Analysis of the mutation to forskolin-resistance in Y1 adrenocortical tumor cells.
Forskolin reduced the plating efficiency of Y1 adrenocortical tumor cells in a concentration-dependent manner--more than 5-orders of magnitude at 10 uM forskolin and at least 6-orders of magnitude at 50 uM forskolin. This effect was related to the diterpene's ability to increase adenylate cyclase activity and adenosine 3',5'-monophosphate (cAMP) levels in Y1 cells. Stable, forskolin-resistant mutants were isolated following growth of Y1 cells for 3 to 4 weeks in the presence of 10 uM forskolin. These mutants were stable, were present in the population at a ratio of approximately 15 mutants per million cells and appeared to result from a defect in cAMP accumulation rather than cAMP action. The forskolin-resistant phenotype was associated with a reduced ability of forskolin to stimulate adenylate cyclase activity in intact cells and in cell homogenates. The adenylate cyclase system of forskolin-resistant mutants was responsive to NaF, but was virtually insensitive to corticotropin (ACTH). As determined by a modified fluctuation analysis, the forskolin-resistant phenotype arose by spontaneous mutation at a frequency consistent with a mutational event at a single genetic locus (2 mutants per million cells per generation). These results indicate that the mutation which rendered Y1 cells insensitive to ACTH likely was the same as that which led to forskolin-resistance. Furthermore, the mutation seemed to behave dominantly. Although the gene product altered by the mutation is unknown, it does not appear to be the catalytic subunit of the enzyme.
D000262:Adenylyl Cyclases / Q000378:metabolism; D000306:Adrenal Cortex Neoplasms / Q000235:genetics* / Q000378:metabolism; D000324:Adrenocorticotropic Hormone / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D005576:Colforsin; D000242:Cyclic AMP / Q000378:metabolism; D004224:Diterpenes / Q000494:pharmacology*; D004351:Drug Resistance; D051379:Mice; D009154:Mutation
D004224:Diterpenes; D005576:Colforsin; D000324:Adrenocorticotropic Hormone; D000242:Cyclic AMP; D000262:Adenylyl Cyclases