Structural Basis of Poly(ADP-ribose) Recognition by the Multizinc Binding Domain of Checkpoint with Forkhead-associated and RING Domains (CHFR)

Oberoi, Jasmeen; Richards, Mark W.; Crumpler, Simon; Brown, Nathan; Blagg, Julian; Bayliss, Richard

doi:10.1074/jbc.m110.159855

Cited by 57 publications

(56 citation statements)

References 28 publications

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“…We first expressed and purified the recombinant GST-ECT2 BRCT domain in vitro and performed in vitro binding assay by incubating recombinant GST-ECT2 BRCT with PAR. Since CHFR directly interacts with PAR via its C-terminal PBZ motif, [16][17] we used GST and recombinant CHFR as the negative and positive control in this binding assay, respectively. Purified recombinant proteins were dot-blotted on the nitrocellulose membrane after incubation with PAR.…”

Section: Resultsmentioning

confidence: 99%

Poly(ADP-ribosyl)ation is recognized by ECT2 during mitosis

Li¹,

Bian²,

Yu³

2014

Cell Cycle

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Poly(ADP-ribosyl)ation is an unique posttranslational modification and required for spindle assembly and function during mitosis. However, the molecular mechanism of poly(ADP-ribose) (PAR) in mitosis remains elusive. Here, we show the evidence that PAR is recognized by ECT2, a key guanine nucleotide exchange factor in mitosis. The BRCT domain of ECT2 directly binds to PAR both in vitro and in vivo. We further found that a-tubulin is PARylated during mitosis. PARylation of a-tubulin is recognized by ECT2 and recruits ECT2 to mitotic spindle for completing mitosis. Taken together, our study reveals a novel mechanism by which PAR regulates mitosis.

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Section: Resultsmentioning

confidence: 99%

Poly(ADP-ribosyl)ation is recognized by ECT2 during mitosis

Li¹,

Bian²,

Yu³

2014

Cell Cycle

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“…His-tagged human PARP1 was expressed in bacteria and purified by Ni-NTA affinity resin. PAR was synthesized and purified as described previously except for the following modifications (49). PAR was synthesized in a 20-mL incubation mixture containing 100 mM Tris·HCl pH 7.8, 10 mM MgCl 2 , 1 mM NAD + , 10 mM DTT, 60 μg calf thymus histone, 50 μg octameric oligonucleotide GGAATTCC, and 2 mg PARP1.…”

Section: Methodsmentioning

confidence: 99%

The oligonucleotide/oligosaccharide-binding fold motif is a poly(ADP-ribose)-binding domain that mediates DNA damage response

Zhang

Chen

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Oligonucleotide/oligosaccharide-binding (OB) fold is a ssDNA or RNA binding motif in prokaryotes and eukaryotes. Unexpectedly, we found that the OB fold of human ssDNA-binding protein 1 (hSSB1) is a poly(ADP ribose) (PAR) binding domain. hSSB1 exhibits highaffinity binding to PAR and recognizes iso-ADP ribose (ADPR), the linkage between two ADPR units. This interaction between PAR and hSSB1 mediates the early recruitment of hSSB1 to the sites of DNA damage. Mutations in the OB fold of hSSB1 that disrupt PAR binding abolish the relocation of hSSB1 to the sites of DNA damage. Moreover, PAR-mediated recruitment of hSSB1 is important for early DNA damage repair. We have screened other OB folds and found that several other OB folds also recognize PAR. Taken together, our study reveals a PAR-binding domain that mediates DNA damage repair.G enomic integrity is constantly challenged by various types of DNA damage, which are induced by DNA replication errors, environmental hazards, and other genotoxic stress. In response to this stress, cells activate an evolutionarily conserved pathway termed the DNA damage response (DDR), to orchestrate various cellular responses for maintaining genomic stability (1-3). It has been shown that poly(ADP ribosyl)ation plays an important role in DDR (4-6). Upon DNA damage induction, poly(ADP ribose) (PAR) polymerases (PARPs), such as PARP1, the founding member of the PARP family, rapidly detect DNA breaks, including single-strand breaks (SSBs) and double-strand breaks (DSBs), and activate PAR synthesis at or adjacent to DNA lesions (7,8). Recent evidence suggests that DNA damage-induced PAR may serve as a docking signal to recruit DDR factors to DNA lesions (7-14). For example, our recent study showed that the BRCT domains of BARD1 and NBS1 recognize PAR, which facilitates the rapid recruitment of the BRCA1/BARD1 complex and NBS1 to the site of DNA damage (15, 16). These results indicate that other DDR factors may also be recruited to DNA damage sites by PAR. Thus, it is important to identify other "readers" of PAR to elucidate the molecular mechanism of PAR in DDR.Previous studies have shown that human ssDNA-binding protein 1 (hSSB1) plays a key role in . hSSB1 is a 211-residue polypeptide containing an oligonucleotide/oligosaccharide-binding (OB) fold at the N terminus. Following DNA damage, hSSB1 quickly relocates to DNA damage sites and regulates foci formation of other DNA damage repair proteins, such as BRCA1 and RAD51 (17,(21)(22)(23)(24). Thus, depletion of hSSB1 impairs the repair of DSBs. In response to DNA damage, hSSB1 is phosphorylated by ATM and regulates ATM-dependent cell cycle checkpoint activation (17,(21)(22)(23)(24). By protein affinity purifications, hSSB1 was shown to tightly associate with other proteins, such as INTS3, forming a multisubunit complex (21-24). INTS3 is a well-folded protein and previously identified as a member in the INTS complex that regulates RNA processing and gene transcription (25). Like hSSB1, INTS3 is also quickly relocated to DNA lesions in r...

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“…Since CHFR and APLF are known PAR-binding proteins, which bind PAR via their PBZ motifs (Ahel et al 2008;Li et al 2010;Oberoi et al 2010), we used GST, recombinant CHFR, and APLF PBZ motifs as negative and positive controls, respectively, to screen PAR-binding domains. We examined 19 FHA or BRCT domains and found that two FHA domains (from PNKP and APTX), two BRCT domains (from Ligase4 and XRCC1), and an FHA-BRCT fusion domain (from NBS1) interacted with PAR (Fig.…”

Section: A Set Of Brct and Fha Domains Binds Parmentioning

confidence: 99%

The FHA and BRCT domains recognize ADP-ribosylation during DNA damage response

et al. 2013

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Poly-ADP-ribosylation is a unique post-translational modification participating in many biological processes, such as DNA damage response. Here, we demonstrate that a set of Forkhead-associated (FHA) and BRCA1 C-terminal (BRCT) domains recognizes poly(ADP-ribose) (PAR) both in vitro and in vivo. Among these FHA and BRCT domains, the FHA domains of APTX and PNKP interact with iso-ADP-ribose, the linkage of PAR, whereas the BRCT domains of Ligase4, XRCC1, and NBS1 recognize ADP-ribose, the basic unit of PAR. The interactions between PAR and the FHA or BRCT domains mediate the relocation of these domain-containing proteins to DNA damage sites and facilitate the DNA damage response. Moreover, the interaction between PAR and the NBS1 BRCT domain is important for the early activation of ATM during DNA damage response and ATM-dependent cell cycle checkpoint activation. Taken together, our results demonstrate two novel PAR-binding modules that play important roles in DNA damage response.

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Structural Basis of Poly(ADP-ribose) Recognition by the Multizinc Binding Domain of Checkpoint with Forkhead-associated and RING Domains (CHFR)

Cited by 57 publications

References 28 publications

Poly(ADP-ribosyl)ation is recognized by ECT2 during mitosis

Poly(ADP-ribosyl)ation is recognized by ECT2 during mitosis

The oligonucleotide/oligosaccharide-binding fold motif is a poly(ADP-ribose)-binding domain that mediates DNA damage response

The FHA and BRCT domains recognize ADP-ribosylation during DNA damage response

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