A role for RNA processing in regulating expression from transfected genes.
We have examined the expression of cloned genes following their stable integration into the genome of pluripotent embryonal carcinoma stem cells. Transfected genes integrate into the genome as tandem arrays. Expression of reporter genes from these tandem arrays in embryonal carcinoma cells is inefficient probably because genes are subject to repeat-induced gene silencing. We found that expression of reporter genes was significantly enhanced if co-transfected with cloned fragments derived from the murine Pgk-1 gene. The enhanced expression required (a) that the Pgk-1 fragment carries an active promoter, (b) that the promoter drives transcription through a region of more than 12 kbp, and (c) that this transcribed region contains both introns and exons. Reporter gene activity did not require specific Pgk-1 DNA sequences suggesting that the coupled processes of transcription and RNA processing conferred activity on neighboring genes probably by influencing local chromatin structure. Consistent with this idea, the effect of the Pgk-1 gene could be mimicked by exposing cells to butyrate or trichostatin A, inhibitors of histone deacetylase. Thus, the effect of the co-transfected Pgk-1 gene is to inhibit the process of gene inactivation possibly by functioning like an insulator or boundary element in the chromatin.
Ottawa Regional Cancer Center, University of Ottawa, Canada. michael_mcburney@cancercare.on.ca
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McBurney
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Yang
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Jardine
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Cormier
D000818:Animals; D002087:Butyrates / Q000494:pharmacology; D005091:Exons / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic; D056572:Histone Deacetylase Inhibitors; D006655:Histone Deacetylases / Q000378:metabolism; D007438:Introns / Q000235:genetics; D007763:Lac Operon / Q000235:genetics; D051379:Mice; D010735:Phosphoglycerate Kinase / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D012323:RNA Processing, Post-Transcriptional / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D011995:Recombination, Genetic; D012091:Repetitive Sequences, Nucleic Acid; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D001616:beta-Galactosidase / Q000187:drug effects / Q000235:genetics / Q000378:metabolism
D002087:Butyrates; D056572:Histone Deacetylase Inhibitors; D011993:Recombinant Fusion Proteins; D010735:Phosphoglycerate Kinase; D001616:beta-Galactosidase; D006655:Histone Deacetylases