The accumulation of damage caused by oxidative stress has been linked to aging and to the etiology of numerous age-related diseases. The longevity gene, sirtuin 6 (SIRT6), promotes genome stability by facilitating DNA repair, especially under oxidative stress conditions. Here we uncover the mechanism by which SIRT6 is activated by oxidative stress to promote DNA double-strand break (DSB) repair. We show that the stress-activated protein kinase, c-Jun N-terminal kinase (JNK), phosphorylates SIRT6 on serine 10 in response to oxidative stress. This post-translational modification facilitates the mobilization of SIRT6 to DNA damage sites and is required for efficient recruitment of poly (ADP-ribose) polymerase 1 (PARP1) to DNA break sites and for efficient repair of DSBs. Our results demonstrate a post-translational mechanism regulating SIRT6, and they provide the link between oxidative stress signaling and DNA repair pathways that may be critical for hormetic response and longevity assurance. Display omitted •JNK phosphorylates SIRT6 at residue S10 in response to oxidative stress•SIRT6 S10 phosphorylation is required for the stimulation of DNA break repair•SIRT6 S10 phosphorylation stimulates SIRT6 mono-ADP ribosylation activity on PARP1 Van Meter et al. show that SIRT6 is phosphorylated by JNK on serine 10 in response to oxidative stress. SIRT6 S10 phosphorylation promotes DNA break repair by stimulating SIRT6 and PARP1 recruitment to DNA break sites.