Abstract
MicroRNAs (miRNAs) are non-coding regulatory RNAs that play a vital part in the host immune response to pathogen infection. Japanese flounder (Paralichthys olivaceus) is an important aquaculture fish species that has suffered from bacterial diseases, including that caused by Vibrio anguillarum infection. In a previous study, we examined the messenger RNA (mRNA) expression profiles of flounder during V. anguillarum infection and identified 26 hub genes in the flounder immune response. In this study, we performed the micro-transcriptome analysis of flounder spleen in response to V. anguillarum infection at 3 different time points. Approximately 277 million reads were obtained, from which 1218 miRNAs were identified, including 740 known miRNAs and 478 novel miRNAs. Among the miRNAs, 206 were differentially expressed miRNAs (DEmiRs), and 104 of the 206 DEmiRs are novel miRNAs identified for the first time. Most of the DEmiRs were strongly time-dependent. A total of 1355 putative target genes of the DEmiRs (named DETGs) were identified based on integrated analysis of miRNA-mRNA expressions. The DETGs were enriched in multiple functional categories associated with immunity. Thirteen key DEmiRs and 66 immune DETGs formed an intricate regulatory network constituting 106 pairs of miRNAs and DETGs that span five immune pathways. Furthermore, seven of the previously identified hub genes were found to be targeted by 73 DEmiRs, and together they formed interlinking regulatory networks. These results indicate that V. anguillarum infection induces complicated miRNA response with extensive influences on immune gene expression in Japanese flounder.
Highlights
MicroRNAs of ~22 nucleotides represent the most well characterized small non-coding RNAs that regulate many fundamental biological processes such as growth, reproduction, and immunity [1,2]
Small RNA-sequencing is an efficient technology to systemically study the expression profiles of miRNAs under various conditions. Benefiting from this technique, many fish miRNAs involved in the immune response associated with pathogen infection have been identified. These miRNAs include the common carp (Cyprinus carpio) miRNAs induced by Flavobacterium columnare, Nile tilapia (Oreochromis niloticus) miRNAs induced by Streptococcus agalactiae, and half-smooth tongue sole (Cynoglossus semilaevis) and miiuy croaker (Miichthys miiuy) miRNAs induced by Vibrio anguillarum [8,9,10,11]
94.19% were aligned to known miRNAs, 0.27% were predicted to be novel miRNAs, 2.71% were identified as other small RNAs (sRNAs), containing ribosome RNAs, transfer RNA, small nuclear RNA, small nucleolar RNA, and small cytoplasmic RNA, and the remaining 2.83% were tags including unannotated sequences and repeated sequences
Summary
MicroRNAs (miRNAs) of ~22 nucleotides (nt) represent the most well characterized small non-coding RNAs that regulate many fundamental biological processes such as growth, reproduction, and immunity [1,2]. Small RNA-sequencing (sRNA-seq) is an efficient technology to systemically study the expression profiles of miRNAs under various conditions. Benefiting from this technique, many fish miRNAs involved in the immune response associated with pathogen infection have been identified. In Japanese flounder (Paralichthys olivaceus), sRNA-seq has been used to explore miRNAs participating in the infections of megalocytivirus, Edwardsiella tarda, and Streptococcus iniae [12,13,14]. These studies demonstrated that pathogens could significantly affect the expressions of large amounts of host miRNAs [15,16,17]
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