Abstract
MicroRNAs are endogenous highly conserved short (~ 21 nucleotides) non-coding RNA molecules that play key roles in post-transcriptional gene regulation by translational inhibition or by target mRNA cleavage. In this report, using high stringent computational-based methods, a total of 101 putative miRNAs were identified from European sardine fish (Sardina pilchardus Walbaum, 1792). All the precursors of identified sardine miRNAs formed stable stem-loop structures and displayed high minimum free energy index (MFEI) values. For the experimental validation of the computationally predicted miRNAs, a tissue-specific quantitative study of eight randomly selected putative sardine miRNAs (spi-miR9, spi-miR26, spi-miR128, spi-miR129, spi-miR132, spi-miR212, spi-miR219, and spi-miR338) was performed in brain and liver and all the selected miRNAs were found to be overexpressed in brain tissue. Moreover, using RNAhybrid, a total of 83 potential target proteins of the characterized sardine miRNAs were identified those are involved in transcription, cellular development, defense mechanism, and various signaling pathways. To the best of our knowledge, this is the first report of sardine microRNAs and their targets.
Highlights
European sardine (Sardina pilchardus Walbaum, 1792), commonly known as European pilchard or sardine is one of the most abundant small pelagic fish in the world from the Clupeidae family that occurs mostly in the Atlantic Ocean and the Mediterranean Sea (Louro et al 2019) and widely consumed by humans
The evolutionarily conserved sequences of miRNAs across different species simplify the characterization process of new miRNA orthologues through computational based homology analysis (Sharma et al 2019); in silico miRNA identification only based on sequence similarity generates false-positive results and other stringent parameters of the predicted miRNA precursors such as minimum folding free energy (MFE), sequence length, GC content, and the minimum folding free energy index (MFEI) are required to increase the prediction precision (Paul et al 2018)
Since the stable secondary structure of the precursors is considered as one of the important factors to be a miRNA candidate some previously demonstrated strict filtering criteria were applied during secondary structure prediction such as: (1) the precursors must form a stem‐ loop structure containing mature miRNA sequences within one arm (2) the potential miRNA sequences should not be positioned at the terminal loop of the hairpin structures, (3) mature miRNAs should have fewer than nine mismatches with the opposite m iRNA* sequence, and (4) the predicted secondary structures must have low MFE and high MFEI values since it is required for distinguishing the miRNAs from other RNAs molecules (MFEIs of tRNAs, rRNAs or mRNAs candidates are 0.64, 0.59 and 0.62–0.66, respectively) (Zhang et al 2006)
Summary
European sardine (Sardina pilchardus Walbaum, 1792), commonly known as European pilchard or sardine is one of the most abundant small pelagic fish in the world from the Clupeidae family that occurs mostly in the Atlantic Ocean and the Mediterranean Sea (Louro et al 2019) and widely consumed by humans. MicroRNAs (miRNAs) are small endogenous ~ 21-nucleotide (nt) long non-coding regulatory RNA molecules that play a pivotal role in gene expression at the post-transcriptional level. It has been evidenced that miRNAs regulate a wide variety of biological processes such as cell cycle control, cell proliferation and differentiation, organ development, apoptosis, and stress response signaling in both animals and plants (Sun and Lai 2013; Paul et al 2011, 2020). Highly tissue-specific expression patterns during embryogenesis suggest that microRNAs play an important role in the differentiation and maintenance of tissue identity (Ribeiro et al 2014). Experimental validation of the predicted miRNAs is a crucial step to authenticate the prediction (Sharma et al 2019)
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