Identification of Pol IV and RDR2-dependent precursors of 24 nt siRNAs guiding de novo DNA methylation in Arabidopsis

Abstract
Article and author information
Metrics

Abstract

In Arabidopsis thaliana, abundant 24 nt siRNAs guide the cytosine methylation and silencing of transposons and a subset of genes. 24 nt siRNA biogenesis requires nuclear RNA Polymerase IV (Pol IV), RNA-DEPENDENT RNA POLYMERASE 2 (RDR2) and DICER-LIKE3 (DCL3). However, siRNA precursors are mostly undefined. We identified Pol IV and RDR2-dependent RNAs (P4R2 RNAs) that accumulate in dcl3 mutants and are diced into 24 nt RNAs by DCL3 in vitro. P4R2 RNAs are mostly 26-45 nt and initiate with a purine adjacent to a pyrimidine, characteristics shared by Pol IV transcripts generated in vitro. RDR2 terminal transferase activity, also demonstrated in vitro, may account for occasional non-templated nucleotides at P4R2 RNA 3' termini. 24 nt siRNAs primarily correspond to the 5' or 3' ends of P4R2 RNAs, suggesting a model whereby siRNAs are generated from either end of P4R2 duplexes by single dicing events.

Article and author information

Author details

Todd Blevins

: Institut de Biologie Moléculaire des Plantes du CNRS, UPR2357, Université de Strasbourg, Strasbourg, France

Competing interests
The authors declare that no competing interests exist.
Ram Podicheti

Center for Genomics and Bioinformatics, Indiana University, Bloomington, United States

Competing interests
The authors declare that no competing interests exist.
Vibhor Mishra

Department of Biology and Department of Molecular and Cellular Biochemistry, Indiana University, Bloomington, United States

Competing interests
The authors declare that no competing interests exist.
Michelle Marasco

Department of Biology and Department of Molecular and Cellular Biochemistry, Indiana University, Bloomington, United States

Competing interests
The authors declare that no competing interests exist.
Jing Wang

Department of Biology and Department of Molecular and Cellular Biochemistry, Indiana University, Bloomington, United States

Competing interests
The authors declare that no competing interests exist.
Douglas Rusch

Center for Genomics and Bioinformatics, Indiana University, Bloomington, United States

Competing interests
The authors declare that no competing interests exist.
Haixu Tang

School of Informatics and Computing, Indiana University, Bloomington, United States

Competing interests
The authors declare that no competing interests exist.
Craig S Pikaard

Howard Hughes Medical Institute, Indiana University, Bloomington, United States

For correspondence
cpikaard@indiana.edu

Competing interests
The authors declare that no competing interests exist.

Copyright

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.