STAT3 Dysregulation of XRCC1 Results on Altered Base Excision Repair

Base Excision Repair (BER) is a critical DNA repair pathway, repairing base damage, the spontaneous decay of bases, abasic sites, and single-strand DNA breaks. BER requires the sequential action of multiple proteins, including poly(ADP ribose) polymerase 1 (PARP1), X-ray cross complementing group 1 (XRCC1), and DNA polymerase beta (POL ). Expression changes and polymorphisms in BER proteins alter the response of cancer cells to multiple DNA damaging chemotherapeutics, with overexpression of XRCC1 increasing chemoresistance in gastric, gallbladder, and ovarian cancer. However, the transcriptional regulation of XRCC1 remains largely understudied. Here, we identify the signal transducer and activator of transcription 3 (STAT3) as a novel regulator of XRCC1 in TNBC cell line models, the osteosarcoma cell line U2OS, and the human embryonic kidney cell line HEK293T. Glucose, epidermal growth factor (EGF) and interleukin 6 (IL-6) are all known to activate STAT3 and promote transcription We have found that this constitutive activation results in persistent occupancy of STAT3 at the XRCC1 promoter in TNBC cell lines, inducing overexpression of XRCC1 protein. However, the non-tumorigenic HEK293T cell line and osteosarcoma U2OS cell line do not have constitutively activated STAT3 but demonstrate an inducible response to the activation of STAT3 from exogenous stimuli. Using acute exposures to EGF, IL-6, and glucose, we observed increased activated STAT3 and subsequent increases in XRCC1 expression in HEK293T and xvii U2OS cells. Critically, the increased XRCC1 expression induced by high glucose exposure resulted in increased DNA repair, measured through alkaline comet assay, and increased survival following methyl methanesulfonate (MMS) challenge in both HEK293T and U2OS cells. Furthermore, continued exposure to elevated glucose concentrations resulted in persistent STAT3 occupancy at the XRCC1 promoter and elevated XRCC1 expression in both HEK293T and U2OS cells, similar to what is seen in constitutively active TNBC cell lines. Low glucose adaption reversed these effects by reducing STAT3 activation and occupancy at the XRCC1 promoter in the HEK293T U2OS cells. However, low glucose adaptation in the MDA-MB-231, which have constitutively active STAT3, only slightly reduced STAT3 activation. Upstream regulators of STAT3 revealed a correlation between IL-6R expression and the inducibility of STAT3 activation and subsequent XRCC1 expression. High EGFR expression and low IL-6R expression resulted in minimal inducibility of MDA-231. In addition to these findings, we also identified POLB as a target for STAT3 regulation and mapped the STAT3 binding site within the POLB promoter. These results demonstrate that activation of STAT3 regulates XRCC1 expression and altered BER functions across various cell line models. More importantly, it links constitutive activation of STAT3 with dysregulation of XRCC1 and BER in TNBC cells and undercovers a critical mechanism by which XRCC1 can become dysregulated in cancer and promote chemoresistance