repair via homology‐based mechanisms involves nuclease‐dependent DNA end resection, which generates long tracts of single‐stranded DNA required for checkpoint activation and loading of homologous recombination proteins Rad52/51/55/57. While recruitment of the homologous recombination machinery is well characterized, it is not known how its presence at repair loci is coordinated with downstream re‐synthesis of resected DNA. We show that Rad51 inhibits recruitment of proliferating cell nuclear antigen (PCNA), the platform for assembly of the DNA replication machinery, and that unloading of Rad51 by Srs2 helicase is required for efficient PCNA loading and restoration of resected DNA. As a result, srs2Δ mutants are deficient in DNA repair correlating with extensive DNA processing, but this defect in srs2Δ mutants can be suppressed by inactivation of the resection nuclease Exo1. We propose a model in which during re‐synthesis of resected DNA, the replication machinery must catch up with the preceding processing nucleases, in order to close the single‐stranded gap and terminate further resection.

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“…Recently, a BRC repeat variant, named BRCv, was identified in HsRecQ5, and was shown to be essential for RecQ5 to interact with Rad51. 38) BRC repeats interact with Rad51 and promote homologous recombination by regulating the Rad51 recombinase activity in the tumor suppressor protein Brca2 (breast cancer type 2 susceptibility protein). 39) Exceptionally, only Drosophila RecQ5 contains 2 putative BRCv repeats; whereas BRCv is present as a single copy in RecQ5 from most species.…”

Section: Discussionmentioning

confidence: 99%

RecQ5 Interacts with Rad51 and Is Involved in Resistance of <i>Drosophila</i> to Cisplatin Treatment

Maruyama¹,

Ohkita

Nakayama

et al. 2012

Biological & Pharmaceutical Bulletin

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RecQ5 is a member of the RecQ family of DNA helicases. There are 5 RecQ members in humans. Defects in 3 of them, i.e., BLM, WRN, and RTS, cause Bloom, Werner, and Rothmund-Thomson syndromes, respectively. RECQL1 and RECQL5 have not been associated with any human disease, and their precise roles are unknown. Our previous study suggests that the lack of RecQ5, which is the Drosophila homolog of RECQL5, leads to the accumulation of DNA double-stranded breaks (DSBs). It is possible that RecQ5 is involved in DSB repair. However, little is known about this possible function of RecQ5 in DSB repair. Here, we report that Rad51 protein, which plays a critical role in DSB repair, interacted with RecQ5 in vitro and in vivo in Drosophila. The Rad51 protein interacted with the C-terminal region of RecQ5, as shown by the yeast two-hybrid method. Moreover, the C-terminal region of the RecQ5 protein and the central region of Rad51 interacted directly and specifically when examined by the glutathione-S-transferase pull-down method. Consistent with these results, when RecQ5 and Rad51 were co-expressed in Drosophila cells in culture, they became co-localized in nuclei and could be co-immunoprecipitated. Furthermore, RecQ5-deficient flies (recq5) were more sensitive to the chemotherapeutic agent cisplatin compared with wild-type ones. Also, Rad51 mutants (rad51) were more sensitive to cisplatin, with sensitivity similar to that of recq5 rad51 double mutants. These data suggest that RecQ5 and Rad51 in Drosophila functioned for survival after the flies had been treated with cisplatin.Key words RecQ; Rad51; cisplatin; repair; DNA damage; Drosophila RecQ5 is a member of the RecQ family of DNA helicases, in which there are 5 members in humans. Defects in 3 of them, i.e., BLM, WRN, and RTS, cause Bloom, Werner, and Rothmund-Thomson syndromes, respectively, in humans. Genome instability and predisposition to cancer is a common feature of many RecQ helicase-deficient cells. Although RecQL1 and RecQL5 (human homolog of Drosophila RecQ5), the other 2 members of the RecQ family in humans, have not been associated with any human genetic diseases, Recql5-knock out mice show susceptibility to cancer, 1) suggesting that RecQ5 may also play an important role in preventing cancer. However, the precise role of RecQ5 remains unknown.In Drosophila, the frequencies of spontaneous and induced chromosomal aberrations are increased in RecQ5-mutant neuroblasts.2) These data imply that double-stranded break (DSB) damage to DNA accumulates spontaneously in RecQ5 mutants.2) Furthermore, the loss of maternally-derived RecQ5 leads to spontaneous mitotic defects in syncytial embryos. These mitotic defects are derived from anaphase DNA bridges, which link pairs of daughter nuclei. These nuclei concurrently exit from the cycle and are eliminated by Drosophila checkpoint kinase 2 tumor suppressor homolog (DmChk2)-dependent centrosome inactivation.3) DmChk2 responds to DSB DNA lesions. 4) These findings suggest that the lack of RecQ5 results in DSBs, which are recogn...

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A Variant of the Breast Cancer Type 2 Susceptibility Protein (BRC) Repeat Is Essential for the RECQL5 Helicase to Interact with RAD51 Recombinase for Genome Stabilization

Cited by 45 publications

References 40 publications

Differential and Concordant Roles for Poly(ADP-Ribose) Polymerase 1 and Poly(ADP-Ribose) in Regulating WRN and RECQL5 Activities

Differential and Concordant Roles for Poly(ADP-Ribose) Polymerase 1 and Poly(ADP-Ribose) in Regulating WRN and RECQL5 Activities

Unloading of homologous recombination factors is required for restoring double‐stranded DNA at damage repair loci

RecQ5 Interacts with Rad51 and Is Involved in Resistance of <i>Drosophila</i> to Cisplatin Treatment

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