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

Abstract

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, ΔNp63α, 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 ΔNp63α. Anticancer drugs, including doxorubicin, induced ΔNp63α ISGylation and caspase-2 activation, leading to cleavage of ISGylated ΔNp63α in the nucleus and subsequent release of its inhibitory domain to the cytoplasm. ISGylation ablated the ability of ΔNp63α to promote anchorageindependent 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 ΔNp63α and provide a molecular rationale for therapeutic use of doxorubicin against ΔNp63α-mediated cancers.

Caspase-2 at a glance

“…In addition to the proapoptotic protein BID, only a few other caspase-2 substrates with a link to cell death regulation have been identified so far (see supplementary material Table S1 and poster), including the p53-related transcription factor p63 (Jeon et al, 2012). In response to DNA damage, DNp63, a splice variant that inhibits the proapoptotic transcriptional activity of full length p63 (the TAp63 isoform) by dimerization in a dominant-negative manner, is modified by the ubiquitin-like protein insulin-stimulated gene 15 (ISG15), which is required for its subsequent cleavage and inactivation by caspase-2.…”

“…In response to DNA damage, DNp63, a splice variant that inhibits the proapoptotic transcriptional activity of full length p63 (the TAp63 isoform) by dimerization in a dominant-negative manner, is modified by the ubiquitin-like protein insulin-stimulated gene 15 (ISG15), which is required for its subsequent cleavage and inactivation by caspase-2. This cleavage leads to the nuclear export of the inhibitory fragment, allowing TAp63 to induce transcription of relevant target genes like proapoptotic PUMA and NOXA (also known as BBC3 and PMAIP1, respectively) (Jeon et al, 2012). In a xenograft tumor model, no difference in growth was observed between cells expressing either wild-type DNp63, a version of TAp63 resistant to caspase-2 cleavage, or a version of TAp63 that is defective for ISG15 conjugation.…”

“…In a xenograft tumor model, no difference in growth was observed between cells expressing either wild-type DNp63, a version of TAp63 resistant to caspase-2 cleavage, or a version of TAp63 that is defective for ISG15 conjugation. Upon DNA damage, however, wild-type cells are more sensitive (Jeon et al, 2012), suggesting that TAp63 has an important role in the proapoptotic response to DNA damage, and that this requires caspase-2 (see poster).…”

ISG15 regulates IFN-γ immunity in human mycobacterial disease

“…Significant efforts have been made to identify the substrates of ISG15 conjugation and their roles in pathogen infection and tumorigenesis [3,4]. For example, the molecular basis of antiviral effect of ISG15 was described in a recent report of the influenza A viral protein NS1 (NS1A).…”

“…ISG15 modification on NS1A resulted in a disruption of normal function of NS1A, thus inhibiting the virus infection [3]. Meanwhile, a recent report revealed an important role of ISG15 in the enhancement of chemosensitivity during cancer therapy [4]. Treatment of cells with several chemotherapeutic drugs, including doxorubicin and camptothecin, increases ISG15 expression and results in ISGylation of ΔNp63α, an alternative splice variant of p53 family protein p63.…”