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RESEARCH PAPER
Department of Biology, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6018, USA
The Arabidopsis genes, TAS2 and TAS1a, produce structurally similar noncoding transcripts that are transformed into short (21-nucleotide [nt]) and long (24-nt) siRNAs by RNA silencing pathways. Some of these short siRNAs direct the cleavage of protein-coding transcripts, and thus function as trans-acting siRNAs (ta-siRNAs). Using genetic analysis, we defined the pathway by which ta-siRNAs and other short siRNAs are generated from these loci. This process is initiated by the miR173-directed cleavage of a primary poly(A) transcript. The 3' fragment is then transformed into short siRNAs by the sequential activity of SGS3, RDR6, and DCL4: SGS3 stabilizes the fragment, RDR6 produces a complementary strand, and DCL4 cleaves the resulting double-stranded molecule into short siRNAs, starting at the end with the miR173 cleavage site and proceeding in 21-nt increments from this point. The 5' cleavage fragment is also processed by this pathway, but less efficiently. The DCL3-dependent pathway that generates long siRNAs does not require miRNA-directed cleavage and plays a minor role in the silencing of these loci. Our results define the core components of a post-transcriptional gene silencing pathway in Arabidopsis and reveal some of the features that direct transcripts to this pathway.
[Keywords: PTGS; RNAi; miRNA; trans-acting siRNAs]
Received July 7, 2005; revised version accepted August 2, 2005.
Article published online ahead of print. Article and publication date are at http://www.genesdev.org/cgi/doi/10.1101/gad.1352605.
1 Corresponding author.
E-MAIL spoethig{at}sas.upenn.edu; FAX (215) 898-8780.
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