BRCA2: a universal recombinase regulator

Thorslund, Tina; West, Stephen C.

doi:10.1038/sj.onc.1210870

Cited by 141 publications

(128 citation statements)

References 59 publications

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“…A more clearly defined function for Brca2 involves its mediator activity for Rad51 nucleoprotein filament assembly at the site of a DNA break (12,(62)(63)(64)(65)(66). Following exposure of Capan-1 cells to IR, we observed a modest increase in chromatin-associated Rad51 (Fig.…”

Section: Discussionmentioning

confidence: 83%

Cellular Redistribution of Rad51 in Response to DNA Damage

Gildemeister

Sage

Knight

2009

Journal of Biological Chemistry

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Exposure of cells to DNA-damaging agents results in a rapid increase in the formation of subnuclear complexes containing Rad51. To date, it has not been determined to what extent DNA damage-induced cytoplasmic to nuclear transport of Rad51 may contribute to this process. We have analyzed subcellular fractions of HeLa and HCT116 cells and found a significant increase in nuclear Rad51 levels following exposure to a modest dose of ionizing radiation (2 grays). We also observed a DNA damageinduced increase in nuclear Rad51 in the Brca2-defective cell line Capan-1. To address a possible Brca2-independent mechanism for Rad51 nuclear transport, we analyzed subcellular fractions for two other Rad51-interacting proteins, Rad51C and Xrcc3. Rad51C has a functional nuclear localization signal, and although we found that the subcellular distribution of Xrcc3 was not significantly affected by DNA damage, there was a damageinduced increase in nuclear Rad51C. Furthermore, RNA interference-mediated depletion of Rad51C in HeLa and Capan-1 cells resulted in lower steady-state levels of nuclear Rad51 as well as a diminished DNA damage-induced increase. Our results provide important insight into the cellular regulation of Rad51 nuclear entry and a role for Rad51C in this process.Cellular surveillance of genome integrity and repair of DNA damage are essential processes that ensure proper development and survival of all organisms. DNA double-strand breaks (DSBs) 2 are a particularly deleterious form of genome damage and occur following exposure of cells to exogenous mutagens as well as during normal metabolic processes, e.g. antigen receptor gene rearrangement, restart of stalled replication forks, formation of meiotic DNA crossovers, etc. (1, 2). Mammalian cells use two distinct mechanisms for repair of DSBs, non-homologous end joining and homologous recombination (HR). Processes requiring imprecise DNA repair, such as the creation of antibody diversity, exploit the error-prone non-homologous end joining mechanism. In contrast, HR is an error-free DNA repair pathway and is critical for avoidance of unwanted genetic changes during the meiotic exchange of information between paternal and maternal alleles and for error-free repair of broken chromosomes (3). Rad51 is the central enzymatic component of HR. Upon its regulated recruitment to sites of DNA breaks, Rad51 forms a nucleoprotein filament by polymerizing onto single-stranded DNA at the processed break. This filament catalyzes DNA strand exchange with an undamaged sister chromatid or homologous chromosome, which serves as a template for the restoration of missing genetic information (3, 4).Visible nuclear Rad51 clusters, or foci, form during S-phase and appear to localize to sites of replicating DNA (5-7). A dramatic increase in the number and size of nuclear Rad51 foci is a hallmark of the early cellular response to genomic insult (7-10). The appearance of DNA damage-induced nuclear Rad51 foci is blocked in cells with deficiencies in several HR-related proteins, including Brca2 ...

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Section: Discussionmentioning

confidence: 83%

Cellular Redistribution of Rad51 in Response to DNA Damage

Gildemeister

Sage

Knight

2009

Journal of Biological Chemistry

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“…2 A and B), suggesting that DSB end resection is unaffected by heat. In subsequent steps of HR, the RAD51 recombinase forms nucleoprotein filaments on singlestranded DNA with the help of BRCA2 (1,15). Whereas RAD51 and BRCA2 IRIF could be detected 15 min post-irradiation in control cells, incubation at 41°C completely abrogated accumulation of both proteins at DSB sites ( Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Unless stated otherwise, cells were irradiated directly after incubation at elevated temperature by γ-rays from a cesium source ( 137 Cs, 0.70 Gy/min) or by α-particles as previously described (15). Cells from the cervix carcinoma biopsies were irradiated for 1 min with α-particles by placing coverslips with cells upside-down on a 1.8-μm-thick polyester membrane and irradiating from below, through the membrane.…”

Section: Methodsmentioning

confidence: 99%

Mild hyperthermia inhibits homologous recombination, induces BRCA2 degradation, and sensitizes cancer cells to poly (ADP-ribose) polymerase-1 inhibition

Krawczyk

Eppink²,

Essers³

et al. 2011

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

295

288

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Defective homologous recombination (HR) DNA repair imposed by BRCA1 or BRCA2 deficiency sensitizes cells to poly (ADP-ribose) polymerase (PARP)-1 inhibition and is currently exploited in clinical treatment of HR-deficient tumors. Here we show that mild hyperthermia (41-42.5°C) induces degradation of BRCA2 and inhibits HR. We demonstrate that hyperthermia can be used to sensitize innately HR-proficient tumor cells to PARP-1 inhibitors and that this effect can be enhanced by heat shock protein inhibition. Our results, obtained from cell lines and in vivo tumor models, enable the design of unique therapeutic strategies involving localized ondemand induction of HR deficiency, an approach that we term induced synthetic lethality.anti-cancer treatment | RAD51 | double-strand break M any anti-cancer therapies are based on cytotoxicity of DNA double strand breaks (DSBs) induced by ionizing radiation or, indirectly, by chemical agents. However, efficient DSB repair mechanisms protect cells from the genotoxic effects of DSBs, thereby reducing the effectiveness of the therapies. Two major pathways are involved in DSB repair in mammalian cells: homologous recombination (HR) and nonhomologous end joining (NHEJ). HR uses intact homologous DNA sequences, usually the sister chromatid in postreplicative chromatin, to faithfully restore DNA breaks (1), whereas NHEJ operates throughout the entire cell cycle and does not require a DNA template (2). Agents inhibiting DNA repair processes potentiate the cytotoxicity of DSBs in cancer therapy (3). Elevated temperature is one such agent that, via unclear mechanisms, interferes with multiple pathways of DNA repair (4-6) and is clinically applied (7). ResultsTo investigate if HR, among other processes and DSB repair pathways, is influenced by elevated temperature, we used an isogenic set of mouse embryonic stem (ES) cells that are either HR proficient (wild-type) or HR deficient (Rad54 −/− ) due to the disruption of the Rad54 gene, which is important for HR activity (1). We compared radiosensitization of these cells by incubating them at 37°C or 41°C before irradiation. For this and subsequent experiments we chose temperatures below 43°C, because they are relevant in clinical practice (8). Interestingly, we observed that wild-type but not Rad54 −/− cells were radiosensitized by preincubation at 41°C compared with cells incubated at 37°C (Fig. 1A). Similarly, HeLa cells, in which the important HR factors XRCC3 or BRCA2 were down-regulated using siRNA, were refractory to further temperature-mediated radiosensitization (Fig. 1B and Fig. S1). These results suggest that elevated temperature inactivates HR. To directly measure the effect of temperature on HR, we quantitated HR-mediated gene targeting in ES cells (9) and found that the efficiency of gene targeting was significantly reduced by preincubation at 41°C compared with 37°C (Fig. 1C). Similarly, preincubation at 41°C reduced the frequency of spontaneous and mitomycin C-induced sister chromatid exchanges in SW-1573 cells (Fig. S2A), w...

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“…BRCA2 regulates the function of the recombinase enzyme RAD51, which plays a crucial role in the DNA-repair pathway (18,19). Germ-line mutations in BRCA2 are associated with increased susceptibility to breast and ovarian cancers and are responsible for one-third of hereditary breast cancer (20,21).…”

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confidence: 99%

Mapping the physical and functional interactions between the tumor suppressors p53 and BRCA2

Rajagopalan

Andreeva

Rutherford

et al. 2010

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

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p53 maintains genome integrity either by regulating the transcription of genes involved in cell cycle, apoptosis, and DNA repair or by interacting with partner proteins. Here we provide evidence for a direct physical interaction between the tumor suppressors p53 and BRCA2. We found that the transactivation domain of p53 made specific interactions with the C-terminal oligonucleotide/oligosaccharide-binding-fold domains of BRCA2 (BRCA2 CTD ). A second distinct site situated on the p53 DNA-binding domain, bound to a region containing BRC repeats of BRCA2 (BRCA2 ½BRC1-8 ) and may contribute synergistically for high affinity association of intact full-length proteins. Overexpression of BRCA2 and BRCA2 CTD suppressed the transcriptional activity of p53 with a concomitant reduction in the expression of p53-target genes such as Bax and p21. Consequently, p53-mediated apoptosis was significantly attenuated by BRCA2. The observed physical association of p53 and BRCA2 may have important functional implications in the p53 transactivation-independent suppression of homologous recombination and suggests a possible interregulatory role for both proteins in apoptosis and DNA repair.

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BRCA2: a universal recombinase regulator

Cited by 141 publications

References 59 publications

Cellular Redistribution of Rad51 in Response to DNA Damage

Cellular Redistribution of Rad51 in Response to DNA Damage

Mild hyperthermia inhibits homologous recombination, induces BRCA2 degradation, and sensitizes cancer cells to poly (ADP-ribose) polymerase-1 inhibition

Mapping the physical and functional interactions between the tumor suppressors p53 and BRCA2

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