USP14 negatively regulates RIG-I-mediated IL-6 and TNF-α production by inhibiting NF-κB activation

•USP14 knockdown increased RIG-I-mediated pro-inflammatory cytokines production.•USP14 negatively regulates VSV-induced activation of NF-κB signaling.•USP14 deubiquitinates K63-linked RIG-I.•IU1 promotes pro-inflammatory cytokines production in VSV-infected mice in vivo. Ubiquitin specific protease 14 (USP14) is a regulator of protein deubiquitination and proteasome activation, and has been implicated in negative regulation of type I IFN signaling pathway. However, the effect of USP14 on RNA virus-related inflammatory response has not been studied. Retinoic acid-inducible gene I (RIG-I) is the important pattern recognition receptor of the innate immunity to detect RNA viruses or intracellular Poly(I:C)-LMW. Here, we reported that USP14 knockdown increased pro-inflammatory cytokines production in macrophages upon VSV infection or intracellular Poly(I:C)-LMW stimulation. USP14-overexpressed HeLa cells exhibited a decrease in RIG-I-mediated IL-6 and TNF-α expression. IU1, USP14 inhibitor, significantly promotes pro-inflammatory cytokines production in VSV-infected mice in vivo. Furthermore, USP14 was also found to inhibit the RIG-I-triggered NF-κB activation by deubiquitinating K63-linked RIG-I. Thus, our results demonstrate that USP14 is a negative regulator of RIG-I-mediated inflammatory response.