Generation of mammalian cells expressing stably measles virus proteins via bicistronic RNA.
The proteins of measles virus are believed to be cytotoxic, and have never been expressed stably from the cloned genes in cultured cells. We found that measles viral proteins can be expressed via a bicistronic RNA. The dominantly selectable DHFR* protein-coding region encoding a mutant dihydrofolate reductase was inserted into the 3'-untranslated regions of the measles viral genes encoding nucleoprotein (N), matrix (M) protein, and hemagglutinin (H). The tandemly arranged cistrons were placed under control by the inducible promoter of human metallothionein IIA gene, or the noninducible early promoter of simian virus 40. Upon transfecting into mammalian cells, these gene constructs synthesized bicistronic RNAs. The downstream DHFR* gene conferred resistance to methotrexate (MTX). Cells that survived MTX selection expressed stably the N, M, or H protein of measles virus. Expression of N protein was further inducible by cadmium chloride treatment. This system will be useful for studying the protein functions of measles virus, and could be applied to express other potentially toxic gene products.
Department of Microbiology, University of Washington School of Medicine, Seattle 98195.
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Hirano
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Wong
D000818:Animals; D002461:Cell Line, Transformed; D015870:Gene Expression; D005814:Genes, Viral; D005822:Genetic Vectors; D008459:Measles virus / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics*; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics; D014162:Transfection; D015678:Viral Structural Proteins / Q000235:genetics* / Q000633:toxicity; D014773:Virology / Q000379:methods
D012367:RNA, Viral; D015678:Viral Structural Proteins; D013762:Tetrahydrofolate Dehydrogenase