Phorbol ester abrogates up-regulation of DNA polymerase beta by DNA-alkylating agents in Chinese hamster ovary cells.
Mammalian DNA polymerase beta (beta-pol), a DNA repair polymerase, is known to be constitutively expressed in cultured cells, but treatment of cells with the DNA-alkylating agents MNNG or methyl methanesulfonate has been shown to up-regulate beta-pol mRNA level. To further characterize this response, we prepared a panel of monoclonal antibodies and used one of them to quantify beta-pol in whole cell extracts by immunoblotting. We found that treatment of Chinese hamster ovary cells with either DNA-alkylating agent up-regulated the beta-pol protein level 5-10-fold. This induction appeared to be secondary to DNA alkylation, as induction was not observed with a genetically altered cell line overexpressing the DNA repair enzyme O6-methylguanine-methyltransferase. We also found that 12-O-tetradecanoylphorbol-13-acetate (TPA) treatment of wild type Chinese hamster ovary cells increased expression of beta-pol protein (approximately 10-fold). Any interrelationship between this TPA response and the DNA-alkylation response was studied by treatment with combinations of MNNG and TPA. The beta-pol up-regulation observed with MNNG treatment was abrogated by TPA, and conversely the up-regulation observed with TPA treatment was abrogated by MNNG.
Srivastava
D K
DK
Sealy Center for Molecular Science, University of Texas Medical Branch, Galveston 77555-1068, USA.
Rawson
T Y
TY
Showalter
S D
SD
Wilson
S H
SH
D000477:Alkylating Agents / Q000494:pharmacology*; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D016466:CHO Cells; D006224:Cricetinae; D004249:DNA Damage; D004256:DNA Polymerase I / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D004260:DNA Repair; D004336:Drug Antagonism; D018604:Epitope Mapping; D015151:Immunoblotting; D008741:Methyl Methanesulfonate / Q000494:pharmacology; D008769:Methylnitronitrosoguanidine / Q000494:pharmacology; D008969:Molecular Sequence Data; D051381:Rats; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D015854:Up-Regulation / Q000187:drug effects*
D000477:Alkylating Agents; D000911:Antibodies, Monoclonal; D008769:Methylnitronitrosoguanidine; D008741:Methyl Methanesulfonate; D004256:DNA Polymerase I; D013755:Tetradecanoylphorbol Acetate