Abstract 4979: SIRT1 regulates the function of the KRAB-associated protein 1 (KAP1) in DNA damage response signaling

Abstract

Abstract KRAB-Associated Protein 1 (KAP1) is a master regulator for genome stability, heterochromatin formation, and target gene silencing. Many studies have suggested that post-translational modification is required for KAP1 to execute pleiotropic effects. Here, we have identified the physical interaction between KAP1 and SIRT1, which is a widely recognized NAD+-dependent deacetylase in DNA damage response. Therefore, the objective of this study was to characterize the functional significance of KAP1 deacetylation in DNA damage repair signaling. In this study, SIRT1 was identified as a major interacting partner of KAP1 via biochemical approaches. Inhibition of SIRT1 activity by shRNA or selective inhibitor was able to upregulate the total acetylation level on KAP1, suggesting that KAP1 is a novel target of SIRT1. The major SIRT1-regulated KAP1 deacetylation sites (Lys266, Lys377, Lys469, and Lys770) were detected by quantitative mass spectrometric analysis. Next, to explore the functional significance of KAP1 deacetylation in NHEJ-mediated repair, KAP1 depleted cells reconstituted with wild-type KAP1 or 4KR acetylation mutant were used in the in vivo end-joining reporter system. Compare to wild-type KAP1, the 4KR mutant presented higher activity in promoting NHEJ efficiency. Furthermore, 4KR mutant showed stronger binding affinity to 53BP1, a crucial NHEJ promoting factor, and stabilized the 53BP1 foci formation in response to DNA damage. In summary, this study unambiguously established KAP1 as a target of SIRT1 and that the deacetylation of KAP1 on Lys266, Lys377, Lys469, and Lys770 stabilizes the formation of 53BP1 foci on sites of double-strand breaks, thereby promotes NHEJ-mediated repair pathway. Citation Format: Yi-Hui Lin, Zhenkun Lou. SIRT1 regulates the function of the KRAB-associated protein 1 (KAP1) in DNA damage response signaling. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4979. doi:10.1158/1538-7445.AM2015-4979