Isolation of genomic instability genes that are associated with malignancy

在对158例肺癌和10例正常肺的5种混合物的检查中，细胞周期和检查点相关基因通常显示癌症中的mRNA表达增加，而POLD 4显示小细胞肺癌（SCLC）中的mRNA减少。在缺乏POLD 4的情况下，我们观察到DNA修复活性的缺陷、细胞周期的延迟和染色体断裂诱导频率的增加。在siRNA靶位点携带沉默突变的工程化POLD 4的过表达拯救了这些表型，牢固地确立了POLD 4在这些效应中的作用。为了鉴定与POLD 4相互作用的基因产物，我们使用具有POLD 4的免疫沉淀蛋白质级分进行了MS分析，其中检测到ATM、CDC 14和其他细胞周期检查点/进展蛋白。我们进一步鉴定了肺癌细胞系中新的单核苷酸变异（SNV）。其中一个SNV降低了与pol δ催化亚基的物理相互作用能力。这些结果表明，新发现的SNV可能通过改变DNA聚合酶δ的DNA复制能力而参与肿瘤的发生。

In examinations of 158 lung cancers and 5 mixtures of 10 normal lungs, cell cycle- and checkpoint-related genes generally showed mRNA expression increases in cancer, whereas POLD4 showed reduced mRNA in small cell lung cancer (SCLC). In the absence of POLD4, we observed defect in DNA repair activity, delay in cell cycle, and increased frequency of chromosomal break induction. Overexpression of an engineered POLD4 carrying silent mutations at the siRNA target site rescued these phenotypes, firmly establishing the role of POLD4 in these effects. In order to identify gene products that interact with POLD4, we performed an MS analysis using the immunoprecipitated protein fractions with POLD4, where ATM, CDC14 and other cell cycle checkpoint/progression proteins were detected. We further identified novel single nucleotide variations (SNV) in lung cancer cell lines. One of the SNV reduced the physical interaction ability with the catalytic subunit of pol delta. These results suggest that the newly identified SNV could be involved in the carcinogenesis by modifying DNA replication ability of DNA polymerase delta.