Skip to main content
PMC home page
NIHPA Author Manuscripts logoLink to NIHPA Author Manuscripts
. Author manuscript; available in PMC: 2015 Aug 10.
Published in final edited form as: Gene. 2014 May 15;546(2):386–389. doi: 10.1016/j.gene.2014.05.036

Expanding the annotation of zebrafish microRNAs based on smallRNA sequencing

Thomas Desvignes a,*, Michael J Beam a, Peter Batzel a, Jason Sydes a,b, John H Postlethwait a
PMCID: PMC4130647  NIHMSID: NIHMS603464  PMID: 24835514

Abstract

MicroRNAs (miRs) are short non-coding RNAs that fine-tune the regulation of gene expression to coordinate a wide range of biological processes. Because of their role in the regulation of gene expression, miRs are essential players in development by acting on cell fate determination and progression towards cell differentiation and are increasingly relevant to human health and disease. Although the zebrafish Danio rerio is a major model for studies of development, genetics, physiology, evolution, and human biology, the annotation of zebrafish miR-producing genes remains limited. In the present work, we report deep sequencing data of zebrafish smallRNAs from brain, heart, testis, and ovary. Results provide evidence for the expression of 56 un-annotated mir genes and 248 un-annotated mature strands, increasing the number of zebrafish mir genes over those already deposited in miRBase by 16% and the number of mature sequences by 63%. We also describe the existence of three pairs of mirror-mir genes and two mirtron genes, genetic features previously undescribed in non-mammalian vertebrates. This report provides information that substantially increases our knowledge of the zebrafish miRNome and will benefit the entire miR community.

Keywords: miRNA, zebrafish, teleost, smallRNA-seq, mirror-mir, mirtron

Introduction

MicroRNA (miRs) are short non-coding RNAs that fine-tune the regulation of gene expression to coordinate a wide range of biological processes (Ambros, 2004; Kosik, 2010). MicroRNAs are transcribed from mir genes and primary miR transcripts are processed to approximately 22 nucleotide single strand mature forms that function as repressors of transcript translation when bound to the 3’UTR of protein-coding transcripts in association with the RISC (Davis and Hata, 2009; Kim et al., 2009). Due to their role in the regulation of gene expression, miRs are essential players in development by acting on cell fate determination and progression towards cell differentiation (Ivey and Srivastava, 2010). the zebrafish Danio rerio is a major model for studies of development, genetics, physiology, evolution, and human health and disease, and many miR-related discoveries have come from zebrafish investigations (Giraldez et al., 2005; Wienholds et al., 2005; Giraldez et al., 2006; Flynt et al., 2007; He et al., 2009; Mishima et al., 2009; He et al., 2011a, 2011b). For instance, mir140 was shown to regulate zebrafish palatal development by down-regulating the translation of Pdgfra protein in neural crest derived cells (Eberhart et al., 2008). Experiments stimulated by that discovery showed that a single nucleotide polymorphism in the human MIR140 gene leads to improper mature MIR140 production and contributes to nonsyndromic cleft palate susceptibility in patients (Li et al., 2010). Another example of successful transfer of discoveries from zebrafish to human is the discovery of the crucial role of mir451 in zebrafish erythropoiesis (Pase et al., 2009), which was later confirmed in mouse (Rasmussen et al., 2010; Patrick et al., 2010) and recently in human (Kim et al., 2013), thereby highlighting the therapeutic potential of miR inhibitors. Despite the high impact of zebrafish research in many aspects of development and physiology, the annotation of zebrafish miR-producing genes remains limited.

This limitation in our knowledge is a broad problem because the annotation of the zebrafish genome, the most complete among fish (Howe et al., 2013), often drives annotation in all teleost fish, which constitute half of all vertebrate species (Nelson, 2006). Moreover, a largely incomplete characterization of the miR repertoire compromises studies on miRNA evolution, their emergence and retention in vertebrates, especially in the rayfin fish lineage following the teleost genome duplication. Furthermore, uncovering and transferring evolutionarily conserved miR functions from fish models to human health depends on recognizing true fish orthologs of human miRs. Thus, the entire miR community would benefit substantially by a better understanding of the zebrafish miRNome.

In the present report, we provide extensive miRNA sequencing data from several zebrafish tissues demonstrating the expression of new mir genes and the expression of previously un-annotated mature strands. Further, we describe for the first time in non-mammalian vertebrates, the expression of mirror-mirs (Scott et al., 2012; Tyler et al., 2008) and mirtrons (Chung et al., 2011; Ladewig et al., 2012; Okamura et al., 2007).

Materials and methods

Small RNAs were extracted using the Norgen microRNA purification kit from four different organs (brain, heart, testis and ovary). Animals were actively reproductive adult “AB” strain zebrafish (Danio rerio) obtained from the University of Oregon fish facility. From two males, we sampled brain, heart, and testes and made six libraries, three for each individual; from two females, we sampled ovaries and made two libraries, one for each individual. Eight tissue-specific sequencing libraries were prepared and barcoded using the BiooScientific NEXTflex smallRNA Sequencing Kit which uses a 3’ adenylated adapter that ligates onto miRs and other small RNAs with a 3’ hydroxyl group, following the manufacturer’s instructions. Libraries were subsequently sequenced by Illumina HiSeq2500 at the University of Oregon Genomics Core Facility and raw single-end 50nt long reads were deposited in the NCBI Short Reads Archive under the accession number SRP039502. All animal work was performed according to the University of Oregon IACUC approved protocol (#09–1BRRA).

Bioinformatic processing involved removing 3’ adapter sequences and filtering reads based on quality; reads with any base Q<30 were removed using the FASTX-Toolkit (http://hannonlab.cshl.edu/fastx_toolkit/). A custom pipeline was created to remove reads outside of the targeted size range (<15 and >28), count the number of occurrences of each read, and remove reads with low counts (<30 summed across all eight libraries). The script then aligns reads to the Zv9 version of the zebrafish reference genome using blastn (Altschul et al., 1990) allowing up to five mismatches, one gap, and alignments at up to 20 locations in the genome; finally, the script groups reads together based on their genomic locations. Aligned groups were annotated against mature and hairpin sequences already annotated in miRBase (release 20)(Kozomara and Griffiths-Jones, 2013) as well as zebrafish noncoding RNA sequences annotated by Ensembl (release 74)(Flicek et al., 2012).

Putative new mirs were predicted based on read presence in our experimental dataset and secondary structures were computed on the RNAfold web server (Zuker and Stiegler, 1981) using default parameters except for changing the calculation of minimum free energy from 37°C to 28°C, the temperature at which zebrafish are reared (Westerfield, 2000). A tentative name was assigned to newly recognized sequences based on identity with deposited sequences in miRBase (Kozomara and Griffiths-Jones, 2013) for zebrafish or other species.

Results/discussion

The sequencing of small RNAs with 3’ hydroxyls from several tissues of adult zebrafish yielded a total of 59.5 million reads that passed all filtering criteria, among which 47.5 million (79.8%) could be directly annotated from mature zebrafish entries in miRBase due to sequence identity. Among remaining reads, sequences mapping perfectly to the zebrafish reference genome at fewer than 20 locations were further studied for putative new mir genes and/or for mature strand annotation.

The annotation pipeline identified 56 mir genes that do not appear in miRBase, although 43 have of them have been bioinformatically predicted as “Novel miRNA” genes in Ensembl (Flicek et al., 2012); thus, our work increases the number of annotated zebrafish mir genes by 16% compared to the most recent miRBase (Release20) (Kozomara and Griffiths-Jones, 2013) (Table 1). Most of these new mir genes (54/56) are clear homologs to genes belonging to known mir families or described in related species and could thus be easily added to their corresponding group.

Table 1.

Summary of improvements to zebrafish annotations in miRBase.

Zebrafish miRBase
Release 20		 Present
study		 % increase Total
mir genes 346 56 16 402
mature sequences 391 248 63 639
mirror-mir pairs 0 3 N/A 3
mirtrons 0 2 N/A 2
Open in a new tab

Two of the miR-encoding genes we identified in our experimental data, tentatively named dre-mir-733-2 and dre-mir-733b, are paralogs of dre-mir-733 (MI0004778) and originate from vaultRNAs, and a third miR, tentatively named dre-mir-735b, a paralog of dre-mir-735 (MI0004781), actually originates from a Y_RNA gene. Human orthologs (hsa-mir-886 and hsa-mir-1979 respectively) of these zebrafish genes were withdrawn from miRBase after their biogenesis was understood (Stadler et al., 2009; Meiri et al., 2010). The role of vaultRNA and Y_RNA short fragments in translation repression is, however, not fully understood and they may act in a “mir-like” manner (Meiri et al., 2010; Persson et al., 2009; Nicolas et al., 2012), as do some snoRNA-derived or lncRNA-derived miRs (Yang and Lai, 2011). If the vaultRNA and Y_RNA derived short sequences do act like miRs, then production of short RNA fragments from vaultRNA and Y_RNA would be a new non-canonical way of generating miRs (Verhagen and Pruijn, 2011).

Our zebrafish smallRNA-seq dataset contained two completely new mir genes that showed no similarity with sequences yet deposited in miRBase for any species. These two new mirs, along with the 54 mir genes in our experimental dataset that are predicted in Ensembl but not referenced in miRBase, are described in the Supplemental File, which gives a tentative name, genomic localizations, predicted hairpin sequences, corresponding secondary structures, minimum free energy calculated at 28°C, the localization of mature sequences on the hairpin sequence, and secondary structure.

The breadth and depth of our sequencing dataset also allowed us to provide annotation for a total of 248 mature strands, 158 of them corresponding to previously annotated mir genes and 90 from the set of 56 new mir genes not present yet in miRBase and described here above, increasing the total number of annotated zebrafish mature sequences from 391 to 639 (a 63% increase) (Table 1). The Supplemental File gives the sequence, and genomic location of each newly identified miR.

Among the newly annotated structures are miRs with several interesting features. The zebrafish smallRNA-seq dataset clearly contained three pairs of mirror-mirs (Table 1), which are defined as a pair of mir genes originating from overlapping genomic regions but on opposite strands (Tyler et al., 2008; Scott et al., 2012). For example, the expression of mir3120, mirror-mir of mir214, has been described in mammals but in no other species (Scott et al., 2012; Desvignes et al., 2014). Here we show that miR3120, a mature fragment originating from the mirror-mir of mir214, is expressed in adult zebrafish. We also found the pair mir7547 and mir7553, which had been previously identified as independent mirs in the channel catfish Ictalurus punctatus (MI0024671 and MI0024685 respectively)(Xu et al., 2013), but had not been described as a mirror-mir pair, probably due to the lack of a reference genome. The newly annotated zebrafish mir7552b and its mirror gene, mir7552bOS, had also not previously been annotated in zebrafish, even though a homolog of mir7552b is annotated in the channel catfish (MI0024684)(Xu et al., 2013). This finding is of particular interest given that, to our knowledge, mirror-mirs have been described so far only in Drosophila and mammals (Tyler et al., 2008; Scott et al., 2012). Evidence given in this report demonstrating the expression of mirror-mirs in zebrafish suggests that mirror-mirs are likely a type of gene featured in all vertebrates.

Finally, we also report the presence and expression of a pair of 3’-tailed mirtron paralogs located in gria3 ohnologs (gria3a, ENSDARG00000032737; and gria3b, ENSDARG00000037498) (Table 1). These mir genes are co-orthologous mirtrons to the predicted human mirtron gene AL356213.1 (ENSG00000265082), which is located in GRIA3 (ENSG00000125675); they lie at orthologous positions in both zebrafish ohnologs; and they show high similarities with tetrapod mir2985, which is deposited in miRBase in zebra finch Taeniopygia guttata, platypus Ornithorhynchus anatinus and rat Rattus norvegicus as intronic MiRs of the Gria2 genes (Kozomara and Griffiths-Jones, 2013). Similar to mirror-mirs, to our knowledge, mirtrons have only been shown in Drosophila, the nematode Caenorhabditis elegans and mammals (Okamura et al., 2007; Chung et al., 2011; Ladewig et al., 2012). This report is thus the first example of mirtrons in a non-mammalian vertebrate.

Conclusion

The present report analyzing smallRNA deep sequencing data expands the annotation of microRNA genes and mature strands in the laboratory model zebrafish Danio rerio, whose mir gene repertoire remains understudied and poorly characterized in comparison to other species, such as Drosophila, worms, mouse, or human, despite its relevance for microRNA research. Increasing the annottion of zebrafish miRs will contribute to a better understanding of the evolution of mir genes in vertebrates and within the rayfin fish lineage, will help annotate mir genes in other teleost species, and will increase confidence in the annotation of each gene thanks to the description of both mature strands, considered as a criterion for mir gene annotation confidence (Kozomara and Griffiths-Jones, 2013).

Supplementary Material

01. Supplemental File.

Supplemental File: New mir genes and miR mature strand annotation. The first tab, “New mir genes”, lists all our newly identified mir genes based on our sequencing data. We give each new mir gene a tentative name based on homology and conserved synteny searches to identify orthology; its corresponding Ensembl accession number, if it exists or ‘N/A’ if not; the sequence of the predicted precursor miR or hairpin and its genomic location in the zebrafish Zv9 assembly; predicted secondary structure and corresponding minimum free energy calculated at 28°C. The second tab, “Mature miR strands”, lists all the zebrafish mir genes (already in miRBase or newly annotated) with information on corresponding genomic location and mature strand sequence annotation and position within the hairpin. The mature sequence given is the isomiR showing the highest expression in our datasets and might differ slightly from the miRBase entry that is based on sequencing datasets from different tissues. mir genes clustering information is also provided. The mir genes and mature miRs annotated in this study are highlighted in blue and yellow respectively, and sequences that were not found in our sequencing data are highlighted in green.

Highlights.

  • Annotation of 56 new miRNA genes in zebrafish

  • Annotation of 248 new mature miRNA strands in zebrafish

  • First description of mirror-miR pairs outside of Drosophila and mammals

  • First description of 3’-tailed mirtrons outside of Drosophila, worms and mammals

Acknowledgements

Authors would like to thank the University of Oregon fish facility crew, especially Trevor Enright, for help in fish husbandry, and members of the Postlethwait Lab and of the Institute of Neuroscience of the University of Oregon for helpful discussions. This work was funded by NIH grants U01 DE020076, R01 OD011116 awarded to JHP and 5R240D011199 to W. Cresko and JHP.

Abbreviations list

miR

microRNA

smallRNA-seq

small RNA sequencing

UTR

untranslated region

RISC

RNA-induced silencing complex

Footnotes

Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

Contributor Information

Thomas Desvignes, Email: desvignes@uoneuro.uoregon.edu.

Michael J. Beam, Email: mbeam@uoregon.edu.

Peter Batzel, Email: pbatzel@uoregon.edu.

Jason Sydes, Email: sydes@uoregon.edu.

John H. Postlethwait, Email: jpostle@uoneuro.uoregon.edu.

References

  1. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ. Basic local alignment search tool. J. Mol. Biol. 1990;215:403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  2. Ambros V. The functions of animal microRNAs. Nature. 2004;431:350–355. doi: 10.1038/nature02871. [DOI] [PubMed] [Google Scholar]
  3. Chung W-J, Agius P, Westholm JO, Chen M, Okamura K, Robine N, Leslie CS, Lai EC. Computational and experimental identification of mirtrons in Drosophila melanogaster and Caenorhabditis elegans. Genome Res. 2011;21:286–300. doi: 10.1101/gr.113050.110. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Davis BN, Hata A. Regulation of MicroRNA Biogenesis: A miRiad of mechanisms. Cell Commun. Signal. 2009;7:18. doi: 10.1186/1478-811X-7-18. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Desvignes T, Contreras A, Postlethwait JH. Evolution of the miR199–214 cluster and vertebrate skeletal development. RNA Biol. 2014;11:0–1. doi: 10.4161/rna.28141. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Eberhart JK, He X, Swartz ME, Yan Y-L, Song H, Boling TC, Kunerth AK, Walker MB, Kimmel CB, Postlethwait JH. MicroRNA Mirn140 modulates Pdgf signaling during palatogenesis. Nat. Genet. 2008;40:290–298. doi: 10.1038/ng.82. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Flicek P, Ahmed I, Amode MR, Barrell D, Beal K, Brent S, Carvalho-Silva D, Clapham P, Coates G, Fairley S, Fitzgerald S, Gil L, Garcia-Giron C, Gordon L, Hourlier T, Hunt S, Juettemann T, Kahari AK, Keenan S, Komorowska M, Kulesha E, Longden I, Maurel T, McLaren WM, Muffato M, Nag R, Overduin B, Pignatelli M, Pritchard B, Pritchard E, Riat HS, Ritchie GRS, Ruffier M, Schuster M, Sheppard D, Sobral D, Taylor K, Thormann A, Trevanion S, White S, Wilder SP, Aken BL, Birney E, Cunningham F, Dunham I, Harrow J, Herrero J, Hubbard TJP, Johnson N, Kinsella R, Parker A, Spudich G, Yates A, Zadissa A, Searle SMJ. Ensembl 2013. Nucleic Acids Res. 2012;41:D48–D55. doi: 10.1093/nar/gks1236. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Flynt AS, Li N, Thatcher EJ, Solnica-Krezel L, Patton JG. Zebrafish miR-214 modulates Hedgehog signaling to specify muscle cell fate. Nat. Genet. 2007;39:259–263. doi: 10.1038/ng1953. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Giraldez AJ, Cinalli RM, Glasner ME, Enright AJ, Thomson JM, Baskerville S, Hammond SM, Bartel DP, Schier AF. MicroRNAs Regulate Brain Morphogenesis in Zebrafish. Science. 2005;308:833–838. doi: 10.1126/science.1109020. [DOI] [PubMed] [Google Scholar]
  10. Giraldez AJ, Mishima Y, Rihel J, Grocock RJ, Dongen SV, Inoue K, Enright AJ, Schier AF. Zebrafish MiR-430 Promotes Deadenylation and Clearance of Maternal mRNAs. Science. 2006;312:75–79. doi: 10.1126/science.1122689. [DOI] [PubMed] [Google Scholar]
  11. He X, Eberhart JK, Postlethwait JH. MicroRNAs and micromanaging the skeleton in disease, development and evolution. J. Cell. Mol. Med. 2009;13:606–618. doi: 10.1111/j.1582-4934.2009.00696.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. He X, Yan Y-L, DeLaurier A, Postlethwait JH. Observation of miRNA gene expression in zebrafish embryos by in situ hybridization to microRNA primary transcripts. Zebrafish. 2011a;8:1–8. doi: 10.1089/zeb.2010.0680. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. He X, Yan Y-L, Eberhart JK, Herpin A, Wagner TU, Schartl M, Postlethwait JH. miR-196 regulates axial patterning and pectoral appendage initiation. Dev. Biol. 2011b;357:463–477. doi: 10.1016/j.ydbio.2011.07.014. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Howe K, Clark MD, Torroja CF, Torrance J, Berthelot C, Muffato M, Collins JE, Humphray S, McLaren K, Matthews L, McLaren S, Sealy I, Caccamo M, Churcher C, Scott C, Barrett JC, Koch R, Rauch G-J, White S, Chow W, Kilian B, Quintais LT, Guerra-Assuncao JA, Zhou Y, Gu Y, Yen J, Vogel J-H, Eyre T, Redmond S, Banerjee R, Chi J, Fu B, Langley E, Maguire SF, Laird GK, Lloyd D, Kenyon E, Donaldson S, Sehra H, Almeida-King J, Loveland J, Trevanion S, Jones M, Quail M, Willey D, Hunt A, Burton J, Sims S, McLay K, Plumb B, Davis J, Clee C, Oliver K, Clark R, Riddle C, Eliott D, Threadgold G, Harden G, Ware D, Mortimer B, Kerry G, Heath P, Phillimore B, Tracey A, Corby N, Dunn M, Johnson C, Wood J, Clark S, Pelan S, Griffiths G, Smith M, Glithero R, Howden P, Barker N, Stevens C, Harley J, Holt K, Panagiotidis G, Lovell J, Beasley H, Henderson C, Gordon D, Auger K, Wright D, Collins J, Raisen C, Dyer L, Leung K, Robertson L, Ambridge K, Leongamornlert D, McGuire S, Gilderthorp R, Griffiths C, Manthravadi D, Nichol S, Barker G, Whitehead S, Kay M, Brown J, Murnane C, Gray E, Humphries M, Sycamore N, Barker D, Saunders D, Wallis J, Babbage A, Hammond S, Mashreghi-Mohammadi M, Barr L, Martin S, Wray P, Ellington A, Matthews N, Ellwood M, Woodmansey R, Clark G, Cooper J, Tromans A, Grafham D, Skuce C, Pandian R, Andrews R, Harrison E, Kimberley A, Garnett J, Fosker N, Hall R, Garner P, Kelly D, Bird C, Palmer S, Gehring I, Berger A, Dooley CM, Ersan-Urun Z, Eser C, Geiger H, Geisler M, Karotki L, Kirn A, Konantz J, Konantz M, Oberlander M, Rudolph-Geiger S, Teucke M, Osoegawa K, Zhu B, Rapp A, Widaa S, Langford C, Yang F, Carter NP, Harrow J, Ning Z, Herrero J, Searle SMJ, Enright A, Geisler R, Plasterk RHA, Lee C, Westerfield M, de Jong PJ, Zon LI, Postlethwait JH, Nusslein-Volhard C, Hubbard TJP, Crollius HR, Rogers J, Stemple DL. The zebrafish reference genome sequence and its relationship to the human genome. Nature. 2013 doi: 10.1038/nature12111. advance online publication. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Ivey KN, Srivastava D. MicroRNAs as Regulators of Differentiation and Cell Fate Decisions. Cell Stem Cell. 2010;7:36–41. doi: 10.1016/j.stem.2010.06.012. [DOI] [PubMed] [Google Scholar]
  16. Kim M, Tan YS, Cheng W-C, Civin CI. MiR-144 and MiR-451 Regulate Human Erythropoiesis By Targeting RAB14. Blood. 2013;122:942–942. [Google Scholar]
  17. Kim VN, Han J, Siomi MC. Biogenesis of small RNAs in animals. Nat. Rev. Mol. Cell Biol. 2009;10:126–139. doi: 10.1038/nrm2632. [DOI] [PubMed] [Google Scholar]
  18. Kosik KS. MicroRNAs and Cellular Phenotypy. Cell. 2010;143:21–26. doi: 10.1016/j.cell.2010.09.008. [DOI] [PubMed] [Google Scholar]
  19. Kozomara A, Griffiths-Jones S. miRBase: annotating high confidence microRNAs using deep sequencing data. Nucleic Acids Res. 2013;42:D68–D73. doi: 10.1093/nar/gkt1181. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Ladewig E, Okamura K, Flynt AS, Westholm JO, Lai EC. Discovery of hundreds of mirtrons in mouse and human small RNA data. Genome Res. 2012;22:1634–1645. doi: 10.1101/gr.133553.111. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Li L, Meng T, Jia Z, Zhu G, Shi B. Single nucleotide polymorphism associated with nonsyndromic cleft palate influences the processing of miR-140. Am. J. Med. Genet. A. 2010;152A:856–862. doi: 10.1002/ajmg.a.33236. [DOI] [PubMed] [Google Scholar]
  22. Meiri E, Levy A, Benjamin H, Ben-David M, Cohen L, Dov A, Dromi N, Elyakim E, Yerushalmi N, Zion O, Lithwick-Yanai G, Sitbon E. Discovery of microRNAs and other small RNAs in solid tumors. Nucleic Acids Res. 2010;38:6234–6246. doi: 10.1093/nar/gkq376. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Mishima Y, Abreu-Goodger C, Staton AA, Stahlhut C, Shou C, Cheng C, Gerstein M, Enright AJ, Giraldez AJ. Zebrafish miR-1 and miR-133 shape muscle gene expression and regulate sarcomeric actin organization. Genes Dev. 2009;23:619–632. doi: 10.1101/gad.1760209. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Nelson JS. Fishes of the World. John Wiley & Sons; 2006. [Google Scholar]
  25. Nicolas FE, Hall AE, Csorba T, Turnbull C, Dalmay T. Biogenesis of Y RNA-derived small RNAs is independent of the microRNA pathway. FEBS Lett. 2012;586:1226–1230. doi: 10.1016/j.febslet.2012.03.026. [DOI] [PubMed] [Google Scholar]
  26. Okamura K, Hagen JW, Duan H, Tyler DM, Lai EC. The Mirtron Pathway Generates microRNA-Class Regulatory RNAs in Drosophila. Cell. 2007;130:89–100. doi: 10.1016/j.cell.2007.06.028. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Pase L, Layton JE, Kloosterman WP, Carradice D, Waterhouse PM, Lieschke GJ. miR-451 regulates zebrafish erythroid maturation in vivo via its target gata2. Blood. 2009;113:1794–1804. doi: 10.1182/blood-2008-05-155812. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Patrick DM, Zhang CC, Tao Y, Yao H, Qi X, Schwartz RJ, Huang LJ-S, Olson EN. Defective erythroid differentiation in miR-451 mutant mice mediated by 14-3-3ζ. Genes Dev. 2010;24:1614–1619. doi: 10.1101/gad.1942810. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Persson H, Kvist A, Vallon-Christersson J, Medstrand P, Borg Å, Rovira C. The non-coding RNA of the multidrug resistance-linked vault particle encodes multiple regulatory small RNAs. Nat. Cell Biol. 2009;11:1268–1271. doi: 10.1038/ncb1972. [DOI] [PubMed] [Google Scholar]
  30. Rasmussen KD, Simmini S, Abreu-Goodger C, Bartonicek N, Giacomo MD, Bilbao-Cortes D, Horos R, Lindern MV, Enright AJ, O’Carroll D. The miR-144/451 locus is required for erythroid homeostasis. J. Exp. Med. 2010;207:1351–1358. doi: 10.1084/jem.20100458. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Scott H, Howarth J, Lee YB, Wong L-F, Bantounas I, Phylactou L, Verkade P, Uney JB. MiR-3120 Is a Mirror MicroRNA That Targets Heat Shock Cognate Protein 70 and Auxilin Messenger RNAs and Regulates Clathrin Vesicle Uncoating. J. Biol. Chem. 2012;287:14726–14733. doi: 10.1074/jbc.M111.326041. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Stadler PF, Chen JJ-L, Hackermüller J, Hoffmann S, Horn F, Khaitovich P, Kretzschmar AK, Mosig A, Prohaska SJ, Qi X, Schutt K, Ullmann K. Evolution of vault RNAs. Mol. Biol. Evol. 2009;26:1975–1991. doi: 10.1093/molbev/msp112. [DOI] [PubMed] [Google Scholar]
  33. Tyler DM, Okamura K, Chung W-J, Hagen JW, Berezikov E, Hannon GJ, Lai EC. Functionally distinct regulatory RNAs generated by bidirectional transcription and processing of microRNA loci. Genes Dev. 2008;22:26–36. doi: 10.1101/gad.1615208. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Verhagen APM, Pruijn GJM. Are the Ro RNP-associated Y RNAs concealing microRNAs? Y RNA-derived miRNAs may be involved in autoimmunity. BioEssays. 2011;33:674–682. doi: 10.1002/bies.201100048. [DOI] [PubMed] [Google Scholar]
  35. Westerfield M. A guide for the laboratory use of zebrafish (Danio rerio) 4th ed. Eugene: Univ. of Oregon Press; 2000. The zebrafish book. [Google Scholar]
  36. Wienholds E, Kloosterman WP, Miska E, Alvarez-Saavedra E, Berezikov E, Bruijn E, de, Horvitz HR, Kauppinen S, Plasterk RHA. MicroRNA Expression in Zebrafish Embryonic Development. Science. 2005;309:310–311. doi: 10.1126/science.1114519. [DOI] [PubMed] [Google Scholar]
  37. Xu Z, Chen J, Li X, Ge J, Pan J, Xu X. Identification and Characterization of MicroRNAs in Channel Catfish (Ictalurus punctatus) by Using Solexa Sequencing Technology. PLoS ONE. 2013;8:e54174. doi: 10.1371/journal.pone.0054174. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Yang J-S, Lai EC. Alternative miRNA Biogenesis Pathways and the Interpretation of Core miRNA Pathway Mutants. Mol. Cell. 2011;43:892–903. doi: 10.1016/j.molcel.2011.07.024. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Zuker M, Stiegler P. Optimal computer folding of large RNA sequences using thermodynamics and auxiliary information. Nucleic Acids Res. 1981;9:133–148. doi: 10.1093/nar/9.1.133. [DOI] [PMC free article] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

01. Supplemental File.

Supplemental File: New mir genes and miR mature strand annotation. The first tab, “New mir genes”, lists all our newly identified mir genes based on our sequencing data. We give each new mir gene a tentative name based on homology and conserved synteny searches to identify orthology; its corresponding Ensembl accession number, if it exists or ‘N/A’ if not; the sequence of the predicted precursor miR or hairpin and its genomic location in the zebrafish Zv9 assembly; predicted secondary structure and corresponding minimum free energy calculated at 28°C. The second tab, “Mature miR strands”, lists all the zebrafish mir genes (already in miRBase or newly annotated) with information on corresponding genomic location and mature strand sequence annotation and position within the hairpin. The mature sequence given is the isomiR showing the highest expression in our datasets and might differ slightly from the miRBase entry that is based on sequencing datasets from different tissues. mir genes clustering information is also provided. The mir genes and mature miRs annotated in this study are highlighted in blue and yellow respectively, and sequences that were not found in our sequencing data are highlighted in green.

NIHMS603464-supplement-01.xls (264KB, xls)

RESOURCES