Abstract
MicroRNAs (miRNAs) are small RNA molecules that play key roles in regulation of various biological processes. In order to better understand the biological significance of miRNAs in the context of Aeromonas hydrophila infection in Megalobrama amblycephala, small RNA libraries obtained from fish liver at 0 (non-infection), 4, and 24 h post infection (poi) were sequenced using Illumina deep sequencing technology. A total of 11,244,207, 9,212,958, and 7,939,157 clean reads were obtained from these three RNA libraries, respectively. Bioinformatics analysis identified 171 conserved miRNAs and 62 putative novel miRNAs. The existence of ten randomly selected novel miRNAs was validated by RT-PCR. Pairwise comparison suggested that 61 and 44 miRNAs were differentially expressed at 4 and 24 h poi, respectively. Furthermore, the expression profiles of nine randomly selected miRNAs were validated by qRT-PCR. MicroRNA target prediction, gene ontology (GO) annotation, and Kyoto Encylopedia of Genes and Genomes (KEGG) analysis indicated that a variety of biological pathways could be affected by A. hydrophila infection. Additionally, transferrin (TF) and transferrin receptor (TFR) genes were confirmed to be direct targets of miR-375. These results will expand our knowledge of the role of miRNAs in the immune response of M. amblycephala to A. hydrophila infection, and facilitate the development of effective strategies against A. hydrophila infection in M. amblycephala.
Highlights
MicroRNAs, typically ~22 nucleotides in length, are small non-coding RNAs, and produced from longer, hairpin-shaped precursors by two RNase III proteins, Dicer and Drosha with the assistance of DiGeorge syndrome critical region 8 (DGCR8) [1,2]
Mature miRNAs are loaded onto Argonaute proteins (AGOs) to form RNA-induced silencing complex (RISC) in which miRNAs function as a guide by base pairing with target mRNAs, leading to cleavage of target mRNAs or translation repression
The small RNA transcriptome of A. hydrophila-infected M. amblycephala was analyzed in the liver because the organ plays important roles in immune responses in both mammals and fish, in fish responses to A. hydrophila infection [26,27,28,29,30]
Summary
MicroRNAs (miRNAs), typically ~22 nucleotides (nt) in length, are small non-coding RNAs, and produced from longer, hairpin-shaped precursors (pre-miRNAs) by two RNase III proteins, Dicer and Drosha with the assistance of DiGeorge syndrome critical region 8 (DGCR8) [1,2]. A multitude of studies demonstrate that miRNAs play crucial roles in the regulation of diverse biological processes, including cell differentiation, apoptosis, and immune defense [13,14]. Increasing studies have revealed that miRNAs are crucial for host responses to pathogen infection in eukaryotes. A subset of miRNAs, including miR-146, miR-155, miR-21 and let-7 family members, have been found to contribute to immune responses during bacterial infections [15,16,17]. Recent studies demonstrate that miRNAs play pivotal roles in the metabolism of iron, an essential element for the growth and survival of most organisms including fish and bacteria, and a key regulator of host-pathogen interactions [18,19,20,21]
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