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Genome-wide identification of targets and function of individual MicroRNAs in mouse embryonic stem cells.

TitleGenome-wide identification of targets and function of individual MicroRNAs in mouse embryonic stem cells.
Publication TypeJournal Article
Year of Publication2010
AuthorsHanina SA, Mifsud W, Down TA, Hayashi K, O'Carroll D, Lao K, Miska EA, M Surani A
JournalPLoS Genet
Date Published2010 Oct
Keywords3' Untranslated Regions, Animals, Argonaute Proteins, Cells, Cultured, DEAD-box RNA Helicases, Embryonic Stem Cells, Endoribonucleases, Eukaryotic Initiation Factor-2, Gene Expression Profiling, Gene Regulatory Networks, Genome, Green Fluorescent Proteins, Mice, Mice, 129 Strain, Mice, Knockout, MicroRNAs, Mutation, Oligonucleotide Array Sequence Analysis, Proto-Oncogene Proteins c-myc, Recombinant Fusion Proteins, Reverse Transcriptase Polymerase Chain Reaction, Ribonuclease III, RNA, Messenger, RNA-Binding Proteins, Transfection

Mouse Embryonic Stem (ES) cells express a unique set of microRNAs (miRNAs), the miR-290-295 cluster. To elucidate the role of these miRNAs and how they integrate into the ES cell regulatory network requires identification of their direct regulatory targets. The difficulty, however, arises from the limited complementarity of metazoan miRNAs to their targets, with the interaction requiring as few as six nucleotides of the miRNA seed sequence. To identify miR-294 targets, we used Dicer1-null ES cells, which lack all endogenous mature miRNAs, and introduced just miR-294 into these ES cells. We then employed two approaches to discover miR-294 targets in mouse ES cells: transcriptome profiling using microarrays and a biochemical approach to isolate mRNA targets associated with the Argonaute2 (Ago2) protein of the RISC (RNA Induced Silencing Complex) effector, followed by RNA-sequencing. In the absence of Dicer1, the RISC complexes are largely devoid of mature miRNAs and should therefore contain only transfected miR-294 and its base-paired targets. Our data suggest that miR-294 may promote pluripotency by regulating a subset of c-Myc target genes and upregulating pluripotency-associated genes such as Lin28.

Alternate JournalPLoS Genet.
PubMed ID20975942
PubMed Central IDPMC2958809
Grant ListG0800784 / / Medical Research Council / United Kingdom
RG49135 / / Wellcome Trust / United Kingdom
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