BRCA1 modulates the autophosphorylation status of DNA-PKcs in S phase of the cell cycle.

Anthony J Davis,Jun Yang,Kyung-Jong Lee,Linfeng Chi,Eiichiro Mori,David J Chen,Sairei So,Kazi Fattah

doi:10.1093/nar/gku824

Abstract

Non-homologous end-joining (NHEJ) and homologous recombination (HR) are the two prominent pathways responsible for the repair of DNA double-strand breaks (DSBs). NHEJ is not restricted to a cell-cycle stage, whereas HR is active primarily in the S/G2 phases suggesting there are cell cycle-specific mechanisms that play a role in the choice between NHEJ and HR. Here we show NHEJ is attenuated in S phase via modulation of the autophosphorylation status of the NHEJ factor DNA-PKcs at serine 2056 by the pro-HR factor BRCA1. BRCA1 interacts with DNA-PKcs in a cell cycle-regulated manner and this interaction is mediated by the tandem BRCT domain of BRCA1, but surprisingly in a phospho-independent manner. BRCA1 attenuates DNA-PKcs autophosphorylation via directly blocking the ability of DNA-PKcs to autophosphorylate. Subsequently, blocking autophosphorylation of DNA-PKcs at the serine 2056 phosphorylation cluster promotes HR-required DNA end processing and loading of HR factors to DSBs and is a possible mechanism by which BRCA1 promotes HR.

Highlights

DNA double-strand breaks (DSBs) are the most deleterious type of DNA lesion because if unrepaired or misrepaired they can promote chromosomal aberrations resulting in genomic instability, which is a driving force for tumorigenesis [1]
DNA-PKcs localizes to DSBs but its autophosphorylation at serine 2056 is attenuated in S phase of the cell cycle
To expand on this previous result, we examined if recruitment of other canonical non-homologous end-joining (NHEJ) factors to DSBs is attenuated in S phase

Summary

Introduction

DNA double-strand breaks (DSBs) are the most deleterious type of DNA lesion because if unrepaired or misrepaired they can promote chromosomal aberrations resulting in genomic instability, which is a driving force for tumorigenesis [1]. Recruitment of DNA-PKcs to the DSB mediates the formation of the DNA-PK complex (Ku70/80 with DNAPKcs) and results in activation of its catalytic activity, which is required for NHEJ. DNA-PKcs is phosphorylated and autophosphorylated at a number of sites in response to DNA damage with the best characterized being the serine 2056 and threonine 2609 phosphorylation clusters [7]. Phosphorylation of these two clusters is important for NHEJ as ablation of either phosphorylation cluster causes increased radiosensitivity and less efficient DSB repair [8,9,10,11,12]. If required, the two DNA termini are processed and ligated by the XRCC4-DNA Ligase IV complex

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