Evolution of the myosin heavy chain gene MYH14 and its intronic microRNA miR-499: muscle-specific miR-499 expression persists in the absence of the ancestral host gene

<jats:title>Abstract</jats:title> <jats:sec> <jats:title>Background</jats:title> <jats:p>A novel sarcomeric myosin heavy chain gene, <jats:italic>MYH14</jats:italic>, was identified following the completion of the human genome project. <jats:italic>MYH14</jats:italic> contains an intronic microRNA, miR-499, which is expressed in a slow/cardiac muscle specific manner along with its host gene; it plays a key role in muscle fiber-type specification in mammals. Interestingly, teleost fish genomes contain multiple <jats:italic>MYH14</jats:italic> and miR-499 paralogs. However, the evolutionary history of <jats:italic>MYH14</jats:italic> and miR-499 has not been studied in detail. In the present study, we identified <jats:italic>MYH14</jats:italic>/miR-499 loci on various teleost fish genomes and examined their evolutionary history by sequence and expression analyses.</jats:p> </jats:sec> <jats:sec> <jats:title>Results</jats:title> <jats:p>Synteny and phylogenetic analyses depict the evolutionary history of <jats:italic>MYH14</jats:italic>/miR-499 loci where teleost specific duplication and several subsequent rounds of species-specific gene loss events took place. Interestingly, miR-499 was not located in the <jats:italic>MYH14</jats:italic> introns of certain teleost fish. An <jats:italic>MYH14</jats:italic> paralog, lacking miR-499, exhibited an accelerated rate of evolution compared with those containing miR-499, suggesting a putative functional relationship between <jats:italic>MYH14</jats:italic> and miR-499. In medaka, <jats:italic>Oryzias latipes</jats:italic>, miR-499 is present where <jats:italic>MYH14</jats:italic> is completely absent in the genome. Furthermore, by using <jats:italic>in situ</jats:italic> hybridization and small RNA sequencing, miR-499 was expressed in the notochord at the medaka embryonic stage and slow/cardiac muscle at the larval and adult stages. Comparing the flanking sequences of <jats:italic>MYH14</jats:italic>/miR-499 loci between torafugu <jats:italic>Takifugu rubripes</jats:italic>, zebrafish <jats:italic>Danio rerio</jats:italic>, and medaka revealed some highly conserved regions, suggesting that <jats:italic>cis</jats:italic>-regulatory elements have been functionally conserved in medaka miR-499 despite the loss of its host gene.</jats:p> </jats:sec> <jats:sec> <jats:title>Conclusions</jats:title> <jats:p>This study reveals the evolutionary history of the <jats:italic>MYH14</jats:italic>/miRNA-499 locus in teleost fish, indicating divergent distribution and expression of <jats:italic>MYH14</jats:italic> and miR-499 genes in different teleost fish lineages. We also found that medaka miR-499 was even expressed in the absence of its host gene. To our knowledge, this is the first report that shows the conversion of intronic into non-intronic miRNA during the evolution of a teleost fish lineage.</jats:p> </jats:sec>