Identification of CHMP5 as an anti-apoptotic gene by shRNA-based RNAi-screening

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RNA interference (RNAi) is a natural mechanism by which small interfering RNA (siRNA)or short hairpin RNA (shRNA) can mediate degradation of a target RNA molecule in a sequence-specific manner. Here we describe a loss-of-function screen for 288 functionally uncharacterized human genes sensitive for inhibiting apoptosis in HEK293 cells. Using a shRNA-based RNAi library subset, we identified a new anti-apoptotic function for CHMP5 (Charged Multivesicular Body Protein5), which we could validate by caspase inhibitor- and gain of function-assays. In addition, based on NF-kB-luciferase reporter assays we elucidate inhibitory effects of CHMP5 for the NF-kB (Neuclear Factor-kB) pathway. Investigating the effect of CHMP5 gene activity modulation on death receptor TNFR1 (Tumor Necrosis Factor Receptor1) protein turnover, we observed decreased cellular surface expression of TNFR1 in CHMP5 over-expressing cells. Taken together our data reveal new anti-apoptotic function for CHMP5 by modulating the TNFR1 death receptor turnover and thereby affecting the NFkB signalling pathway.