Expression profile analyses of mettl8 in Oryzias latipes

Abstract

Methyltransferase-like 8 (mettl8) is a protein-coding gene that may demonstrate nucleic acid or protein methyltransferase activity. Although several members of the METTL protein family have been reported, the expression and function of this family are still poorly understood, especially in fish. Medaka (Oryzias latipes) is an important model organism with relatively complete genome information, and more and more genetic toolkits are available for this fish. The popularity of medaka among developmental biologists has led to important insights into vertebrate development. Here, we report the DNA sequence and expression of mettl8 in medaka. The full-length cDNA of medaka mettl8 is 1266 bp, and its predicted open reading frame codes for a protein with 393 amino acids. The predicted molecular mass was 45.8 kDa, and the theoretical isoelectric point was 8.61. It had a conserved methyltransferase domain in METTL8 proteins. Homology analysis revealed that medaka METTL8 clustered in close proximity with the METTL8 of Austrofundulus limnaeus and Nothobranchius furzeri within the Cyprinodontiformes branch, and the protein structure of METTL8 was highly conserved. During embryogenesis, the mettl8 transcript was highly expressed in early stages, while it persisted at a detectable level until the larvae stage. In adult fish, the RT-PCR result indicated that mettl8 mRNA was expressed in the brain, eye, skin, liver, intestine, ovary, and testis. Slice in situ hybridization analysis showed that mettl8 was highly expressed in the eye, intestine, ovary, and testis. The expression and distribution of mettl8 during embryogenesis were also demonstrated by whole mount in situ hybridization. The results indicated that the mettl8 is expressed significantly in the eye, somite, and otic vesicles. Immunofluorescence and Western blot analyses showed that METTL8 protein was present in both the nuclei and cytoplasm. This study lays a foundation for further research on the function of fish mettl8.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2
Fig. 3
Fig. 4

References

  1. Alexandrov A, Martzen MR, Phizicky EM (2002) Two proteins that form a complex are required for 7-methylguanosine modification of yeast tRNA RNA. 8:1253–1266

  2. Badri KR, Zhou Y, Dhru U, Aramgam S, Schuger L (2008) Effects of the SANT domain of tension-induced/inhibited proteins (TIPs), novel partners of the histone acetyltransferase p300, on p300 activity and TIP-6-induced adipogenesis. Mol Cell Biol 28:6358–6372. https://doi.org/10.1128/MCB.00333-08

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  3. Brawand D et al (2014) The genomic substrate for adaptive radiation in African cichlid fish. Nature 513:375–381. https://doi.org/10.1038/nature13726

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  4. Carmel L, Chorev M (2012) The function of introns. Front Genet 3:55

    PubMed  PubMed Central  Google Scholar 

  5. Chen XW, Jiang S, Gu YF, Shi ZY (2014) Molecular characterization and expression of cyp19a gene in Carassius auratus. J Fish Biol 85:516–522. https://doi.org/10.1111/jfb.12418

    CAS  Article  PubMed  Google Scholar 

  6. Cui Q et al (2017) m(6)A RNA methylation regulates the self-renewal and tumorigenesis of glioblastoma stem cells. Cell Rep 18:2622–2634. https://doi.org/10.1016/j.celrep.2017.02.059

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  7. Diez-Roux G, Banfi S, Sultan M, Geffers L, Anand S, Rozado D, Magen A, Canidio E, Pagani M, Peluso I, Lin-Marq N, Koch M, Bilio M, Cantiello I, Verde R, de Masi C, Bianchi SA, Cicchini J, Perroud E, Mehmeti S, Dagand E, Schrinner S, Nürnberger A, Schmidt K, Metz K, Zwingmann C, Brieske N, Springer C, Hernandez AM, Herzog S, Grabbe F, Sieverding C, Fischer B, Schrader K, Brockmeyer M, Dettmer S, Helbig C, Alunni V, Battaini MA, Mura C, Henrichsen CN, Garcia-Lopez R, Echevarria D, Puelles E, Garcia-Calero E, Kruse S, Uhr M, Kauck C, Feng G, Milyaev N, Ong CK, Kumar L, Lam M, Semple CA, Gyenesei A, Mundlos S, Radelof U, Lehrach H, Sarmientos P, Reymond A, Davidson DR, Dollé P, Antonarakis SE, Yaspo ML, Martinez S, Baldock RA, Eichele G, Ballabio A (2011) A high-resolution anatomical atlas of the transcriptome in the mouse embryo. PLoS Biol 9:e1000582. https://doi.org/10.1371/journal.pbio.1000582

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  8. Duff MO et al (2015) Genome-wide identification of zero nucleotide recursive splicing in Drosophila. Nature 521:376–379. https://doi.org/10.1038/nature14475

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  9. Gu H et al (2018) The STAT3 target mettl8 regulates mouse ESC differentiation via inhibiting the JNK pathway. Stem Cell Reports 10:1807–1820

    CAS  Article  Google Scholar 

  10. Gu YF, Fang Y, Jin Y, Dong WR, Xiang LX, Shao JZ (2011) Discovery of the DIGIRR gene from teleost fish: a novel Toll-IL-1 receptor family member serving as a negative regulator of IL-1 signaling. J Immunol 187:2514–2530

    CAS  Article  Google Scholar 

  11. Heyn H, Esteller M (2015) An adenine code for DNA: a second life for N6-methyladenine. Cell 161:710–713. https://doi.org/10.1016/j.cell.2015.04.021

    CAS  Article  PubMed  Google Scholar 

  12. Ignatova VV, Jansen P, Baltissen MP, Vermeulen M, Schneider R (2019) The interactome of a family of potential methyltransferases in HeLa cells. Sci Rep 9:6584. https://doi.org/10.1038/s41598-019-43010-2

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  13. Jakkaraju S, Zhe X, Pan D, Choudhury R, Schuger L (2005) TIPs are tension-responsive proteins involved in myogenic versus adipogenic differentiation. Dev Cell 9:39–49. https://doi.org/10.1016/j.devcel.2005.04.015

    CAS  Article  PubMed  Google Scholar 

  14. Katz JE, Dlakic M, Clarke S (2003) Automated identification of putative methyltransferases from genomic open reading frames. Mol Cell Proteomics 2:525–540

    CAS  Article  Google Scholar 

  15. Liu J et al (2014) A METTL3-METTL14 complex mediates mammalian nuclear RNA N6-adenosine methylation. Nat Chem Biol 10:93–95. https://doi.org/10.1038/nchembio.1432

    CAS  Article  Google Scholar 

  16. Lo L, Zhang Z, Hong N, Peng J, Hong Y (2008) 3640 unique EST clusters from the medaka testis and their potential use for identifying conserved testicular gene expression in fish and mammals. PLoS One 3:e3915

    Article  Google Scholar 

  17. Ma CJ, Ding JH, Ye TT, Yuan BF, Feng YQ (2019) AlkB homologue 1 demethylates N(3)-methylcytidine in mRNA of mammals. ACS Chem Biol 14:1418–1425. https://doi.org/10.1021/acschembio.8b01001

    CAS  Article  PubMed  Google Scholar 

  18. Niewmierzycka A, Clarke S (1999) S-Adenosylmethionine-dependent methylation in Saccharomyces cerevisiae identification of a novel protein arginine methyltransferase. J Biol Chem 274:814–824. https://doi.org/10.1074/jbc.274.2.814

    CAS  Article  PubMed  Google Scholar 

  19. Pendleton KE, Chen B, Liu K, Hunter OV, Xie Y, Tu BP, Conrad NK (2017) The U6 snRNA m(6)A methyltransferase METTL16 regulates SAM synthetase intron retention. Cell 169:824–835 e814. https://doi.org/10.1016/j.cell.2017.05.003

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  20. Shimazu T, Barjau J, Sohtome Y, Sodeoka M, Shinkai Y (2014) Selenium-based S-adenosylmethionine analog reveals the mammalian seven-beta-strand methyltransferase METTL10 to be an EF1A1 lysine methyltransferase. PLoS One 9:e105394

    Article  Google Scholar 

  21. Siedlecki P, Zielenkiewicz P (2006) Mammalian DNA methyltransferases. Acta Biochim Pol 53:245–256

    CAS  Article  Google Scholar 

  22. Tobi EW et al (2018) DNA methylation as a mediator of the association between prenatal adversity and risk factors for metabolic disease in adulthood. Sci Adv 4:eaao4364

    Article  Google Scholar 

  23. Tooley CE, Petkowski JJ, Muratore-Schroeder TL, Balsbaugh JL, Shabanowitz J, Sabat M, Minor W, Hunt DF, Macara IG (2010) NRMT is an alpha-N-methyltransferase that methylates RCC1 and retinoblastoma protein. Nature 466:1125–1128

    Article  Google Scholar 

  24. Walter RB, Li HY, Intano GW, Kazianis S, Walter CA (2002) Absence of global genomic cytosine methylation pattern erasure during medaka (Oryzias latipes) early embryo development. Comp Biochem Physiol B Biochem Mol Biol 133:597–607. https://doi.org/10.1016/s1096-4959(02)00144-6

    Article  PubMed  Google Scholar 

  25. Wang H et al (2019) Mettl3-mediated mRNA m(6)A methylation promotes dendritic cell activation. Nat Commun 10:1898. https://doi.org/10.1038/s41467-019-09903-6

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  26. Wang X, Bhandari RK (2019) DNA methylation dynamics during epigenetic reprogramming of medaka embryo. Epigenetics 14:611–622. https://doi.org/10.1080/15592294.2019.1605816

    Article  PubMed  PubMed Central  Google Scholar 

  27. Xie S et al (2019) An integrated analysis of mRNA and miRNA in skeletal muscle from myostatin-edited Meishan pigs. Genome 62:305–315. https://doi.org/10.1139/gen-2018-0110

    CAS  Article  PubMed  Google Scholar 

  28. Xu H, Gui J, Hong Y (2005) Differential expression of vasa RNA and protein during spermatogenesis and oogenesis in the gibel carp (Carassius auratus gibelio), a bisexually and gynogenetically reproducing vertebrate. Dev Dyn 233:872–882. https://doi.org/10.1002/dvdy.20410

    CAS  Article  PubMed  Google Scholar 

  29. Xu L, Liu X, Sheng N, Oo KS, Liang J, Chionh YH, Xu J, Ye F, Gao YG, Dedon PC, Fu XY (2017) Three distinct 3-methylcytidine (m(3)C) methyltransferases modify tRNA and mRNA in mice and humans. J Biol Chem 292:14695–14703. https://doi.org/10.1074/jbc.M117.798298

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  30. Yeon SY, Jo YS, Choi EJ, Kim MS, Yoo NJ, Lee SH (2018) Frameshift mutations in repeat sequences of ANK3, HACD4, TCP10L, TP53BP1, MFN1, LCMT2, RNMT, TRMT6, METTL8 and METTL16 genes in colon cancers. Pathol Oncol Res 24:617–622. https://doi.org/10.1007/s12253-017-0287-2

    CAS  Article  PubMed  Google Scholar 

Download references

Acknowledgments

We would like to express our sincere thanks to Dr. Yunhan Hong from the National University of Singapore for providing the medaka fish and experimental materials for in situ hybridization in 2013.

Funding

This work was supported by grants from the National Natural Science Foundation of China, 81874142, to Yi-Feng Gu and the Natural Science Foundation of Shanghai (grant number 12ZR1412900).

Author information

Affiliations

Authors

Contributions

X.W.C. and Y.F.G drafted the experiments. Z.W.Z., W.P., Y.F.G, and Y.H.B. performed the experiments. Y.F.G and Y.W.S. analyzed the data. W.P. and X.W.C. wrote the paper. All authors read and approved the final version of the manuscript.

Corresponding authors

Correspondence to Yifeng Gu or Xiaowu Chen.

Ethics declarations

Competing interests

The authors declare that they have no competing interests.

Additional information

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

Figure S1

Phylogenetic analysis of METTL8family. The tree was generated using the neighbor-joining method by MEGA X software. Deduced amino acid sequences were aligned using ClustalX and the phylogenetic tree was constructed using bootstrap maximum likelihood tree (1000 replicates) method. medaka mettl1was used as an outgroup. (PNG 989 kb)

Figure S2

RT-PCR analysis of medakamettl8RNA expression. (A) Different developing stages. (B) Different adult organs. β-actinwas used as a control gene. (PNG 105 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Pang, W., Zhao, Z., Shen, Y. et al. Expression profile analyses of mettl8 in Oryzias latipes. Fish Physiol Biochem 46, 971–979 (2020). https://doi.org/10.1007/s10695-020-00764-1

Download citation

Keywords

  • Whole mount in situ hybridization
  • Methyltransferase-like 8
  • Methyltransferase
  • Oryzias latipes