A Novel microRNA of Japanese Flounder Regulates Antimicrobial Immunity Involving a Bacteria-Binding CSF3.

Wenrui Li,Bin Sun,Xiaolu Guan,Li Sun

doi:10.3389/fimmu.2021.723401

Abstract

MicroRNAs (miRNAs) are small non-coding RNAs that regulate diverse biological processes including immunity. In a previous high-throughput RNA sequencing study, a novel miRNA, pol-miR-novel_642, was identified from Japanese flounder (Paralichthys olivaceus), a farmed fish species with important economic value. In this study, we investigated the regulatory mechanism and the function of pol-miR-novel_642 and its target gene. We found that pol-miR-novel_642 targeted, in a sequence-specific manner, a flounder gene encoding an uncharacterized protein that is a structural homologue of murine granulocyte colony stimulating factor 3 (CSF3). The expression of pol-miR-novel_642 and its target gene (named PoCSF3-1) was regulated, in different manners, by the bacterial pathogen Edwardsiella tarda and the viral pathogen megalocytivirus. Overexpression of pol-miR-novel_642 or interference with PoCSF3-1 expression in flounder cells strongly potentiated E. tarda infection. Consistently, in vivo knockdown of PoCSF3-1 enhanced bacterial dissemination in flounder tissues but blocked viral replication, whereas in vivo overexpression of PoCSF3-1 inhibited bacterial dissemination and facilitated viral infection. Overexpression/knockdown of PoCSF3-1 and pol-miR-novel_642 also affected the activation of autophagy. Recombinant PoCSF3-1 (rPoCSF3-1) interacted with and inhibited the growth of Gram-negative bacteria in a manner relying on a PoCSF3-1-characteristic structural motif that is absent in mouse CSF3. rPoCSF3-1 also regulated the proliferation, inflammatory response, and immune defense of flounder head kidney leukocytes in a structure-dependent fashion. Together, these results reveal the function of a novel miRNA-CSF3 regulatory system of flounder, and add new insights into the role and mechanism of fish miRNA and CSF3 in antimicrobial immunity.

Highlights

MicroRNAs are a type of small noncoding RNAs that negatively regulate the expression of protein-coding genes [1,2,3]
When flounder FG-9307 cells were transfected with pol-miR-novel_642 mimic, PoCSF3-1 expression was significantly suppressed at both mRNA and protein levels (Figures 1B, C). These results indicate that PoCSF3-1 is a target gene of pol-miR-novel_642
Accumulating evidences have demonstrated that pathogen infection can induce extensive host miRNA response, which provides a potential mechanism for host antibacterial defense and pathogen manipulation of the host system [54,55,56]

Summary

Introduction

MicroRNAs (miRNAs) are a type of small noncoding RNAs that negatively regulate the expression of protein-coding genes [1,2,3]. MiRNAs most often inhibit their target genes by blocking mRNA translation or decreasing mRNA stability via interaction with the 3’ untranslated regions (3’UTRs) of the target genes [4, 5]. Recent studies showed that some miRNAs interact with the coding regions of the target genes to modulate gene expression [1, 6,7,8]. MiRNAs regulate diverse biological processes and signal pathways involved in infection and immunity [9,10,11,12,13]. MiRNAs associated with bacterial and viral infection have been identified in various species, and some of the miRNAs were found to function by targeting specific immune genes such as IL-1 receptor-associated kinase 4 and signal transducer and activator of transcription (STAT) 3 [19,20,21,22,23]

Methods

Results

Conclusion