Ablating Ku70 phosphorylation results in defective DNA damage repair and spontaneous induction of hepatocellular carcinoma

Abstract

Multiple pathways mediate the repair of DNA double-strand break (DSB), with numerous mechanisms responsible for driving choice between the pathways. Previously, we reported that phosphorylation of the non-homologous end joining (NHEJ) factor, Ku70, is required for the dissociation of the Ku heterodimer from DNA ends to allow DSB repair via homologous recombination (HR). A knock-in mouse, in which phosphorylation is ablated in the three conserved sites of Ku70 (Ku703A/3A), was generated in order to test the hypothesis that Ku70 phosphorylation is required for initiation of HR and that blocking this process results in enhanced genomic instability and tumorigenesis. Here, we show that Ku703A/3A mice develop spontaneous and have accelerated chemical-induced hepatocellular carcinoma (HCC) compared to wild-type (Ku70+/+) littermates. The HCC tumors from the Ku703A/3A mice have increased {gamma}H2AX and 8-oxo-G staining, suggesting DNA repair is decreased in these mice. Spontaneous transformed cell lines from Ku703A/3A mice are more radiosensitive, have a significant decrease in DNA end resection, and are more sensitive to the DNA cross-linking agent mitomycin C compared to cells from Ku70+/+ littermates. Collectively, these findings demonstrate that phosphorylation-mediated dissociation of Ku heterodimer from DNA ends is required for efficient DNA damage repair and disruption of this process results in genomic instability and accelerated development of HCC.