Roles of p97/VCP in linear ubiquitin mediated NF-kappaB activation

LUBAC泛素连接酶由HOIP、HOIL-1L和Sharpin三个亚基组成。LUBAC具有多个结构域，但存在于HOIP N-末端区域的pub结构域的作用尚未解决。PUB结构域已被确定为p97/VCP的结合域。由于p97/VCP已被证明负向调节NF-kappaB的激活，我们研究了pub结构域在LUBAC介导的NF-kappaB激活中的作用。令我们惊讶的是，在表达HOIP突变体的细胞中，不能结合p97/VCP的细胞中，核因子-kappaB的激活和肿瘤坏死因子-α介导的NEMO的线性多泛素化都得到了增强。然后我们寻找与HOIP的pub结构域结合的蛋白质，并发现两种脱泛素酶OTULIN和CyLD，它们特异性地切割线性链作为HOIP pub的相互作用蛋白。OTULIN和CyLD都直接与HOIP的pub结构域结合，并下调LUBAC介导的NF-kappaB的激活。

The LUBAC ubiquiin ligase is composed of three subunits, HOIP, HOIL-1L and SHARPIN. LUBAC possesses multiple domains, but the role of the PUB domain that exists in the N-terminal region of HOIP had not been addressed. The PUB domain has been identified as a binding domain of p97/VCP. Since p97/VCP has shown to regulate NF-kappaB activation negatively, we examined the role of the PUB domain in LUBAC mediated NF-kappaB activation. To our surprise, in cells expressing a HOIP mutant, which cannot bind p97/VCP, both NF-kappaB activation and linear polyubiquitination of NEMO mediated by TNF-alpha were augmented. We then sought proteins bound to the PUB domain of HOIP and found that two deubiquitinases, OTULIN and CYLD, that specifically cleaved linear chains as interacting proteins of HOIP PUB. Both OTULIN and CYLD bind to the PUB domain of HOIP directly and down-regulated LUBAC-mediated NF-kappaB activation.