NDLI: Amplification of the UMP synthase gene and enzyme overproduction in pyrazofurin-resistant rat hepatoma cells. Molecular cloning of a cDNA for UMP synthase.

Content Provider	Journal of Biological Chemistry (JBC)
Author	Kanalas, J. J. Suttle, D. P.
Abstract	The levels of UMP synthase protein and mRNA are increased in rat hepatoma cells that have acquired resistance to pyrazofurin, a potent inhibitor of pyrimidine biosynthesis. A cDNA plasmid library was prepared from partially purified poly(A)+ mRNA isolated from the resistant cell line. Recombinant plasmids with inserts complementary to UMP synthase mRNA were selected by differential hybridization with cDNA prepared from wild type and resistant cell mRNA and analysis of hybrid-selected mRNA by in vitro translation reactions. One plasmid, pUMPS-2, contains a 850-base pair insert and was used to analyze UMP synthase gene sequences in the wild type and resistant cell lines. Blot hybridization of restricted genomic DNA demonstrated amplification of the UMP synthase gene in the resistant cells. The number of UMP synthase genes is increased 15-fold as determined by a modified dot hybridization procedure. Previous studies have shown that the resistant cells have a 16-fold increase in UMP synthase mRNA but a 40-fold increase in synthase activity (Suttle, D.P. (1983) J. Biol. Chem. 258, 7707-7713). To further investigate this discrepancy between the amount of increase in DNA and mRNA versus the increase in enzyme activity, we have determined the relative rate of synthesis and degradation of UMP synthase. The rate of synthesis was 13-fold faster in the resistant cells. The degradation rate was not significantly different between the two cell lines. These data indicate that gene amplification is the major factor contributing to the enzyme overproduction in the pyrazofurin-resistant cells.
Related Links	http://www.jbc.org/content/259/3/1848.abstract
Ending Page	1853
Starting Page	1848
Page Count	6
File Format	HTM / HTML PDF
ISSN	00219258
Journal	Journal of Biological Chemistry (JBC)
Issue Number	3
Volume Number	259
e-ISSN	1083351X
Language	English
Publisher	American Society for Biochemistry and Molecular Biology
Publisher Date	1984-02-10
Access Restriction	Open
Content Type	Text
Resource Type	Article
Subject	Cell Biology Biochemistry Molecular Biology

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