RNA sequence analysis defines Dicer's role in mouse embryonic stem cells
- *Center for Cancer Research and
- †Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139; and
- ‡Crick-Jacobs Center for Theoretical and Computational Biology, Salk Institute, 10010 North Torrey Pines Road, La Jolla, CA 92037
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Contributed by Phillip A. Sharp, September 27, 2007 (received for review August 13, 2007)
Abstract
Short RNA expression was analyzed from Dicer-positive and Dicer-knockout mouse embroyonic stem (ES) cells, using high-throughput pyrosequencing. A correlation of miRNA quantification with sequencing frequency estimates that there are 110,000 miRNAs per ES cell, the majority of which can be accounted for by six distinct miRNA loci. Four of these miRNA loci or their human homologues have demonstrated roles in cell cycle regulation or oncogenesis, suggesting that a major function of the miRNA pathway in ES cells may be to shape their distinct cell cycle. Forty-six previously uncharacterized miRNAs were identified, most of which are expressed at low levels and are less conserved than the set of known miRNAs. Low-abundance short RNAs matching all classes of repetitive elements were present in cells lacking Dicer, although the production of some SINE- and simple repeat-associated short RNAs appeared to be Dicer-dependent. These and other Dicer-dependent sequences resembled miRNAs. At a depth of sequencing that approaches the total number of 5′ phosphorylated short RNAs per cell, miRNAs appeared to be Dicer's only substrate. The results presented suggest a model in which repeat-associated miRNAs serve as host defenses against repetitive elements, a function canonically ascribed to other classes of short RNA.
Footnotes
- §To whom correspondence should be addressed. E-mail: sharppa{at}mit.edu
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Author contributions: J.M.C. and A.C.S. contributed equally to this work; J.M.C., A.C.S., and P.A.S. designed research; J.M.C. and A.C.S. performed research; G.W.Y. contributed new reagents/analytic tools; J.M.C., A.C.S., G.W.Y., and P.A.S. analyzed data; and J.M.C., A.C.S., and P.A.S. wrote the paper.
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The authors declare no conflict of interest.
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Data deposition: The data reported in this paper have been deposited in the Gene Expression Omnibus (GEO) database, www.ncbi.nlm.nih.gov/geo (accession no. GSE9306). Analyzed sequences are provided in SI Tables 9–13.
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This article contains supporting information online at www.pnas.org/cgi/content/full/0709193104/DC1.
- Abbreviation:
- miRNA,
- microRNA.
- © 2007 by The National Academy of Sciences of the USA
