Site-specific acetylation of polynucleotide kinase 3'-phosphatase regulates its distinct role in DNA repair pathways

Mammalian polynucleotide kinase 3 ' -phosphatase (PNKP), a DNA end-processing enzyme with 3 ' -phosphatase and 5 ' -kinase activities, is in- v olv ed in multiple DNA repair pathw a y s, including base e x cision (BER), single-strand break (SSBR), and double-strand break repair (DSBR). Ho w e v er, little is kno wn as to ho w PNKP functions in such diverse repair processes. Here we report that PNKP is acetylated at K1 42 (AcK1 42) by p300 constitutively but at K226 (AcK226) by CBP, only after DSB induction. Co-immunoprecipitation analysis using AcK142 or AcK226 PNKP- specific antibodies sho w ed that A cK142-PNKP associates only with BER / SSBR, and AcK226 PNKP with DSBR proteins. Despite the modest effect of acetylation on PNKP's enzymatic activity in vitro , cells expressing non-acetylable PNKP (K142R or K226R) accumulated DNA damage in transcribed genes. Intriguingly, in striatal neuronal cells of a Huntington's Disease (HD)-based mouse model, K142, but not K226, was acetylated. This is consistent with the reported degradation of CBP, but not p300, in HD cells. Moreo v er, transcribed genomes of HD cells progressively accumulated DSBs. Chromatin-immunoprecipitation analysis demonstrated the association of Ac-PNKP with the transcribed genes, consistent with PNKP's role in transcription-coupled repair. Thus, our findings demonstrate that acetylation at two lysine residues, located in different domains of PNKP, regulates its distinct role in BER / SSBR versus DSBR.