Evaluation of the functional impact of germline BRCA1 variants that are likely to be associated to breast and ovarian cancer could help to investigate the mechanism of BRCA1 tumorigenesis. Expression of pathogenic BRCA1 missense variants increased homologous recombination (HR) and gene reversion (GR) in yeast. We thought to exploit yeast genetics to shed light on BRCA1-induced genome instability and tumorigenesis. We determined the effect on GR of several neutral and pathogenic BRCA1 variants in the yeast strain RSY6wt and its isogenic DSB repair mutants, such as mre11∆, rad50∆ and rad51∆. In the RSY6wt, four out of five pathogenic and two out of six neutral variants significantly increased GR; rad51∆ strain, the pathogenic variants C61G and A1708E induced a weak but significant increase in GR. On the other hand, in rad50∆ mutant expressing the pathogenic variants localised at the BRCT domain, a further GR increase was seen. The neutral variant N132K and the VUS A1789T induced a weak GR increase in mre11∆ mutant. Thus, BRCA1 missense variants require specific genetic functions and presumably induced GR by different mechanisms. As DNA repair is regulated by cell cycle, we determined the effect on GR of BRCA1 variants in cell cycle-arrested RSYwt cells. GR is highly BRCA1-inducible in S-phase-arrested cells as compared to G1 or G2. Sequence analysis of genomic DNA from ILV1 revertant clones showed that BRCA1-induced ilv1-92 reversion by base substitution when GR is at least 6-fold over the control. Our study demonstrated that BRCA1 may interfere with yeast DNA repair functions that are active in S-phase causing high level of GR. In addition, we confirmed here that yeast could be a reliable model to investigate the mechanism and genetic requirements of BRCA1-induced genome instability. Finally, developing yeast-based assays to characterise BRCA1 missense variants could be useful to design more precise therapies.

中文翻译：

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BRCA1错义变体对酿酒酵母DNA双链断裂修复突变体和细胞周期阻滞细胞基因回复的影响。

评估可能与乳腺癌和卵巢癌有关的种系BRCA1变体的功能影响可能有助于研究BRCA1肿瘤发生的机制。致病性BRCA1错义变体的表达增加了酵母中的同源重组（HR）和基因回复（GR）。我们认为利用酵母遗传学来阐明BRCA1诱导的基因组不稳定性和肿瘤发生。我们确定了酵母菌株RSY6wt及其同基因DSB修复突变体中的几个中性和致病性BRCA1变体对GR的影响，例如mre11∆，rad50∆和rad51∆。在RSY6wt中，五分之四的病原体和六分之二的中性变体显着增加了GR。rad51∆菌株，致病性变体C61G和A1708E诱导了GR的弱但显着增加。另一方面，在表达定位于BRCT结构域的致病变异的rad50Δ突变体中，GR进一步增加。中性变体N132K和VUS A1789T导致mre11Δ突变体的GR弱增加。因此，BRCA1错义变体需要特定的遗传功能，并可能通过不同的机制诱导GR。由于DNA修复受细胞周期调控，因此我们确定了细胞周期停滞的RSYwt细胞对BRCA1变体GR的影响。与G1或G2相比，GR在S期停滞细胞中高度BRCA1诱导。来自ILV1回复克隆的基因组DNA的序列分析表明，当GR至少比对照高6倍时，BRCA1诱导的碱基取代引起的ilv1-92回复。我们的研究表明，BRCA1可能会干扰酵母DNA修复功能（在S期中起作用），从而导致高水平的GR。此外，我们在这里证实酵母可能是研究BRCA1诱导的基因组不稳定的机制和遗传学要求的可靠模型。最后，开发基于酵母的测定法来表征BRCA1错义变体可能对设计更精确的疗法有用。