The Recessive Breast Cancer Predisposition Variant In XRCC4 Abrogates DNA Damage Response Via Disturbing Its Nuclear Localization

The incidence of breast cancer is generally attributed to both genetic and environmental factors. Genetic variants in a network of genes involved in DNA damage repair (DDR) are known to predispose to breast cancer. XRCC4 plays a crucial role in the non-homologous end-joining pathway (NHEJ) to maintain genome stability, making it as a potential candidate for yet unexplained breast cancer susceptibility.Association study was conducted through a two-stage case-control study in a Chinese female population. We examined the variant’s effect on XRCC4 nuclear localization in both cellular and tissue level. We silenced the core XRCC4 complex to reveal which partner facilitates heterozygote’s nucleus localization. Finally, we used comet assays, colony formation assay and metaphase spreads assay to assess the variant’s impact on DDR, All statistical tests were two-sided.A recessive missense variant, rs3734091 (c.739G>T, p.Ala247Ser) in XRCC4, was significantly associated with an increased risk of breast cancer (odds ratio [OR]= 3.92, P=0.007). This association was more significant in patients with triple-negative breast cancer (TNBC) (OR=18.65, P<0.0001), but not luminal or HER-2+breast cancer. This p.Ala247Ser alternation partially disturbed the nuclear localization of XRCC4 exclusively in homozygote harboring rs3734091-TT genotype but not in heterozygote. Similar aberrant distributions were also observed at the tissue level in breast tumors. Furthermore, we revealed that wild-type XRCC4 could facilitate the nucleus localization of p.Ala247Ser mutant in heterozygote. Remarkably, functional analysis demonstrated that p.Ala247Ser significantly reduced the in vivo NHEJ repair efficiency compared to the wild-type XRCC4 via colony formation assay and NHEJ reporter assay. Metaphase-spread assay shows that XRCC4A247S leads to chromosomal abnormalities, suggesting that reduced nuclear accumulation by XRCC4A247S variant impaired its functions in response to DNA damage and ultimately perturbed genomic stability.Our findings provide the biological rationality why XRCC4 rs3734091 confers an increased susceptibility to breast cancer exclusively under a recessive model and highlight the importance of genetic pathogenesis in mammary carcinogenesis.