PARP1 regulates the genomic ribonucleotide processing activity of TOP1 to prevent the formation of toxic TOP1-DNA adducts and the associated mutations.

Ribonucleotides are frequently incorporated into our genome during replication. Canonically, RNase H2 is responsible for the removal of these embedded ribonucleotides. Alternatively, DNA topoisomerase 1 (TOP1) has also been shown to have genomic ribonucleotide processing activity. When this process occurs at short tandem repeat (STR) sequences, it can lead to 2-5 bp deletions. These deletions are the result of two sequential cuts by TOP1 at sites of ribonucleotide incorporation. In this study, we have determined that PARP1 regulates the TOP1-mediated excision of ribonucleotides by preventing the formation of TOP1-DNA adducts that occur through a second cleavage following the initial ribonucleotide cut by TOP1. We biochemically defined the mechanism by which this regulatory inhibition of TOP1 occurs, which involves both PARP1 physically restricting TOP1 from the cleavage site followed by the inhibitory PARylation of TOP1. We also show that this activity means that PARP1 prevents the TOP1-dependent deletions at STRs in cells. In the absence of both a functional RNase H2 complex and PARP1, we demonstrated that cells appear to be in a senescent state provoked by the accumulation of TOP1-DNA adducts, which are a result of TOP1 being unimpeded to remove genomic ribonucleotides. Our work has elucidated the role of PARP1 in preventing the deleterious consequences of the processing of genomic ribonucleotides by TOP1. Understanding this mechanism could help us develop therapies that better sensitize tumors to PARP inhibitors, especially in cancers that present loss-of-function RNase H2 mutations (seen in certain chronic lymphocytic leukemia and prostate cancers).