Journal
GENES & DEVELOPMENT
Volume 17, Issue 16, Pages 2036-2047Publisher
COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT
DOI: 10.1101/gad.1081603
Keywords
cytosine methylation; RNA-directed DNA methylation; RNA silencing; small RNAs; dicer
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Funding
- NIEHS NIH HHS [ES 07141, T32 ES007141] Funding Source: Medline
- NIGMS NIH HHS [R01 GM061148, GM61148] Funding Source: Medline
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In plants, replication of RNA viruses and RNA from highly transcribed transgenes can trigger DNA methylation. These systems accumulate diced small RNA (smRNA) products of double-stranded RNA (dsRNA) precursors, but it is not known which RNA species directs methylation. The methylated PAI tryptophan biosynthetic genes in Arabidopsis allow the study of methylation signals for endogenous genes with lower expression levels. The PAI genes are arranged as a tandem inverted repeat plus two singlet genes at unlinked loci. Here we show that the predominant PAI transcript initiates at a novel unmethylated promoter that lies upstream of one of the inverted repeat PAI genes. Suppressed transcription from the upstream promoter using transgene-directed silencing reduces methylation on the singlet PAI genes, but not on the inverted repeat, consistent with an RNA methylation signal. RNA gel blots detect normal PAI transcripts and dsRNA read-through species, but not diced smRNAs, suggesting that either precursor dsRNAs or subdetectable levels of smRNAs, below the threshold to effectively degrade PAI transcripts, serve as the PAI methylation signal. Thus, the lower expression endogenous gene system allows dissection of a RNA-directed methylation pathway distinct from RNA degradation pathways.
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