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이용선 (Yong Sun Lee) 저자 이메일 보기
University of Virginia
 
조회 1890  인쇄하기 주소복사 트위터 공유 페이스북 공유 
Depletion of Human Micro-RNA miR-125b Reveals That It Is Critical for the Proliferation of Differentiated Cells but Not for the Down-regulation of Putative Targets during Differentiation
Yong Sun Lee‡, Hak Kyun Kim‡§, Sangmi Chung¶, Kwang-Soo Kim¶ and Anindya Dutta‡∥

Department of Biochemistry and Molecular Genetics, University of Virginia, Charlottesville, Virginia 22908, §Institute of Molecular Biology and Genetics, School of Biological Sciences, Seoul National University, Seoul 151-742, Korea, and Molecular Neurobiology Laboratory, McLean Hospital, Harvard Medical School, Belmont, Massachusetts 02478

To whom correspondence should be addressed. Tel.: 434-924-1227; Fax: 434-924-5069.

Abstract

Micro-RNAs are small non-coding RNAs that regulate target gene expression post-transcriptionally through base pairing with the target messenger RNA. Functional characterization of micro-RNAs awaits robust experimental methods to knock-down a micro-RNA as well as to assay its function in vivo. In addition to the recently developed method to sequester micro-RNA with 2′-O-methyl antisense oligonucleotide, we report that small interfering RNA against the loop region of a micro-RNA precursor can be used to deplete the micro-RNA. The depletion of miR-125b by this method had a profound effect on the proliferation of adult differentiated cancer cells, and this proliferation defect was rescued by co-transfected mature micro-RNA. This technique has unique advantages over the 2′-O-methyl antisense oligonucleotide and can be used to determine micro-RNA function, assay micro-RNAs in vivo, and identify the contribution of a predicted micro-RNA precursor to the pool of mature micro-RNA in a given cell. miR-125b and let-7 micro-RNAs are induced, whereas their putative targets, lin-28 and lin-41, are decreased during in vitro differentiation of Tera-2 or embryonic stem cells. Experimental increase or decrease of micro-RNA concentrations did not, however, affect the levels of the targets, a finding that is explained by the fact that the down-regulation of the targets appears to be mostly at the transcriptional level in these in vitro differentiation systems. Collectively these results reveal the importance of micro-RNA depletion strategies for directly determining micro-RNA function in vivo.

Footnotes

1 The abbreviations used are: miRNA, micro-RNA; siRNA, small interfering RNA; NP, neuronal precursor; EC, embryonic carcinoma; ES cell, embryonic stem cell; gapdh, glyceraldehyde-3-phosphate dehydrogenase gene; EB, embryoid bodies.

2 Y. S. Lee and A. Dutta, unpublished data.

* This work was partly supported by a post-doctoral fellowship program of the Korea Science and Engineering Foundation (to Y. S. L.) and National Institutes of Health Grant RO1 CA89406 (to A. D.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Received October 28, 2004. Revision received February 15, 2005.

논문정보
- 형식: Research article
- 게재일: 2005년 04월 (BRIC 등록일 2009-11-05)
- 연구진: 국내+국외 연구진
- 피인용횟수: 120회 이상 인용된 논문
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