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- Title
A stepwise pathway for biogenesis of 24-nt secondary siRNAs and spreading of DNA methylation.
- Authors
Daxinger, Lucia; Kanno, Tatsuo; Bucher, Etienne; van der Winden, Johannes; Naumann, Ulf; Matzke, Antonius J. M.; Matzke, Marjori
- Abstract
We used a transgene system to study spreading of RNA-directed DNA methylation (RdDM) during transcriptional gene silencing in Arabidopsis thaliana. Forward and reverse genetics approaches using this system delineated a stepwise pathway for the biogenesis of secondary siRNAs and unidirectional spreading of methylation from an upstream enhancer element into downstream sequences. Trans-acting, hairpin-derived primary siRNAs induce primary RdDM, independently of an enhancer-associated ‘nascent’ RNA, at the target enhancer region. Primary RdDM is a key step in the pathway because it attracts the secondary siRNA-generating machinery, including RNA polymerase IV, RNA-dependent RNA polymerase2 and Dicer-like3 (DCL3). These factors act in a turnover pathway involving a nascent RNA, which normally accumulates stably in non-silenced plants, to produce cis-acting secondary siRNAs that induce methylation in the downstream region. The identification of DCL3 in a forward genetic screen for silencing-defective mutants demonstrated a strict requirement for 24-nt siRNAs to direct methylation. A similar stepwise process for spreading of DNA methylation may occur in mammalian genomes, which are extensively transcribed in upstream regulatory regions.
- Subjects
SMALL interfering RNA; DNA; METHYLATION; GENE silencing; RNA polymerases; GENOMES
- Publication
EMBO Journal, 2009, Vol 28, Issue 1, p48
- ISSN
0261-4189
- Publication type
Article
- DOI
10.1038/emboj.2008.260