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Cell polarity protein Par3 complexes with DNA-PK via Ku70 and regulates DNA double-strand break repair

机译：细胞极性蛋白Par3通过Ku70与DNA-PK复合并调节DNA双链断裂修复

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摘要

The partitioning-defective 3 (Par3), a key component in the conserved Par3/Par6/aPKC complex, plays fundamental roles in cell polarity. Herein we report the identification of Ku70 and Ku80 as novel Par3-interacting proteins through an in vitro binding assay followed by liquid chromatography-tandem mass spectrometry. Ku70/Ku80 proteins are two key regulatory subunits of the DNA-dependent protein kinase (DNA-PK), which plays an essential role in repairing double-strand DNA breaks (DSBs). We determined that the nuclear association of Par3 with Ku70/Ku80 was enhanced by y-irradiation (IR), a potent DSB inducer. Furthermore, DNA-PKcs, the catalytic subunit of DNA-PK, interacted with the Par3/Ku70/Ku80 complex in response to IR. Par3 over-expression or knockdown was capable of up- or downregulat-ing DNA-PK activity, respectively. Moreover, the Par3 knockdown cells were found to be defective in random plasmid integration, defective in DSB repair following IR, and radiosensitive, phenotypes similar to that of Ku70 knockdown cells. These findings identify Par3 as a novel component of the DNA-PK complex and implicate an unexpected link of cell polarity to DSB repair.
机译：缺陷分区3（Par3）是保守的Par3 / Par6 / aPKC复合体中的关键成分，在细胞极性中起着基本作用。在这里，我们报告通过体外结合测定，然后通过液相色谱-串联质谱法鉴定Ku70和Ku80为新型Par3相互作用蛋白。 Ku70 / Ku80蛋白是DNA依赖性蛋白激酶（DNA-PK）的两个关键调节亚基，在修复双链DNA断裂（DSB）中起着至关重要的作用。我们确定通过有效的DSB诱导物y辐射（IR）增强了Par3与Ku70 / Ku80的核缔合。此外，DNA-PKcs，DNA-PK的催化亚基，响应于IR与Par3 / Ku70 / Ku80复合物相互作用。 Par3过表达或敲低分别能够上调或下调DNA-PK活性。此外，发现Par3敲低细胞在随机质粒整合中存在缺陷，在IR后DSB修复中存在缺陷，并且具有与Ku70敲低细胞相似的放射敏感性表型。这些发现将Par3鉴定为DNA-PK复合物的新成分，并暗示了细胞极性与DSB修复的意想不到的联系。

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