Probing tumor phenotypes using stable and regulated synthetic microRNA precursors

Dickins, R. A., Hemann, M. T., Zilfou, J. T., Simpson, D. R., Ibarra, I., Hannon, G. J., Lowe, S. W. (November 2005) Probing tumor phenotypes using stable and regulated synthetic microRNA precursors. Nat Genet, 37 (11). pp. 1289-95. ISSN 1061-4036 (Print)

URL: http://www.ncbi.nlm.nih.gov/pubmed/16200064
DOI: 10.1038/ng1651

Abstract

RNA interference is a powerful method for suppressing gene expression in mammalian cells. Stable knock-down can be achieved by continuous expression of synthetic short hairpin RNAs, typically from RNA polymerase III promoters. But primary microRNA transcripts, which are endogenous triggers of RNA interference, are normally synthesized by RNA polymerase II. Here we show that RNA polymerase II promoters expressing rationally designed primary microRNA-based short hairpin RNAs produce potent, stable and regulatable gene knock-down in cultured cells and in animals, even when present at a single copy in the genome. Most notably, by tightly regulating Trp53 knock-down using tetracycline-based systems, we show that cultured mouse fibroblasts can be switched between proliferative and senescent states and that tumors induced by Trp53 suppression and cooperating oncogenes regress upon re-expression of Trp53. In practice, this primary microRNA-based short hairpin RNA vector system is markedly similar to cDNA overexpression systems and is a powerful tool for studying gene function in cells and animals.

Item Type: Paper
Uncontrolled Keywords: Animals Cell Aging Cell Proliferation Fibroblasts metabolism Genetic Vectors Mice MicroRNAs genetics Neoplasms metabolism Oncogenes Phenotype Promoter Regions (Genetics) genetics Protein Synthesis Inhibitors pharmacology RNA Interference RNA Polymerase II genetics RNA Small Interfering pharmacology Tetracycline pharmacology Transcription Genetic Tumor Suppressor Protein p53 genetics metabolism
Subjects: bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > RNAi
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > shRNA
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > suppressor
CSHL Authors:
Communities: CSHL labs > Lowe lab
CSHL labs > Hannon lab
Watson School > Publications
Depositing User: CSHL Librarian
Date: November 2005
Date Deposited: 13 Jan 2012 16:10
Last Modified: 19 Sep 2014 15:44
Related URLs:
URI: http://repository.cshl.edu/id/eprint/22556

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