The Mechanism On How RIG-? And MAVS Activate Downstream Signaling Pathway

RIG-I-MAVS-mediated antiviral signaling plays a key role in innate immune defense.Once recongnizing and binding to cytosolic RNA,RIG-I undergoes a conformational change to facilitate its activation.Besides the ligand RNA,what is crucial for RIG-I activation remains a hot spot in the field.There are plenty of reports on RIG-I regulation,which mostly focus on protein-protein interaction and post-translation modification.However,the mechanism on how ubiquitination mediates RIG-I activation is elusive and controversial.In the previous studies,we found that Ube2D3 or Ube2 N works together with Riplet to ubiquitinate RIG-I in human cell lines redundantly.The detailed mechanism underlying Riplet-Ube2D3-and Riplet-Ube2N-mediated RIG-I ubiquitination needs further investigation.In this dissertation,we found that Riplet-Ube2 N is responsible for the activation of RIG-I in mice.Using the in vitro biochemistry assay,we learned that Riplet-Ube2D3 mediates K63-linked ubiquitination of RIG-I,while Riplet-Ube2 N promoted unanchored K63-linked polyubiquitin chain formation.Activated RIG-I interacts with MAVS and mediates its oligomerization.Then activated MAVS recruits downstream signal moleculars TBK1 and IKK,which stimulate the nuclear translocation of IRF3 and NF-?B.These two transcription factors can induce the expression of type-I interferons and other cytokins,which will provide defense against pathogens.MAVS can be divided into three parts(Region I/II/III),and Region I/II is responsible for the activation of IKK/NF-?B,while Region III leads the activation of TBK1/IRF3.But the detailed mechanism on how Region III activating TBK1/IRF3 still remains mysterious and needs further studies.In this dissertation,we utilised the active tetrameric form of MAVS-Region III to set up an in vitro assay,and identified a regulatory protein TRAF3IP3 for MAVS activity.Following the further functional and mechanical analysis,we uncoverd TRAF3IP3 can specifically interact with active MAVS-Region III and TRAF3.The deficiency of TRAF3IP3 crippled IFNB and ISG54 induction,but had no effect on IL-6activation in response to virus infection.Upon RNA virus infection,TRAF3IP3 is accumulated on mitochondria and binds to activated MAVS,which promotes MAVS recruitment of downstream player TRAF3,thus functioning as a positive factor for MAVS-mediated antiviral response.In conclusion,we revealed the mechanisms underlying the ubiquitination and activation of RIG-I and identified TRAF3IP3 as an important regulator for MAVS-mediated TBK1/IRF3 activation.These findings have laid solid foundation for following studies,especially those on autoimmune diseases caused by RIG-I and MAVS mutations.