USP2Negatively Regulates Nf-κB Activation By Deubiquitinating TRAF6

Vertebrates are constantly threatened by the invasion of microorganisms and have evolved systems of immune defense to eliminate infective pathogens in the body. The mammalian immune system is comprised of two branches:innate and acquired immunity. The innate immune system is the first line of host defense against pathogens and is mediated by phagocytes including macrophages and dendritic cells (DCs). Acquired immunity is involved in elimination of pathogens in the late phase of infection as well as the generation of immunological memory. The innate immune system recognizes microorganisms'pathogen-associated molecular pattern (PAMP) via a limited number of germline-encoded pattern-recognition receptors (PRRs). PRRs are inluding Toll-like receptor (TLR), RIG-I-like receptor (RLR), NOD-like receptor (NLR) and DNA sensor etc. When the ligands bind to PRRs, they can recruit adaptor proteins to transduct immune signaling and activat immune responds. They are involved in different pathways and therefore a number of transcriptional factors can be activatied including IRF3, IRF7, API and NF-κB which can promote the transcription of pro-inflammatory cytokines and type I interferons and potently activate immune responds.The transcription factor, nuclear factor-kappa B (NF-κB), plays critical roles in many biological processes especially immunity. The signaling to NF-κB activation is subtly regulated to avoid harmful effects. In this report, we identified USP2a (ubiquitin-specific protease2isoform a) as a novel negative regulator in TLR/IL-1β and SeV-induced NF-κB activation. Overexpression of USP2a inhibited IL-1β and SeV-induced NF-κB activation and the transcription of inflammatory cytokines. We also generated HCT116USP2gene knock out cells and performed several experiments in this cells. We found that knockout of USP2a can promote the phosphorylation of TAK1/IKK complex, phosphorylation and degradation of IκBα, NF-κB activation as well as cytokine production. We also discovered that USP2was persistently associated with TRAF6(tumor necrosis factor receptor-associated factor6), and this association was no affectd by the stimulus such as IL-1β. USP2a is a deubiquiting enzyme. We proved that USP2a removed IL-1β or SeV-induced K63-linked polyubiquitin chains of TRAF6and knock out of USP2a have the opposite effect. Furthermore, the residues of USP2responsible for its role were also clarified. These results provide a relative universal mechanism in the negative regulation of NF-κB activation.Ubiquitination is an important modification in immune responds, and deubiquitination is a key regulation mechanism in immune pathways. There are over100DUBs discovered by researchers, but the functions of most DUBs are still unknown to us. It is quite necessary to have more studies on DUBs, including USP2a which functions are still no fully understood. Comparing with other DUBs we known the proteins belonging to this family have many interaction targets, but for USP2a, we only discovered TRAF6. USP2a is an isoform of USP2protein. There are other isoforms of USP2, and their function is less studied. Besides, the detailed TRAF6deubiquitination mechanism through USP2a is not fully understood. All of aboye questions are needed to further study.