Association of BRCA1 with the hRad50-hMre11-p95 complex and the DNA damage response

Department of Molecular Medicine, Institute of Biotechnology, University of Texas Health Science Center at San Antonio, 15355 Lambda Drive, San Antonio, TX 78245, USA.
Science (Impact Factor: 33.61). 08/1999; 285(5428):747-50.
Source: PubMed


BRCA1 encodes a tumor suppressor that is mutated in familial breast and ovarian cancers. Here, it is shown that BRCA1 interacts in vitro and in vivo with hRad50, which forms a complex with hMre11 and p95/nibrin. Upon irradiation, BRCA1 was detected in discrete foci in the nucleus, which colocalize with hRad50. Formation of irradiation-induced foci positive for BRCA1, hRad50, hMre11, or p95 was dramatically reduced in HCC/1937 breast cancer cells carrying a homozygous mutation in BRCA1 but was restored by transfection of wild-type BRCA1. Ectopic expression of wild-type, but not mutated, BRCA1 in these cells rendered them less sensitive to the DNA damage agent, methyl methanesulfonate. These data suggest that BRCA1 is important for the cellular responses to DNA damage that are mediated by the hRad50-hMre11-p95 complex.

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Available from: Chi-Fen Chen, Jan 07, 2014
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    • "However, their enhanced association at G2 suggests a function during late replication or during recombination-associated events at the telomere [38], [66]. While BRCA1 is essential for recruitment of recombination proteins to promote strand processing and invasion, and to form recombination intermediates, its recruitment to telomeres requires RAD50 during S-phase [42], [53], [61]. Our data indicate that BLM is also part of stable complexes with RAD50, albeit predominantly in G2 and suggest that BLM-BRCA1 complexes may be distinct from BRCA1-RAD50 complexes required during recombination initiation (Figures 5–8). "
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    ABSTRACT: Fifteen percent of tumors utilize recombination-based alternative lengthening of telomeres (ALT) to maintain telomeres. The mechanisms underlying ALT are unclear but involve several proteins involved in homologous recombination including the BLM helicase, mutated in Bloom's syndrome, and the BRCA1 tumor suppressor. Cells deficient in either BLM or BRCA1 have phenotypes consistent with telomere dysfunction. Although BLM associates with numerous DNA damage repair proteins including BRCA1 during DNA repair, the functional consequences of BLM-BRCA1 association in telomere maintenance are not completely understood. Our earlier work showed the involvement of BRCA1 in different mechanisms of ALT, and telomere shortening upon loss of BLM in ALT cells. In order to delineate their roles in telomere maintenance, we studied their association in telomere metabolism in cells using ALT. This work shows that BLM and BRCA1 co-localize with RAD50 at telomeres during S- and G2-phases of the cell cycle in immortalized human cells using ALT but not in cells using telomerase to maintain telomeres. Co-immunoprecipitation of BRCA1 and BLM is enhanced in ALT cells at G2. Furthermore, BRCA1 and BLM interact with RAD50 predominantly in S- and G2-phases, respectively. Biochemical assays demonstrate that full-length BRCA1 increases the unwinding rate of BLM three-fold in assays using a DNA substrate that models a forked structure composed of telomeric repeats. Our results suggest that BRCA1 participates in ALT through its interactions with RAD50 and BLM.
    PLoS ONE 08/2014; 9(8):e103819. DOI:10.1371/journal.pone.0103819 · 3.23 Impact Factor
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    • "Through its interaction with BRCA2/Rad51, BRCA1 promotes HR, which often occurs in proliferating cells [23], [24]. In addition, BRCA1 is responsible for NHEJ by associating with the Ku80 RAD50-MRE11-NBS1 complex through its BRCT domains at the C-terminus [16], [25]. Data from our previous study also confirmed that BRCA1 phosphorylation regulates the fidelity of NHEJ by checkpoint kinase 2 [9]. "
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    ABSTRACT: Retinal post-mitotic neurocytes display genomic instability after damage induced by physiological or pathological factors. The involvement of BRCA1, an important factor in development and DNA repair in mature retinal neurocytes remains unclear. Thus, we investigated the developmental expression profile of BRCA1 in the retina and defined the role of BRCA1 in DNA repair in retinal neurocytes. Our data show the expression of BRCA1 is developmentally down-regulated in the retinas of mice after birth. Similarly, BRCA1 is down-regulated after differentiation induced by TSA in retinal precursor cells. An end-joining activity assay and DNA fragmentation analysis indicated that the DNA repair capacity is significantly reduced. Moreover, DNA damage in differentiated cells or cells in which BRCA1 is silenced by siRNA interference is more extensive than that in precursor cells subjected to ionizing radiation. To further investigate non-homologous end joining (NHEJ), the major repair pathway in non-divided neurons, we utilized an NHEJ substrate (pEPI-NHEJ) in which double strand breaks are generated by I-SceI. Our data showed that differentiation and the down-regulation of BRCA1 respectively result in a 2.39-fold and 1.68-fold reduction in the total NHEJ frequency compared with that in cells with normal BRCA1. Furthermore, the analysis of NHEJ repair junctions of the plasmid substrate indicated that BRCA1 is involved in the fidelity of NHEJ. In addition, as expected, the down-regulation of BRCA1 significantly inhibits the viability of retina precursor cells. Therefore, our data suggest that BRCA1 plays a critical role in retinal development and repairs DNA damage of mature retina neurocytes.
    PLoS ONE 06/2014; 9(6):e99371. DOI:10.1371/journal.pone.0099371 · 3.23 Impact Factor
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    • "Humans that have a haploinsufficiency for the gene encoding TFII-I are characterised by a craniofacial phenotype along with cognitive deficits (Pober, 2010). The carboxyl terminus of BRCA1, referred to as the BRCT domain, possesses autonomic transcriptional activation functions, and the BRCT domain has been shown to be involved in DNA double strand damage repair and homologous recombination (HR) (Zhong et al, 1999). We previously reported that TFII-I stimulates the transactivation function of the BRCT domain and the BRCA1-mediated stimulation of SIRT1 promoter activity in the mammalian homologue of yeast Sir2 (silent information regulator 2) (Tanikawa et al, 2011). "
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    ABSTRACT: Background: In multicellular organisms, precise control of cell cycle and the maintenance of genomic stability are crucial to prevent chromosomal alterations. The accurate function of the DNA damage pathway is maintained by DNA repair mechanisms including homologous recombination (HR). Herein, we show that both TFII-I and DBC1 mediate cellular mechanisms of cell-cycle regulation and DNA double strand damage repair. Methods: Regulation of cell cycle by TFII-I and DBC1 was investigated using Trypan blue dye exclusion test, luciferase assay, and flow cytometry analysis. We also analysed the role of TFII-I and DBC1 in DNA double strand damage repair after irradiation by immunofluorescence study, clonogenicity assay, and HR assay. Results: Flow cytometry analysis revealed a novel function that siRNA-mediated knockdown of endogenous DBC1 resulted in G2/M phase arrest. We also have shown that both endogenous TFII-I and DBC1 activate DNA repair mechanisms after irradiation because irradiation-induced foci formation of TFII-I-γH2AX was observed, and the depletion of endogenous TFII-I or DBC1 resulted in the inhibition of normal HR efficiency. Conclusion: These results reveal novel mechanisms by which TFII-I and DBC1 can modulate cellular fate by affecting cell-cycle control as well as HR pathway.
    British Journal of Cancer 11/2013; 109(12). DOI:10.1038/bjc.2013.532 · 4.84 Impact Factor
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