Ctenopharyngodon idellus Mex3C activates the innate immune by promoting the K63-linked ubiquitination of RIG-I

Tingting Yu; Chuxin Wu; Dongming Li; Xiaowen Xu; Miaomiao Li; Quanling Zhang; Chen Guo; Yujiao Chen; Chengyu Hu; Zeyin Jiang

doi:10.1016/j.fsi.2025.110519

Ctenopharyngodon idellus Mex3C activates the innate immune by promoting the K63-linked ubiquitination of RIG-I

Fish Shellfish Immunol. 2025 Jun 27:165:110519. doi: 10.1016/j.fsi.2025.110519. Online ahead of print.

Authors

Tingting Yu¹, Chuxin Wu², Dongming Li³, Xiaowen Xu⁴, Miaomiao Li⁴, Quanling Zhang⁴, Chen Guo⁴, Yujiao Chen⁴, Chengyu Hu⁴, Zeyin Jiang⁵

Affiliations

¹ State Key Laboratory of Food Science and Resources, Nanchang University, Nanchang, 330047, Jiangxi, China; Department of Bioscience, College of Life Science, Nanchang University, Nanchang, 330031, Jiangxi, China.
² Yuzhang Normal University, Nanchang, 330103, China.
³ Fuzhou Medical College, Nanchang University, Fuzhou, 344000, Jiangxi, China.
⁴ Department of Bioscience, College of Life Science, Nanchang University, Nanchang, 330031, Jiangxi, China.
⁵ State Key Laboratory of Food Science and Resources, Nanchang University, Nanchang, 330047, Jiangxi, China. Electronic address: Jiangzy20230915@163.com.

PMID: 40582531
DOI: 10.1016/j.fsi.2025.110519

Abstract

The cytoplasmic RNA sensor RIG-I-like receptors (RLRs) play a pivotal role in initiating antiviral innate immunity. In this study, we identified the grass carp (Ctenopharyngodon idellus) Mex3C gene, named CiMex3C which encodes a protein with a RING domain and two heterogeneous nuclear ribonucleoprotein K homology (KH) domains. Our study revealed that CiMex3C responded to multiple stimuli including GCRV, poly(I:C), B-DNA, Z-DNA, characterized by the heightened sensitivity to GCRV stimulation. Notably, CiMex3C co-localized with RIG-I in the cytoplasm of C.idellus kidney (CIK) cells following viral challenge. Further investigation revealed that CiMex3C overexpression significantly enhanced K63-linked ubiquitination of RIG-I. CiMex3C exhibited E3 ubiquitin ligase activity through its conserved RING domain. It directly catalyzed the K63-linked ubiquitin chains on RIG-I. Conversely, siRNA-mediated knockdown of CiMex3C in CIK cells markedly attenuated the RIG-I ubiquitination and suppressed IFN I production. These findings provided the empirical evidences that CiMex3C functions as a post-translational modulator of RIG-I activation, establishing its essential role in antiviral innate immune response in teleosts.

Keywords: IFN I; Innate immunity; Mex3C; RIG-I; Ubiquitination.