Cancer metabolism in radiation sensitization – complementary roles of O-GlcNAc transferase and PARP1

For double-strand breaks (DSBs) formed by radiation, the onset of 5' to 3' end resection is a deciding factor in repair pathway choice, favoring homologous recombination (HR) over non-homologous end-joining (NHEJ). Studying HR-proficient MCF7 breast cancer cells, we confirmed a role for PARP1 in promoting DSB repair and limiting resection stress and identified the hexosamine biosynthetic pathway (HBP)-dependent post-translational modification O-GlcNAcylation as an independent regulator. Using pharmacological and genetic perturbations of O-linked β-N-acetylglucosamine (O-GlcNAc) transferase (OGT) and O-GlcNAcase (OGA), we showed that O-GlcNAcylation can limit end resection as measured by BrdU and RPA staining, recruitment of HR proteins BRCA1 and RAD51, and accumulation of cytosolic DNA in S/G2-phase cells. These effects were independent of PARP1 but required the histone methyltransferase EZH2. Loss of OGT or EZH2 phenocopied PARP inhibition, leading to hyper-resection after irradiation. The OGA inhibitor PUGNAc suppressed hyper-resection due to PARP1 knockout whereas treatment with the PARP inhibitor veliparib exacerbated defects in OGT- or EZH2-deficient cells. In each case, increased resection correlated with cytosolic DNA accumulation, suggesting a link to inflammatory signaling. These findings implicate the Warburg effect, via the HBP and O-GlcNAcylation, in favoring NHEJ over HR and suggest that disrupting EZH2 may sensitize HR-proficient tumor cells to radiation via resection-dependent mechanisms. Our results highlight the potential of targeting cancer-associated metabolic reprogramming to overwhelm HR repair and drive resection stress. Combining PARP inhibition with blockade of O-GlcNAcylation or EZH2 might offer a strategy to radiosensitize proliferating HR-proficient cancers while sparing non-cycling normal tissues.