Optineurin Is Required for CYLD-Dependent Inhibition of TNFα-Induced NF-κB Activation

The nuclear factor kappa B (NF-kappa B) regulates genes that function in diverse cellular processes like inflammation, immunity and cell survival. The activation of NF-kappa B is tightly controlled and the deubiquitinase CYLD has emerged as a key negative regulator of NF-kappa B signalling. Optineurin, mutated in certain glaucomas and amyotrophic lateral sclerosis, is also a negative regulator of NF-kappa B activation. It competes with NEMO (NF-kappa B essential modulator) for binding to ubiquitinated RIP (receptor interacting protein) to prevent NF-kappa B activation. Recently we identified CYLD as optineurin-interacting protein. Here we have analysed the functional significance of interaction of optineurin with CYLD. Our results show that a glaucoma-associated mutant of optineurin, H486R, is altered in its interaction with CYLD. Unlike wild-type optineurin, the H486R mutant did not inhibit tumour necrosis factor alpha (TNF alpha)-induced NF-kappa B activation. CYLD mediated inhibition of TNF alpha-induced NF-kappa B activation was abrogated by expression of the H486R mutant. Upon knockdown of optineurin, CYLD was unable to inhibit TNF alpha-induced NF-kappa B activation and showed drastically reduced interaction with ubiquitinated RIP. The level of ubiquitinated RIP was increased in optineurin knockdown cells. Deubiquitination of RIP by over-expressed CYLD was abrogated in optineurin knockdown cells. These results suggest that optineurin regulates NF-kappa B activation by mediating interaction of CYLD with ubiquitinated RIP thus facilitating deubiquitination of RIP.