Nucleosome unwrapping and PARP1 allostery drive affinities for chromatin and DNA breaks.

Poly(ADP-ribose) polymerase 1 (PARP1) detects DNA strand breaks that occur in duplex DNA and chromatin. Here, correlative optical tweezers and fluorescence microscopy reveal how single molecules of PARP1 identify single-strand breaks (i.e., nicks), undamaged nucleosome core particles (NCP) and NCPs containing DNA nicks. Fluorescently-tagged PARP1 or PARP2 from nuclear extracts binds nicks with nanomolar affinity but does not engage undamaged dsDNA regions. In contrast, PARP1 avidly binds undamaged NCPs, and partial NCP unwrapping induced by DNA tension significantly increases PARP1 on rate and affinity. Catalytically dead PARP1 or EB-47 inhibition greatly increases PARP1 affinity to DNA nicks and undamaged NCP, implicating a mechanism where PARP1 reverse allostery regulates PARP1 retention to undamaged chromatin. We also monitor ADP-ribosylation in real time upon PARP1 binding undamaged or nicked NCPs. These results provide key mechanistic insights into domain allostery and how pharmacological intervention alters PARP1 binding dynamics for therapeutic impacts.