von-Hippel Lindau protein (pVHL) suppresses tumorigenesis in the kidney, in part through regulation of hypoxia-inducible factor alpha (HIF alpha). However, HIF has been proposed to be necessary but insufficient for renal tumorigenesis. p53 was implicated as a transcription factor that is regulated by pVHL, but the molecular mechanism by which pVHL regulates p53 on DNA damage is unknown. We demonstrated that checkpoint kinase-2 (Chk2) binds to the beta-domain of pVHL and phosphorylates Ser 111 on DNA damage. Notably, this modification enhances pVHL-mediated transactivation of p53 by recruiting p300 and Tip60 to the chromatin of p53 target gene. Further, the naturally occurring pVHL mutants pVHL-S111R and pVHL-S111C showed diminished binding to coactivators, ultimately retarding p53-mediated growth arrest and apoptosis. In this study, we determined the molecular mechanism by which pVHL transactivates p53 on DNA damage and demonstrated that p53-related pVHL subtype mutants regulate tumorigenecity in VHL diseases.