Negative Regulation Of TLR3-triggered Innate Immune Response By E3 Ligase RNF170

Innate immune response serves as the first line of host defense against invading pathogens.However,deficiency or overreaction of innate immune responses will lead to related diseases.So,focusing on the sophisticated regulating mechanism of innate immune responses is extremely important.After the invasion of different pathogens,pattern recognition receptors(PRRs)associate with related ligands,then activate the downstream transcription factors NF-?B,MAPK and IRF3,inducing the production of type I interferons and proinflammatory cytokines.Pattern recognition receptors(PRRs)include RIG-I-like receptors,Toll-like receptors,NOD-like receptors and some other DNA virus receptors.Among all Toll-like receptors,TLR3 can recognize dsRNA,dsRNA is produced by the replication of RNA and DNA viruses,meanwhile,necrotic cells also produce dsRNA.Thus,the regulation and mechanism study of TLR3 signaling is meaningful.Post-translational modifications(PTMs)play a vital role in innate immune responses.It has reported that phosphorylate Tyr759 and Tyr858 of TLR3 is crucial for the recruitment of TRIF to TLR3 and subsequent downstream signaling via NF-?B and IRF3.To find out the new regulators of TLR3-triggered innate immune response,Through MS analysis on TLR3-interacting proteins,we found a E3 ubiquitin ligase,ring finger protein 170(RNF170).By knocking out Rnf170 gene expression in cells and mice,over-expressing of RNF170 in cells,we found that RNF170 negatively regulated TLR3-induced signaling pathway,suppressing the production of type I interferons and proinflammatory cytokines.In vivo experiments,Rnf170-deficient mice exhibited higher inflammatory-death rate compared with wildtype mice after the peritoneal injection of poly(I:C).RNF170 has two functional domains,RING and TM domain,TLR3 has three domains,LRR,TM and TIR domain.We found only full-length RNF170 interacted with TIR domain of TLR3.In our mechanism study,RNF170 as a E3 ubiquitin ligase could increase K48-linked polyubiquitination level of TLR3 and promote TLR3 degradation,inhibiting the downstream-signaling transcriptional activity.There are ten lysine sites of TLR3 TIR domain,by using immunoprecipitation and dual-luciferase reporter assay,we found that RNF170 could deliver the Lys-48-linked ubiquitin to the Lys766 residue of TLR3.In conclusion,we find a new regulator RNF170 of TLR3-triggered innate immune responses which plays an important role.Hence,our study has important values for clinical instruction and translation and provide new potential targets for development of new drugs.