XRCC4 in G1 suppresses homologous recombination in S/G2, in G1 checkpoint-defective cells

Saintigny, Yannick; Delacôte, Fabien; Boucher, Didier; Averbeck, D.; Lopez, Bernard S.

doi:10.1038/sj.onc.1210075

Cited by 32 publications

(27 citation statements)

References 61 publications

(92 reference statements)

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“…Importantly, expression of kdAKT1 did not induce any change in RAD51 foci formation when compared to the parental cells (Figure 2d). Because IR-induced RAD51 foci formation is cell-cycle dependent, we analyzed the cell-cycle distribution of irradiated cells by flow cytometry (Saintigny et al, 2007; Supplementary data S3). We did not observe any significant differences in the cell-cycle distributions of the different clones up to 24 h after exposure to 6 Gy IR (Supplementary data S3).…”

Section: Resultsmentioning

confidence: 99%

AKT1 represses gene conversion induced by different genotoxic stresses and induces supernumerary centrosomes and aneuploidy in hamster ovary cells

Plo

Lopez

2009

Oncogene

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The oncogenic kinase AKT1 is frequently overexpressed or activated in sporadic breast and ovarian cancers. In human breast tumors, we have previously shown that AKT1 represses homologous recombination (HR) induced by one double-strand break (DSB). To further analyze the impact of AKT1 on HR, we ectopically expressed wildtype or mutant forms of AKT1 in a hamster ovary cell line containing an intrachromosomal substrate for monitoring HR. In this cell line, AKT1 repressed HR induced by different genotoxic stresses including ionizing radiation, UV-C and one single DSB introduced into the intrachromosomal substrate. Consistently, AKT1 disrupted RAD51 foci formation, showing that AKT1 specifically affects gene conversion. Concomitantly, AKT1 represses both BRCA1 foci formation and HR stimulation resulting from BRCA1 overexpression, showing that AKT1 affects BRCA1-mediated HR functions, also in another species (hamster) and in another type of cell tissue (ovary cells). Finally, consistent with the HR defects, active AKT1 expression induces supernumerary centrosomes and aneuploidy. In addition to its impact on cell proliferation and apoptosis, the present data propose a novel oncogenic function for AKT1, by producing genomic instability as a consequence of HR repression.

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Section: Resultsmentioning

confidence: 99%

AKT1 represses gene conversion induced by different genotoxic stresses and induces supernumerary centrosomes and aneuploidy in hamster ovary cells

Plo

Lopez

2009

Oncogene

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“…Consequently, NHEJ is often thought to act preferentially in the G 1 phase. However, this increased sensitivity might be due to the inability of homologous recombination to act in G 1 , and thus to compensate for NHEJ deficiency (Saintigny et al, 2007). In addition, NHEJ has also been proposed to act in S phase (Stamato et al, 1988;Saintigny et al, 2001;Couedel et al, 2004;Mills et al, 2004) and throughout the cell cycle (Rothkamm et al, 2003).…”

Section: Introductionmentioning

confidence: 99%

S-phase progression stimulates both the mutagenic KU-independent pathway and mutagenic processing of KU-dependent intermediates, for nonhomologous end joining

2007

Self Cite

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We used intrachromosomal substrates to directly monitor the effect of the cell cycle on the efficiency and the accuracy of nonhomologous end joining (NHEJ) in mammalian cells. We show that both KU and KU-independent (KU-alt) pathways are efficient when maintaining cells in G 1 /S, in G 2 /M or during dynamic progression through S phase. In addition, the accuracy of NHEJ is barely altered when the cells are blocked in G 1 /S or in G 2 /M. However, progression through S phase increases the frequency of deletions, which is a hallmark of the KUalt pathway. Moreover, we show that the intermediates that are generated by the KU-dependent end joining of non-fully complementary ends, and which contain mismatches, nicks or gap intermediates, are less accurately processed when the cells progress through S phase. In conclusion, both KU and KU-alt processes are active throughout the cell cycle, but the repair is more error prone during S phase, both by increasing the mutagenic KU-alt pathway and decreasing the accuracy of the repair of the intermediates generated by the KU-dependent pathway.

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“…Thus, the ensuing XRCC4-alt DSB repair pathways might culminate in homologous recombination (22,23) or XRCC4-alt NHEJ.…”

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confidence: 99%

Defects in XRCC4 and KU80 differentially affect the joining of distal nonhomologous ends

Guirouilh‐Barbat

Rass

Plo

et al. 2007

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

152

183

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XRCC4-null mice have a more severe phenotype than KU80-null mice. Here, we address whether this difference in phenotype is connected to nonhomologous end-joining (NHEJ). We used intrachromosomal substrates to monitor NHEJ of two distal doublestrand breaks (DSBs) targeted by I-SceI, in living cells. In xrcc4-defective XR-1 cells, a residual but significant end-joining process exists, which primarily uses microhomologies distal from the DSB. However, NHEJ efficiency was strongly reduced in xrcc4-defective XR-1 cells versus complemented cells, contrasting with KU-deficient xrs6 cells, which showed levels of end-joining similar to those of complemented cells. Nevertheless, sequence analysis of the repair junctions indicated that the accuracy of end-joining was strongly affected in both xrcc4-deficient and KU-deficient cells. More specifically, these data showed that the KU80/XRCC4 pathway is conservative and not intrinsically error-prone but can accommodate non-fully complementary ends at the cost of limited mutagenesis.double-strand break repair ͉ genome rearrangements D NA double-strand breaks (DSBs) are harmful lesions generated by a variety of endogenous or exogenous stresses, potentially leading to genomic rearrangement. Nonhomologous endjoining (NHEJ) is a prominent pathway for DSB repair (1). Canonical NHEJ involves the successive intervention of the KU80-KU70 heterodimer, DNA-PKcs-Artemis, and, finally, ligase IV (Lig4) associated with its cofactors XRCC4 and Cernunnos/Xlf (2, 3). KU-independent NHEJ (KU-alt) has been described in vitro in both acellular extracts and cultured cells (1, 4-6).Alternative, XRCC4-independent DSB repair pathways (XRCC4-alt) have also been described, using episomic plasmid in cultured cells, using pulse field gel electrophoresis, or in in vitro biochemical experiments (5-10). One hypothesis could propose that XRCC4 and KU are implicated in the same canonical NHEJ pathway, whereas the alternate pathway is independent of both KU and XRCC4. However, in transgenic mice, the inactivation of XRCC4 or Lig4 results in a more severe phenotype than the inactivation of KU (11), suggesting that XRCC4 might have an additional essential function; however, studies show that XRCC4 and Lig4 do not have roles outside of NHEJ, whereas in contrast, KU acts in other processes such as transcription, apoptosis, and responses to the cell microenvironment (12)(13)(14).Alternatively, these varying phenotypes in mice may actually result from differences in DSB repair efficiencies, indicating that defects in XRCC4 might be more deleterious for DSB repair than defects in KU. What challenges this hypothesis, however, is that substantial class switch recombination (CSR) has recently been shown to occur in mouse B cells without XRCC4, whereas no CSR was recorded in cells devoid of KU (15-18).The relative contributions of XRCC4 and KU80 versus the XRCC4-alt and KU-alt pathways, respectively, to DSB repair remain unclear in wild-type cells. Contrasting results were obtained in living cells, using an episomic plasm...

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XRCC4 in G1 suppresses homologous recombination in S/G2, in G1 checkpoint-defective cells

Cited by 32 publications

References 61 publications

AKT1 represses gene conversion induced by different genotoxic stresses and induces supernumerary centrosomes and aneuploidy in hamster ovary cells

AKT1 represses gene conversion induced by different genotoxic stresses and induces supernumerary centrosomes and aneuploidy in hamster ovary cells

S-phase progression stimulates both the mutagenic KU-independent pathway and mutagenic processing of KU-dependent intermediates, for nonhomologous end joining

Defects in XRCC4 and KU80 differentially affect the joining of distal nonhomologous ends

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