CRFB5a, a Subtype of Japanese Eel (Anguilla japonica) Type I IFN Receptor, Regulates Host Antiviral and Antimicrobial Functions through Activation of IRF3/IRF7 and LEAP2

Tianyu Wang; Peng Lin; Yilei Wang; Xiaojian Lai; Pengyun Chen; Fuyan Li; Jianjun Feng

doi:10.3390/ani13193157

CRFB5a, a Subtype of Japanese Eel (Anguilla japonica) Type I IFN Receptor, Regulates Host Antiviral and Antimicrobial Functions through Activation of IRF3/IRF7 and LEAP2

Animals (Basel). 2023 Oct 9;13(19):3157. doi: 10.3390/ani13193157.

Authors

Tianyu Wang¹, Peng Lin¹, Yilei Wang¹, Xiaojian Lai¹, Pengyun Chen¹, Fuyan Li¹, Jianjun Feng^{1

2}

Affiliations

¹ Key Laboratory of Healthy Mariculture for the East China Sea, Ministry of Agriculture and Rural Affairs, Fisheries College, Jimei University, Xiamen 361021, China.
² The Open Program of Key Laboratory of Cultivation and High-Value Utilization of Marine Organisms in Fujian Province, Xiamen 361000, China.

Abstract

IFNAR1, one of the type I IFN receptors, is crucial to mammalian host defense against viral invasion. However, largely unknown is the immunological role of the fish teleost protein IFNAR1, also known as CRFB5. We have successfully cloned the whole cDNA of the Japanese eel's (Anguilla japonica) CRFB5a homolog, AjCRFB5a. The two fibronectin-3 domains and the transmembrane region (238-260 aa) of AjCRFB5a are normally present, and it shares a three-dimensional structure with zebrafish, Asian arowana, and humans. According to expression analyses, AjCRFB5a is highly expressed in all tissues found, particularly the liver and intestine. In vivo, Aeromonas hydrophila, LPS, and the viral mimic poly I:C all dramatically increased AjCRFB5a expression in the liver. Japanese eel liver cells were reported to express AjCRFB5a more strongly in vitro after being exposed to Aeromonas hydrophila or being stimulated with poly I: C. The membranes of Japanese eel liver cells contained EGFP-AjCRFB5a proteins, some of which were condensed, according to the results of fluorescence microscopy. Luciferase reporter assays showed that AjCRFB5a overexpression strongly increased the expression of immune-related genes in Japanese eel liver cells, such as IFN1, IFN2, IFN3, IFN4, IRF3, IRF5, and IRF7 of the type I IFN signaling pathway, as well as one of the essential antimicrobial peptides LEAP2, in addition to significantly inducing human IFN-promoter activities in HEK293 cells. Additionally, RNA interference (RNAi) data demonstrated that knocking down AjCRFB5a caused all eight of those genes to drastically lower their expression in Japanese eel liver cells, as well as to variable degrees in the kidney, spleen, liver, and intestine. Our findings together showed that AjCRFB5a participates in the host immune response to bacterial infection by inducing antimicrobial peptides mediated by LEAP2 and favorably modulates host antiviral immune responses by activating IRF3 and IRF7-driven type I IFN signaling pathways.

Keywords: Anguilla japonica; CRFB5a; LEAP2; RNAi; type I IFN.

Grants and funding

This work was financially supported by the “Nature Science Foundation of Fujian Province” (No. 2020J01671, 2021J01830), “the Open Program of Key Laboratory of Cultivation and High-value Utilization of Marine Organisms in Fujian Province” (No. 2021fjscq03), “Engineering Research Center of the Modern Technology for Eel Industry, Ministry of Education, China” (No. RE202307, RE202208), and “Key Laboratory and Healthy Mariculture for the East China Sea, Ministry of Agriculture and Rural Affairs, China” (No. 2020ESHML02).