U6 promoter-driven siRNAs with four uridine 3′ overhangs efficiently suppress targeted gene expression in mammalian cells

• Published in: Nature biotechnology Vol. 20; no. 5; pp. 497 - 500
• Main Authors: Taira, Kazunari, Miyagishi, Makoto
• Format: Journal Article
• Language: English
• Published: New York, NY Nature 01.05.2002; Nature Publishing Group
• Subjects: Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy; Animals; Applied cell therapy and gene therapy; Biological and medical sciences; Biotechnology; Blotting, Northern; Blotting, Western; Caenorhabditis elegans; Dose-Response Relationship, Drug; Drosophila; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation; Gene therapy; Gene Transfer Techniques; Genes, Reporter; Genetic Vectors; Health. Pharmaceutical industry; HeLa Cells; Humans; Industrial applications and implications. Economical aspects; Mammals; Medical sciences; Microscopy, Fluorescence; Models, Genetic; Nematoda; Oligonucleotides, Antisense - pharmacology; Protein synthesis; RNA, Small Interfering; RNA, Small Nuclear - metabolism; RNA, Untranslated - metabolism; Transfection; Transfusions. Complications. Transfusion reactions. Cell and gene therapy; Uridine - chemistry; Gene silencing; Vertebrata; Cell line; Mammalia; Double stranded RNA; RNA interference; Small interfering RNA; Gene expression; Gene therapy; Vector; Medical application
• ISSN: 1087-0156; 1546-1696
• DOI: 10.1038/nbt0502-497

The first evidence for gene disruption by double-stranded RNA (dsRNA) came from careful analysis in Caenorhabditis elegans. This phenomenon, called RNA interference (RNAi), was observed subsequently in various organisms, including plants, nematodes, Drosophila, and protozoans. Very recently, it has been reported that in mammalian cells, 21- or 22-nucleotide (nt) RNAs with 2-nt 3' overhangs (small inhibitory RNAs, siRNAs) exhibit an RNAi effect. This is because siRNAs are not recognized by the well-characterized host defense system against viral infections, involving dsRNA-dependent inhibition of protein synthesis. However, the current method for introducing synthetic siRNA into cells by lipofection restricts the range of applications of RNAi as a result of the low transfection efficiencies in some cell types and/or short-term persistence of silencing effects. Here, we report a vector-based siRNA expression system that can induce RNAi in mammalian cells. This technical advance for silencing gene expression not only facilitates a wide range of functional analysis of mammalian genes but might also allow therapeutic applications by means of vector-mediated RNAi.