A DCL3 dicing code within Pol IV-RDR2 transcripts diversifies the siRNA pool guiding RNA-directed DNA methylation

  1. Andrew Loffer
  2. Jasleen Singh
  3. Akihito Fukudome
  4. Vibhor Mishra
  5. Feng Wang
  6. Craig S Pikaard  Is a corresponding author
  1. Indiana University Bloomington, United States
  2. Howard Hughes Medical Institute, Indiana University, United States

Abstract

In plants, selfish genetic elements including retrotransposons and DNA viruses are transcriptionally silenced by RNA-directed DNA methylation. Guiding the process are short interfering RNAs (siRNAs) cut by DICER-LIKE 3 (DCL3) from double-stranded precursors of ~30 bp that are synthesized by NUCLEAR RNA POLYMERASE IV (Pol IV) and RNA-DEPENDENT RNA POLYMERASE 2 (RDR2). We show that Pol IV's choice of initiating nucleotide, RDR2's initiation 1-2 nt internal to Pol IV transcript ends and RDR2's terminal transferase activity collectively yield a code that influences which precursor end is diced and whether 24 or 23 nt siRNAs are produced. By diversifying the size, sequence, and strand specificity of siRNAs derived from a given precursor, alternative patterns of DCL3 dicing allow for maximal siRNA coverage at methylated target loci.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting file; Source Data files have been provided for all figures.

Article and author information

Author details

  1. Andrew Loffer

    Department of Biology, Indiana University Bloomington, Bloomington, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Jasleen Singh

    Department of Biology, Indiana University Bloomington, Bloomington, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Akihito Fukudome

    Department of Biology, Indiana University Bloomington, Bloomington, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8924-1035
  4. Vibhor Mishra

    Department of Biology, Indiana University Bloomington, Bloomington, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Feng Wang

    Department of Biology, Indiana University Bloomington, Bloomington, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Craig S Pikaard

    Department of Biology, Howard Hughes Medical Institute, Indiana University, Bloomington, United States
    For correspondence
    cpikaard@indiana.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8204-7459

Funding

National Institutes of Health (GM077590)

  • Craig S Pikaard

Howard Hughes Medical Institute (Investigator:Pikaard)

  • Akihito Fukudome
  • Vibhor Mishra
  • Feng Wang
  • Craig S Pikaard

Indiana University Foundation (Carlos O. Miller Graduate Student Fellowship)

  • Andrew Loffer
  • Jasleen Singh

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

Reviewing Editor

  1. Daniel Zilberman, Institue of Science and Technology, Austria

Publication history

  1. Received: August 23, 2021
  2. Preprint posted: September 6, 2021 (view preprint)
  3. Accepted: January 28, 2022
  4. Accepted Manuscript published: January 31, 2022 (version 1)
  5. Version of Record published: February 15, 2022 (version 2)

Copyright

© 2022, Loffer 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. Andrew Loffer
  2. Jasleen Singh
  3. Akihito Fukudome
  4. Vibhor Mishra
  5. Feng Wang
  6. Craig S Pikaard
(2022)
A DCL3 dicing code within Pol IV-RDR2 transcripts diversifies the siRNA pool guiding RNA-directed DNA methylation
eLife 11:e73260.
https://doi.org/10.7554/eLife.73260
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