Introduction and expression of the bacterial PaeR7 methylase gene in mammalian cells

Kwoh, T. J., Kwoh, D. Y., McCue, A. W., Davis, G. R., Patrick, D., Gingeras, T. R. (1986) Introduction and expression of the bacterial PaeR7 methylase gene in mammalian cells. Proc Natl Acad Sci U S A, 83 (20). pp. 7713-7. ISSN 0027-8424 (Print)0027-8424 (Linking)

URL: http://www.ncbi.nlm.nih.gov/pubmed/3463994

Abstract

An approach is devised for studying the role of DNA methylation in eukaryotic gene expression. The approach is based on the expression of site-specific bacterial methylase genes in animal cells. A model system using the cloned PaeR7 (an isoschizomer of Xho I) methylase gene was constructed to test the feasibility of this approach. Expression plasmids for the PaeR7 methylase gene were introduced into mouse Ltk- cells by cotransfection with the cloned chicken thymidine kinase (tk) gene. Several of the cell strains derived from Tk+ colonies were found to express the PaeR7 gene as judged by four criteria: the cellular DNA of these strains showed increased resistance to cleavage by Xho I; these strains contained cellular proteins that comigrated with pure PaeR7 methylase protein, as visualized by immunoblotting; PaeR7 methylase activity was found in vitro in crude extracts of total cellular protein from these strains; and murine adenovirus genomes grown on cells expressing PaeR7 methylase showed resistance to cleavage to PaeR7 endonuclease. The potential applications of this approach for the study of cellular and viral gene regulation, DNA repair, and restriction modification are discussed.

Item Type: Paper
Uncontrolled Keywords: Animals Cell Line DNA metabolism DNA Cytosine 5 Methyltransferase analysis genetics DNA Repair DNA Viral metabolism Genes Bacterial Methylation Mice Nucleic Acid Hybridization Plasmids Transfection
Subjects: bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > DNA methylation
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > genes, structure and function > gene expression
CSHL Authors:
Communities: CSHL labs > Gingeras lab
Depositing User: CSHL Librarian
Date Deposited: 14 Mar 2012 14:02
Last Modified: 31 Mar 2014 13:52
PMCID: PMC386791
Related URLs:
URI: http://repository.cshl.edu/id/eprint/25237

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