Advertisement

Profiling of N6-Methyladenosine in Zika Virus RNA and Host Cellular mRNA

  • Gianluigi Lichinchi
  • Tariq M. RanaEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1870)

Abstract

N6-Methylation of adenosine (m6A) in viral RNA was first identified in Rous sarcoma virus, influenza virus, and SV40 virus several decades ago. More recently, human immunodeficiency virus-1, hepatitis C virus, and Zika virus (ZIKV) RNAs have also been reported to be modified by m6A, highlighting the significance of this posttranscriptional RNA modification. Nevertheless, the functions of m6A and their roles in the context of viral infections are not completely understood. Similarly, the mechanisms by which viral infections may alter the dynamics of the host RNA methylome have yet to be elucidated. Here, we describe an experimental protocol to obtain m6A profiles of ZIKV RNA and the host cell mRNA using methylated RNA immunoprecipitation–sequencing (MeRIP-Seq).

Key words

N6-Methylation of adenosine RNA methylome ZIKV RNA Methylated RNA immunoprecipitation–sequencing 

References

  1. 1.
    Linder B, Grozhik AV, Olarerin-George AO, Meydan C, Mason CE, Jaffrey SR (2015) Single-nucleotide-resolution mapping of m6A and m6Am throughout the transcriptome. Nat Methods 12(8):767–772.  https://doi.org/10.1038/nmeth.3453CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Meyer KD, Saletore Y, Zumbo P, Elemento O, Mason CE, Jaffrey SR (2012) Comprehensive analysis of mRNA methylation reveals enrichment in 3′ UTRs and near stop codons. Cell 149(7):1635–1646.  https://doi.org/10.1016/j.cell.2012.05.003CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Dominissini D, Moshitch-Moshkovitz S, Schwartz S, Salmon-Divon M, Ungar L, Osenberg S, Cesarkas K, Jacob-Hirsch J, Amariglio N, Kupiec M, Sorek R, Rechavi G (2012) Topology of the human and mouse m6A RNA methylomes revealed by m6A-seq. Nature 485(7397):201–206.  https://doi.org/10.1038/nature11112CrossRefGoogle Scholar
  4. 4.
    Batista PJ, Molinie B, Wang J, Qu K, Zhang J, Li L, Bouley DM, Lujan E, Haddad B, Daneshvar K, Carter AC, Flynn RA, Zhou C, Lim KS, Dedon P, Wernig M, Mullen AC, Xing Y, Giallourakis CC, Chang HY (2014) m(6)A RNA modification controls cell fate transition in mammalian embryonic stem cells. Cell Stem Cell 15(6):707–719.  https://doi.org/10.1016/j.stem.2014.09.019CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Kane SE, Beemon K (1985) Precise localization of m6A in Rous sarcoma virus RNA reveals clustering of methylation sites: implications for RNA processing. Mol Cell Biol 5(9):2298–2306CrossRefGoogle Scholar
  6. 6.
    Bokar JA, Shambaugh ME, Polayes D, Matera AG, Rottman FM (1997) Purification and cDNA cloning of the AdoMet-binding subunit of the human mRNA (N6-adenosine)-methyltransferase. RNA 3(11):1233–1247PubMedPubMedCentralGoogle Scholar
  7. 7.
    Schwartz S, Mumbach MR, Jovanovic M, Wang T, Maciag K, Bushkin GG, Mertins P, Ter-Ovanesyan D, Habib N, Cacchiarelli D, Sanjana NE, Freinkman E, Pacold ME, Satija R, Mikkelsen TS, Hacohen N, Zhang F, Carr SA, Lander ES, Regev A (2014) Perturbation of m6A writers reveals two distinct classes of mRNA methylation at internal and 5′ sites. Cell Rep 8(1):284–296.  https://doi.org/10.1016/j.celrep.2014.05.048CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Ping XL, Sun BF, Wang L, Xiao W, Yang X, Wang WJ, Adhikari S, Shi Y, Lv Y, Chen YS, Zhao X, Li A, Yang Y, Dahal U, Lou XM, Liu X, Huang J, Yuan WP, Zhu XF, Cheng T, Zhao YL, Wang X, Rendtlew Danielsen JM, Liu F, Yang YG (2014) Mammalian WTAP is a regulatory subunit of the RNA N6-methyladenosine methyltransferase. Cell Res 24(2):177–189.  https://doi.org/10.1038/cr.2014.3CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Patil DP, Chen CK, Pickering BF, Chow A, Jackson C, Guttman M, Jaffrey SR (2016) m(6)A RNA methylation promotes XIST-mediated transcriptional repression. Nature 537(7620):369–373.  https://doi.org/10.1038/nature19342CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Jia G, Fu Y, Zhao X, Dai Q, Zheng G, Yang Y, Yi C, Lindahl T, Pan T, Yang YG, He C (2011) N6-methyladenosine in nuclear RNA is a major substrate of the obesity-associated FTO. Nat Chem Biol 7(12):885–887.  https://doi.org/10.1038/nchembio.687CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Zheng G, Dahl JA, Niu Y, Fedorcsak P, Huang CM, Li CJ, Vagbo CB, Shi Y, Wang WL, Song SH, Lu Z, Bosmans RP, Dai Q, Hao YJ, Yang X, Zhao WM, Tong WM, Wang XJ, Bogdan F, Furu K, Fu Y, Jia G, Zhao X, Liu J, Krokan HE, Klungland A, Yang YG, He C (2013) ALKBH5 is a mammalian RNA demethylase that impacts RNA metabolism and mouse fertility. Mol Cell 49(1):18–29.  https://doi.org/10.1016/j.molcel.2012.10.015CrossRefGoogle Scholar
  12. 12.
    Krug RM, Morgan MA, Shatkin AJ (1976) Influenza viral mRNA contains internal N6-methyladenosine and 5′-terminal 7-methylguanosine in cap structures. J Virol 20(1):45–53PubMedPubMedCentralGoogle Scholar
  13. 13.
    Finkel D, Groner Y (1983) Methylations of adenosine residues (m6A) in pre-mRNA are important for formation of late simian virus 40 mRNAs. Virology 131(2):409–425CrossRefGoogle Scholar
  14. 14.
    Lichinchi G, Gao S, Saletore Y, Gonzalez GM, Bansal V, Wang Y, Mason CE, Rana TM (2016) Dynamics of the human and viral m(6)A RNA methylomes during HIV-1 infection of T cells. Nat Microbiol 1:16011.  https://doi.org/10.1038/nmicrobiol.2016.11CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Lichinchi G, Zhao BS, Wu Y, Lu Z, Qin Y, He C, Rana TM (2016) Dynamics of Human and Viral RNA Methylation during Zika Virus Infection. Cell Host Microbe 20(5):666–673.  https://doi.org/10.1016/j.chom.2016.10.002CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Kennedy EM, Bogerd HP, Kornepati AV, Kang D, Ghoshal D, Marshall JB, Poling BC, Tsai K, Gokhale NS, Horner SM, Cullen BR (2016) Posttranscriptional m(6)A Editing of HIV-1 mRNAs Enhances Viral Gene Expression. Cell Host Microbe 19(5):675–685.  https://doi.org/10.1016/j.chom.2016.04.002CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Tirumuru N, Zhao BS, Lu W, Lu Z, He C, Wu L (2016) N(6)-methyladenosine of HIV-1 RNA regulates viral infection and HIV-1 Gag protein expression. Elife 5. doi: https://doi.org/10.7554/eLife.15528
  18. 18.
    Gokhale NS, McIntyre ABR, McFadden MJ, Roder AE, Kennedy EM, Gandara JA, Hopcraft SE, Quicke KM, Vazquez C, Willer J, Ilkayeva OR, Law BA, Holley CL, Garcia-Blanco MA, Evans MJ, Suthar MS, Bradrick SS, Mason CE, Horner SM (2016) N6-methyladenosine in flaviviridae viral RNA genomes regulates infection. Cell Host Microbe 20(5):654–665.  https://doi.org/10.1016/j.chom.2016.09.015CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Pediatrics and Institute for Genomic MedicineUniversity of California San DiegoLa JollaUSA

Personalised recommendations