Multiomics analysis revealed miRNAs as potential regulators of the immune response in Carassius auratus gills to Aeromonas hydrophila infection

Abstract

The gill of fish is an important immune organ for pathogen defense, but its microRNA (miRNA) expression and regulatory mechanism remain unclear. In this study, we report on the histopathological and immunohistochemical features of the gills of the crucian carp Carassius auratus challenged with Aeromonas hydrophila. Small RNA libraries of the gills were constructed and sequenced on the Illumina HiSeq 2000 platform. A total of 1,165 differentially expressed miRNAs (DEMs) were identified in gills, of which 539 known and 7 unknown DEMs were significantly screened (p < 0.05). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses revealed that the potential target genes/proteins were primarily involved in 33 immune-related pathways, in which the inflammatory responses were focused on the Toll-like receptor (TLR), mitogen-activated protein kinase (MAPK), and nuclear factor kappa B (NF-κB) signaling pathways. Moreover, the expression levels of 14 key miRNAs (e.g., miR-10, miR-17, miR-26a, miR-144, miR-145, and miR-146a) and their target genes (e.g., TNFα, TLR4, NF-κB, TAB1, PI3K, and IRAK1) were verified. In addition, the protein levels based on isobaric tags for relative and absolute quantification (iTRAQ) were significantly associated with the results of the quantitative real-time PCR (qRT-PCR) analysis (p < 0.01). miR-17/pre-miR-17 were identified in the regulation expression of the NF-κB target gene, and the phylogenetic tree analysis showed that the pre-miR-17 of C. auratus with the closest similarity to the zebrafish Danio rerio is highly conserved in teleosts. This is the first report of the multi-omics analysis of the miRNAs and proteins in the gills of C. auratus infected with A. hydrophila, thus enriching knowledge on the regulation mechanism of the local immune response in Cyprinidae fish.

Multi-omics analysis of miRNA-mediated intestinal microflora changes in crucian carp Carassius auratus infected with Rahnella aquatilis

“…As previously reported ( 20 ), the differential expression level of the miRNAs was analyzed. The candidate gene functional enrichment of the differentially expressed miRNAs (DEMs) was analyzed using Gene Ontology (GO) and KEGG.…”

“…As an important molecular regulator of fish, the immune system protects the host from infections by pathogens. MiRNAs regulate systemic and mucosal immunity by targeting pattern recognition receptors and downstream signaling factors ( 19 , 20 ). Several kinds of miRNAs (e.g., miR-21b-3p, miR-146a-3p, and miR-155-5p) were found to effectively inhibit harmful inflammation or promote early immune response in Atlantic salmon Salmo salar ( 21 ).…”

“…Very recently, the integration of miRNA and mRNA analyses revealed the potential roles of the identified known miRNAs that are involved in the regulation of glycometabolism-related pathways in common carp Cyprinus carpio ( 26 ). Previous studies revealed that mucosal immune responses in crucian carp include the TLR, NOD-like receptor (NLR), and complement and coagulation pathways ( 20 , 27 , 28 ). However, there are few transcriptome analyses of miRNAs (e.g., miR-17, miR-26a, miR-144, and miR-146a) related to the involvement in the mucosal immune response of miRNA and the intestinal microflora regulation function in crucian carp ( 20 ).…”

“…Previous studies revealed that mucosal immune responses in crucian carp include the TLR, NOD-like receptor (NLR), and complement and coagulation pathways ( 20 , 27 , 28 ). However, there are few transcriptome analyses of miRNAs (e.g., miR-17, miR-26a, miR-144, and miR-146a) related to the involvement in the mucosal immune response of miRNA and the intestinal microflora regulation function in crucian carp ( 20 ).…”

“…In previous studies by our group, a new pathogenic R. aquatilis KCL-5 strain was isolated from diseased crucian carp ( 27 , 30 – 32 ). Subsequently, we detailedly studied the mucosal immunity (skin and gill) response of bacterial septicemia of crucian carp through multi-omics analysis ( 20 , 28 , 41 ). However, the molecular mechanism of the intestinal microflora, inflammation, and immune response-mediated miRNA–target gene axis in the mucosal immune regulation infected with R. aquatilis is still unclear.…”