DNA damage is known to trigger key cellular defense pathways such as those involved in DNA repair. Here we provide evidence for a previously unrecognized pathway regulating transcription in response to DNA damage and show that this regulation is mediated by the abundant nuclear enzyme poly(ADP-ribose) polymerase. We found that poly(ADP-ribose) polymerase reduced the rate of transcription elongation by RNA polymerase II, suggesting that poly(ADP-ribose) polymerase negatively regulates transcription, possibly through the formation of poly(ADP-ribose) polymerase-RNA complexes. In damaged cells, poly(ADP-ribose) polymerase binds to DNA breaks and automodifies itself in the presence of NAD ؉ , resulting in poly(ADP-ribose) polymerase inactivation. We found that automodification of poly(ADP-ribose) polymerase in response to DNA damage resulted in the up-regulation of transcription, presumably because automodified poly-(ADP-ribose) polymerase molecules were released from transcripts, thereby relieving the block on transcription. Because agents that damage DNA damage RNA as well, up-regulation of RNA synthesis in response to DNA damage may provide cells with a mechanism to compensate for the loss of damaged transcripts and may be critical for cell survival after exposure to DNA-damaging agents. P oly(ADP-ribose) polymerase (PARP) is a highly abundant nuclear protein (for reviews, see refs. 1-6), the physiological role of which is not yet clear. In damaged cells, PARP binds to DNA breaks and becomes enzymatically activated (1, 5). In the presence of its substrate, NAD ϩ , activated PARP automodifies itself through the addition of ADP-ribosyl polymers (1, 5). This automodification inactivates PARP and leads to its dissociation from DNA breaks (7), a prerequisite for DNA repair (8-11). In contrast, in undamaged cells, PARP is found to be associated with regions actively transcribed by RNA polymerase II (POL II) (12), as well as with nucleoli, where ribosomal RNA is transcribed by RNA polymerase I (12). Furthermore, treatment of nuclei with RNase results in the release of PARP (13). In addition, exposure of cells to the transcription inhibitors 5,6-dichloro-1-␤-ribofuranosylbenzimidazole or actinomycin D disperses PARP from nucleoli (14). These observations suggest that PARP may play a role in transcription. Here we demonstrate that PARP mediates a link between DNA damage and elongation of transcription by RNA polymerase II. Materials and MethodsCell-Free Transcription͞DNA Repair Assay. Lymphoblastoid GMO1953C cells (NIGMS Human Mutant Cell Repository, NJ) were cultured in RPMI medium 1640, and whole cell-free extracts were prepared as described by Manley et al. (15). The reactions were carried out with 50 g of cell-free extract, 0.125 g of either pGf1a or p⌬Gf1a plasmid (16), and 0.125 g of ␥-irradiated pBluescript KS (ϩ) plasmid (pBS; Stratagene), prepared as described previously (10). The transcription-DNA repair assay reaction conditions have been described previously (16). Briefly, the reaction was carried out in a mix...