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Nat Struct Mol Biol. 2010 Feb;17(2):173-9. doi: 10.1038/nsmb.1745. Epub 2010 Jan 10.

Comprehensive discovery of endogenous Argonaute binding sites in Caenorhabditis elegans.

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Department of Biology, Stem Cell Program, University of California, San Diego, La Jolla, California, USA.


MicroRNAs (miRNAs) regulate gene expression by guiding Argonaute proteins to specific target mRNA sequences. Identification of bona fide miRNA target sites in animals is challenging because of uncertainties regarding the base-pairing requirements between miRNA and target as well as the location of functional binding sites within mRNAs. Here we present the results of a comprehensive strategy aimed at isolating endogenous mRNA target sequences bound by the Argonaute protein ALG-1 in C. elegans. Using cross-linking and ALG-1 immunoprecipitation coupled with high-throughput sequencing (CLIP-seq), we identified extensive ALG-1 interactions with specific 3' untranslated region (UTR) and coding exon sequences and discovered features that distinguish miRNA complex binding sites in 3' UTRs from those in other genic regions. Furthermore, our analyses revealed a striking enrichment of Argonaute binding sites in genes important for miRNA function, suggesting an autoregulatory role that may confer robustness to the miRNA pathway.

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