Chemosensitivity is controlled by p63 modification with ubiquitin-like protein ISG15

Identification of the cellular mechanisms that mediate cancer cell chemosensitivity is important for developing new cancer treatment strategies. Several chemotherapeutic drugs increase levels of the posttranslational modifier ISG15, which suggests that ISGylation could suppress oncogenesis. However, how ISGylation of specific target proteins controls tumorigenesis is unknown. Here, we identified proteins that are ISGylated in response to chemotherapy. Treatment of a human mammary epithelial cell line with doxorubicin resulted in ISGylation of the p53 family protein p63. An alternative splice variant of p63, Delta Np63 alpha, suppressed the transactivity of other p53 family members, and its expression was abnormally elevated in various human epithelial tumors, suggestive of an oncogenic role for this variant. We showed that ISGylation played an essential role in the downregulation of Delta Np63 alpha. Anticancer drugs, including doxorubicin, induced Delta Np63 alpha ISGylation and caspase-2 activation, leading to cleavage of ISGylated Delta Np63 alpha in the nucleus and subsequent release of its inhibitory domain to the cytoplasm. ISGylation ablated the ability of Delta Np63 alpha to promote anchorage-independent cell growth and tumor formation in vivo as well to suppress the transactivities of proapoptotic p53 family members. These findings establish ISG15 as a tumor suppressor via its conjugation to Delta Np63 alpha and provide a molecular rationale for therapeutic use of doxorubicin against Delta Np63 alpha-mediated cancers.