PARG inhibition elicits cytotoxicity by sequestering XRCC1 and other PAR-binding proteins into PAR-dependent nuclear condensates

DNA breaks activate PARP1/2 to synthesize poly(ADP-ribose) (PAR), which relax chromatin and recruit DNA repair factors. PAR is normally short-lived and rapidly degraded by poly(ADP-ribose) glycohydrolase (PARG). While PARP1/2 inhibitors target homologous recombination (HR)-deficient cancers, predictive biomarkers for PARG inhibition (PARGi) remain undefined. Using genome-wide CRISPR screens, we find that PARGi is synthetically lethal with loss of multiple PAR-binding factors (XRCC1-LIG3/POLB, ALC1/CHD1L, ARH3, PARG itself) but not HR-deficiency. Conversely, loss of PARP1, NMNAT1 (nuclear NAD⁺ synthesis), or UNG confers resistance. Mechanistically, PARGi induces reversible PARP1- and PAR-dependent nuclear condensates in undamaged cells. While lacking detectable DNA breaks, these condensates sequester various PAR-binding proteins (e.g. XRCC1, ALC1, FUS), reducing their nuclear availability and impairing recruitment to bona fide DNA lesions. These findings identify PAR-binding protein sequestration as the mechanism underlying PARGi cytotoxicity, explain the paradoxical essentiality of PARG but not PARP1, and establish PAR-binding factor deficiency as a biomarker for PARGi. While PARP inhibitors are well characterized, predictive biomarkers for PARG inhibition are undefined. Here, the authors propose a unified mechanism in which the sequestration of PAR-binding proteins underlies PARG inhibitor-induced cytotoxicity.