Nucleic Acids Research, 1985, Vol. 13, No. 10 3599-3615
© 1985
Articles |
Minichromosome assembly of non-integrated plasmid DNA transfected into mammalian cells
Biochemistiy/Biophysics Program and Program in Genetics and Cell Biology, Washington State University Pullman, WA 99164-4350, USA 1Eukaryotic Molecular Biology Group, Cancer Research Campaign, Biochemistry, Imperial College of Science and Technology London 5W7 2AZ, UK 2Laboratory of Mo-lecular Biology Group, Cancer Research Institute, Division of Cancer Biology and Diagnosis, National Institutes of Health Bethesda, MD 20205, USA
Received September 24, 1984. Revised April 22, 1985. Accepted April 22, 1985.
The nucleoprotein structures formed on various plasmid expression vectors transfected into mammalian cells by both the calcium phosphate and DEAE-dextran methods have been studied. We demonstrate by a variety of means that mammalian cells are capable of rapidly assembling non-integrated circular plasmids (both replicating and non-replicating) into typical "minichromosomes" containing nucleosomes with a 190 bp repetitive spacing. Treatment of recipient cells with sodium butyrate for a short period of time (12–16 h) immediately following transfection markedly increased the DNase I digestion sensitivity of the newly assembled plasmid chromatin. Furthermore, minichromosomes isolated from such butyrate-treated cells are depleted in histone H1 and Contain highly acetylated forms of histone H4. These findings are entirely consistent with our earlier speculation (Gorman et al., Nucleic Acids Res. 11, 1044; 1983) that appropriate butyrate treatment might stimulate transient expression of newly transfected genes by facilitating their assembly into an "active" type of chromatin structure.
*Present Address: Genentech, Inc., 460 Point San Bruno Blvd., South San Francisco, CA 94080, USA
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