Genome-wide identification of Ago2 binding sites from mouse embryonic stem cells with and without mature microRNAs
Author(s)
Leung, Anthony; Young, Amanda G.; Bhutkar, Arjun (AJ); Zheng, Grace X.; Bosson, Andrew David; Nielsen, Cydney B.; Sharp, Phillip A.; ... Show more Show less
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MicroRNAs (miRNAs) are 19–22-nucleotide noncoding RNAs that post-transcriptionally regulate mRNA targets. We have identified endogenous miRNA binding sites in mouse embryonic stem cells (mESCs), by performing photo-cross-linking immunoprecipitation using antibodies to Argonaute (Ago2) followed by deep sequencing of RNAs (CLIP-seq). We also performed CLIP-seq in Dicer[superscript −/−] mESCs that lack mature miRNAs, allowing us to define whether the association of Ago2 with the identified sites was miRNA dependent. A significantly enriched motif, GCACUU, was identified only in wild-type mESCs in 3′ untranslated and coding regions. This motif matches the seed of a miRNA family that constitutes ~68% of the mESC miRNA population. Unexpectedly, a G-rich motif was enriched in sequences cross-linked to Ago2 in both the presence and absence of miRNAs. Expression analysis and reporter assays confirmed that the seed-related motif confers miRNA-directed regulation on host mRNAs and that the G-rich motif can modulate this regulation.
Date issued
2011-01Department
Massachusetts Institute of Technology. Computational and Systems Biology Program; Massachusetts Institute of Technology. Department of Biology; Koch Institute for Integrative Cancer Research at MITJournal
Nature Structural and Molecular Biology
Publisher
Nature Publishing Group
Citation
Leung, Anthony K L et al. “Genome-wide Identification of Ago2 Binding Sites from Mouse Embryonic Stem Cells with and Without Mature microRNAs.” Nature Structural & Molecular Biology 18.2 (2011): 237–244.
Version: Author's final manuscript
ISSN
1545-9993
1545-9985