Genome-wide siRNA screen for mediators of NF-?B activation.

ABSTRACT: Although canonical NF?B is frequently critical for cell proliferation, survival, or differentiation, NF?B hyperactivation can cause malignant, inflammatory, or autoimmune disorders. Despite intensive study, mammalian NF?B pathway loss-of-function RNAi analyses have been limited to specific protein classes. We therefore undertook a human genome-wide siRNA screen for novel NF?B activation pathway components. Using an Epstein Barr virus latent membrane protein (LMP1) mutant, the transcriptional effects of which are canonical NF?B-dependent, we identified 155 proteins significantly and substantially important for NF?B activation in HEK293 cells. These proteins included many kinases, phosphatases, ubiquitin ligases, and deubiquinating enzymes not previously known to be important for NF?B activation. Relevance to other canonical NF?B pathways was extended by finding that 118 of the 155 LMP1 NF-?B activation pathway components were similarly important for IL-1?-, and 79 for TNF?-mediated NF?B activation in the same cells. MAP3K8, PIM3, and six other enzymes were uniquely relevant to LMP1-mediated NF?B activation. Most novel pathway components functioned upstream of I?B kinase complex (IKK) activation. Robust siRNA knockdown effects were confirmed for all mRNAs or proteins tested. Although multiple ZC3H-family proteins negatively regulate NF?B, ZC3H13 and ZC3H18 were activation pathway components. ZC3H13 was critical for LMP1, TNF?, and IL-1? NF?B-dependent transcription, but not for IKK activation, whereas ZC3H18 was critical for IKK activation. Down-modulators of LMP1 mediated NF?B activation were also identified. These experiments identify multiple targets to inhibit or stimulate LMP1-, IL-1?-, or TNF?-mediated canonical NF?B activation.

SUBMITTER: Gewurz BE 

PROVIDER: S-EPMC3289371 | biostudies-literature | 2012 Feb

REPOSITORIES: biostudies-literature