Section of Genetic Oncology, University Hospital and University of Pisa, Pisa, Italy.
Breast Cancer Res Treat. 2011 Jan;125(1):265-72. doi: 10.1007/s10549-010-1112-8. Epub 2010 Aug 25.
Many missense variants in BRCA1 are of unclear clinical significance. Functional and genetic approaches have been proposed for elucidating the clinical significance of such variants. The purpose of this study was to evaluate BRCA1 missense variants for their effect on both homologous recombination (HR) and non homologous end joining (NHEJ). HR frequency evaluation: HeLaG1 cells, containing a stably integrated plasmid that allows us to measure HR events by gene conversion events, were transfected with the pcDNA3β expression vector containing the BRCA1-wild-type (BRCA1 wild type) or the BRCA1-unclassified variants (BRCA1-UCVs). The NHEJ was measured by a random plasmid integration assay. The assays suggested a BRCA1 involvement mainly in the NHEJ. As a matter of fact, the Y179C and the A1789T variant significantly altered the NHEJ activity as compared to the wild type, suggesting that they may be related to BRCA1-associated pathogenicity by affecting this function. The variants N550H and I1766S, and the mutation M1775R did not alter the NHEJ frequency. These data, besides proposing a method for the study of BRCA1 variants' effect on HR and NHEJ, highlighted the need for a range of functional assays to be performed to identify variants with altered function.
许多 BRCA1 中的错义变异体的临床意义尚不清楚。已经提出了功能和遗传方法来阐明此类变异体的临床意义。本研究的目的是评估 BRCA1 错义变异体对同源重组 (HR) 和非同源末端连接 (NHEJ) 的影响。HR 频率评估：含有允许我们通过基因转换事件测量 HR 事件的稳定整合质粒的 HeLaG1 细胞，用含有 BRCA1-野生型 (BRCA1 野生型) 或 BRCA1-未分类变异体 (BRCA1-UCV) 的 pcDNA3β 表达载体转染。NHEJ 通过随机质粒整合测定法进行测量。这些测定结果表明 BRCA1 主要参与 NHEJ。事实上，与野生型相比，Y179C 和 A1789T 变异体显著改变了 NHEJ 活性，表明它们可能通过影响该功能而与 BRCA1 相关的致病性有关。变异体 N550H 和 I1766S 以及突变 M1775R 并未改变 NHEJ 频率。除了提出一种用于研究 BRCA1 变异体对 HR 和 NHEJ 影响的方法外，这些数据还强调需要进行一系列功能测定以识别具有改变功能的变异体。
