Ubiquitination profiling identifies sensitivity factors for IAP antagonist treatment

Evasion of cell death is one crucial capability acquired by tumor cells to ward off anti-tumor therapies, and represents a fundamental challenge to sustaining clinical efficacy for currently available agents. Inhibitor of Apoptosis (IAP) proteins use their ubiquitin E3 ligase activity to promote cancer cell survival by mediating proliferative signaling and blocking cell death in response to diverse stimuli. Using immunoaffinity enrichment and mass spectrometry, ubiquitination sites on thousands of proteins were profiled upon initiation of cell death by IAP antagonists in IAP antagonist-sensitive and -resistant breast cancer cell lines. Our analyses identified hundreds of proteins with elevated levels of ubiquitin-remnant (K-GG) peptides upon activation of cell death by the IAP antagonist BV6. The majority of these were observed in BV6-sensitive, but not -resistant cells. Among these were known proapoptotic regulators including cytochrome C, RIP1 and a selection of proteins known to reside in the mitochondria or regulate NF-kB signaling. Analysis of early timepoints revealed that IAP antagonist treatment stimulated rapid ubiquitination of NF-kB signaling proteins including TRAF2, HOIL1, NEMO as well as c-IAP1 autoubiquitination. Knockdown of several NF-kB pathway members reduced BV6 induced cell death and TNF production in sensitive cell lines. Importantly, RIP1 was found to be constitutively ubiquitinated in sensitive breast cancer cell lines at higher basal level than in res...

Source: BJ Cell - November 21, 2014 Category: Biochemistry Authors: E Varfolomeev, A Izrael-Tomasevic, K Yu, D Bustos, T Goncharov, L D Belmont, A Masselot, C E Bakalarski, D S Kirkpatrick, D Vucic Tags: BJ Signal Source Type: research