RNA binding to human METTL3-METTL14 restricts N6-deoxyadenosine methylation of DNA in vitro

  1. Shan Qi
  2. Javier Mota
  3. Siu-Hong Chan
  4. Johanna Villarreal
  5. Nan Dai
  6. Shailee Arya
  7. Robert A Hromas
  8. Manjeet K Rao
  9. Ivan R Corrêa Jr
  10. Yogesh K Gupta  Is a corresponding author
  1. The University of Texas Health Science Center at San Antonio, United States
  2. New England Biolabs, United States

Abstract

Methyltransferase like-3 (METTL3) and METTL14 complex transfers a methyl group from S-adenosyl-L-methionine to N6 amino group of adenosine bases in RNA (m6A) and DNA (m6dA). Emerging evidence highlights a role of METTL3-METTL14 in the chromatin context, especially in processes where DNA and RNA are held in close proximity. However, a mechanistic framework about specificity for substrate RNA/DNA and their interrelationship remain unclear. By systematically studying methylation activity and binding affinity to a number of DNA and RNA oligos with different propensities to form inter- or intra-molecular duplexes or single-stranded molecules in vitro, we uncover an inverse relationship for substrate binding and methylation and show that METTL3-METTL14 preferentially catalyzes the formation of m6dA in single-stranded DNA (ssDNA), despite weaker binding affinity to DNA. In contrast, it binds structured RNAs with high affinity, but methylates the target adenosine in RNA (m6A) much less efficiently than it does in ssDNA. We also show that METTL3-METTL14-mediated methylation of DNA is largely restricted by structured RNA elements prevalent in long noncoding and other cellular RNAs.

Data availability

The information about coding sequences of human METTL3 (NCBI reference sequence GI: 33301371) and METTL14 (NCBI reference sequence GI: 172045930) used in this study is available at NCBI. Source data are provided as a separate Source Data file. Correspondence and requests for material should be addressed to Y.K.G. (guptay@uthscsa.edu).

Article and author information

Author details

  1. Shan Qi

    Department of Biochemistry and Structural Biology, The University of Texas Health Science Center at San Antonio, San Antonio, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-0175-6267
  2. Javier Mota

    Greehey Children's Cancer Research Institute, The University of Texas Health Science Center at San Antonio, San Antonio, United States
    Competing interests
    No competing interests declared.
  3. Siu-Hong Chan

    RNA Biology, New England Biolabs, Ipswich, United States
    Competing interests
    Siu-Hong Chan, is an employee of New England Biolabs, a manufacturer and vendor of molecular biology reagents..
  4. Johanna Villarreal

    Greehey Children's Cancer Research Institute, The University of Texas Health Science Center at San Antonio, San Antonio, United States
    Competing interests
    No competing interests declared.
  5. Nan Dai

    RNA Biology, New England Biolabs, Ipswich, United States
    Competing interests
    Nan Dai, is an employee of New England Biolabs, a manufacturer and vendor of molecular biology reagents..
  6. Shailee Arya

    Greehey Children's Cancer Research Institute, The University of Texas Health Science Center at San Antonio, San Antonio, United States
    Competing interests
    No competing interests declared.
  7. Robert A Hromas

    Department of Medicine, The University of Texas Health Science Center at San Antonio, San Antonio, United States
    Competing interests
    Robert A Hromas, owns equity in Dialectic Therapeutics and Abfero..
  8. Manjeet K Rao

    Greehey Children's Cancer Research Institute, The University of Texas Health Science Center at San Antonio, San Antonio, United States
    Competing interests
    No competing interests declared.
  9. Ivan R Corrêa Jr

    RNA Biology, New England Biolabs, Ipswich, United States
    Competing interests
    Ivan R Corrêa, is an employee of New England Biolabs, a manufacturer and vendor of molecular biology reagents..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3169-6878
  10. Yogesh K Gupta

    Department of Biochemistry and Structural Biology, The University of Texas Health Science Center at San Antonio, San Antonio, United States
    For correspondence
    guptay@uthscsa.edu
    Competing interests
    Yogesh K Gupta, is founder of Atomic Therapeutics. None of these affiliations affect the authors' impartiality, adherence to journal standards and policies, or availability of data..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6372-5007

Funding

Max and Minnie Tomerlin Voelcker Fund

  • Yogesh K Gupta

San Antonio Area Foundation

  • Yogesh K Gupta

IIMS/CTSA pilot award

  • Yogesh K Gupta

Greehey Children's Cancer Research Institute

  • Shan Qi
  • Yogesh K Gupta

University of Texas System

  • Yogesh K Gupta

National Institute of Allergy and Infectious Diseases (1R01AI161363)

  • Yogesh K Gupta

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Shozeb Haider, University College London, United Kingdom

Version history

  1. Received: February 2, 2021
  2. Preprint posted: January 9, 2022 (view preprint)
  3. Accepted: January 20, 2022
  4. Accepted Manuscript published: January 21, 2022 (version 1)
  5. Version of Record published: February 9, 2022 (version 2)
  6. Version of Record updated: April 4, 2022 (version 3)

Copyright

© 2022, Qi et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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  1. Shan Qi
  2. Javier Mota
  3. Siu-Hong Chan
  4. Johanna Villarreal
  5. Nan Dai
  6. Shailee Arya
  7. Robert A Hromas
  8. Manjeet K Rao
  9. Ivan R Corrêa Jr
  10. Yogesh K Gupta
(2022)
RNA binding to human METTL3-METTL14 restricts N6-deoxyadenosine methylation of DNA in vitro
eLife 11:e67150.
https://doi.org/10.7554/eLife.67150

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