Ku70

Background: Ku70 participates in several pathological processes through mediating repair of DNA double-strand breaks. Our previous study has identified a highly conserved long noncoding RNA cardiac ischemia reperfusion associated Ku70 interacting lncRNA (CIRKIL) that was upregulated in myocardial infarction. The study aims to investigate whether CIRKIL regulates myocardial ischemia/reperfusion (I/R) through binding to Ku70. Methods: CIRKIL transgenic and knockout mice were subjected to 45-minute ischemia and 24-hour reperfusion to establish myocardial I/R model. RNA pull-down and RNA immunoprecipitation assay were used to detect the interaction between CIRKIL and Ku70. Results: The expression of CIRKIL was increased in I/R myocardium and H2O2-treated cardiomyocytes. Overexpression of CIRKIL increased the expression of γH2A.X, a specific marker of DNA double-strand breaks and aggravated cardiomyocyte apoptosis, whereas knockdown of CIRKIL produced the opposite changes. Transgenic overexpression of CIRKIL aggravated cardiac dysfunction, enlarged infarct area, and worsened cardiomyocyte damage in I/R mice. Knockout of CIRKIL alleviated myocardial I/R injury. Mechanistically, CIRKIL directly bound to Ku70 to subsequently decrease nuclear translocation of Ku70 and impair DNA double-strand breaks repair. Concurrent overexpression of Ku70 mitigated CIRKIL overexpression-induced myocardial I/R injury. Furthermore, knockdown of human CIRKIL significantly suppressed cell damage induced by H2O2 in adult human ventricular cardiomyocytes and human induced pluripotent stem cell-derived cardiomyocytes. Conclusions: CIRKIL is a detrimental factor in I/R injury acting via regulating nuclear translocation of Ku70 and DNA double-strand breaks repair. Thus, CIRKIL might be considered as a novel molecular target for the treatment of cardiac conditions associated with I/R injury.

Human Ku70 is a well-known endogenous nuclear protein involved in the non-homologous end joining pathway to repair double-stranded breaks in DNA. However, Ku70 has been studied in multiple contexts and grown into a multifunctional protein. In addition to the extensive functional study of Ku70 in DNA repair process, many studies have emphasized the role of Ku70 in various other cellular processes, including apoptosis, aging, and HIV replication. In this review, we focus on discussing the role of Ku70 in inducing interferons and proinflammatory cytokines as a cytosolic DNA sensor. We explored the unique structure of Ku70 binding with DNA; illustrated, with evidence, how Ku70, as a nuclear protein, responds to extracellular DNA stimulation; and summarized the mechanisms of the Ku70-involved innate immune response pathway. Finally, we discussed several new strategies to modulate Ku70-mediated innate immune response and highlighted some potential physiological insights based on the role of Ku70 in innate immunity.

The mammalian KU70 is a pleiotropic protein functioning in DNA repair and cytoplasmic suppression of apoptosis. We report a regulatory mechanism by which KU70's cytoplasmic function is enabled due to a methylation at K570 of KU70 by SET-domain-containing protein 4 (SETD4). While SETD4 silencing reduces the level of methylated KU70, over-expression of SETD4 enhances methylation of KU70. Mutations of Y272 and Y284 of SETD4 abrogate methylation of KU70. Although SETD4 is predominantly a nuclear protein, the methylated KU70 is enriched in the cytoplasm. SETD4 knockdown enhances staurosporine (STS)-induced apoptosis and cell killing. Over-expression of the wild-type (WT) SETD4, but not the SETD4-Y272/Y284F mutant, suppresses STS-induced apoptosis. The KU70-K570R (mouse Ku70-K568R) mutation dampens the anti-apoptosis activity of KU70. Our study identifies KU70 as a non-histone substrate of SETD4, discovers a post-translational modification of KU70, and uncovers a role for SETD4 and KU70-K570 methylation in the suppression of apoptosis.