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Dicer recognizes the 5¡ä end of RNA for efficient and accurate processing

hallmark of RNA silencing is a class of approximately 22-nucleotide RNAs that are processed from double-stranded RNA precursors by Dicer. Accurate processing by Dicer is crucial for the functionality of microRNAs (miRNAs). The current model posits that Dicer selects cleavage sites by measuring a set distance from the 3¡ä overhang of the double-stranded RNA terminus. Here we report that human Dicer anchors not only the 3¡ä end but also the 5¡ä end, with the cleavage site determined mainly by the distance (~22 nucleotides) from the 5¡ä end (5¡ä counting rule). This cleavage requires a 5¡ä-terminal phosphate group. Further, we identify a novel basic motif (5¡ä pocket) in human Dicer that recognizes the 5¡ä-phosphorylated end. The 5¡ä counting rule and the 5¡ä anchoring residues are conserved in Drosophila Dicer-1, but not in Giardia Dicer. Mutations in the 5¡ä pocket reduce processing efficiency and alter cleavage sites in vitro. Consistently, miRNA biogenesis is perturbed in vivo when Dicer-null embryonic stem cells are replenished with the 5¡ä-pocket mutant. Thus, 5¡ä-end recognition by Dicer is important for precise and effective biogenesis of miRNAs. Insights from this study should also afford practical benefits to the design of small hairpin RNAs.

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