Regulation of the ovalbumin gene: effects of insulin, adenosine 3',5'-monophosphate, and estrogen.
Induction of the ovalbumin gene in chicken oviduct explant cultures requires the presence of a serum component in addition to estrogen. Previous studies have demonstrated that either insulin or related peptides, such as proinsulin or insulin-like growth factor, can eliminate the requirement for serum. In the present study, we determined that half-maximal binding of [125I]iodoinsulin to oviduct cell membranes occurs with 5 X 10(-10) M insulin, and proinsulin is 10-fold less effective than insulin as a competitor. Induction of the ovalbumin gene is half-maximal with 10(-9) M insulin, and both proinsulin and insulin-like growth factor are 10-fold less potent. These results suggest that insulin is the physiological hormone that is required in addition to estrogen to stimulate transcription of the ovalbumin gene. In an effort to understand the mechanism by which insulin regulates the steroid response in this system, we searched for agents that could substitute for insulin. We found that cyclic nucleotide derivatives, such as 8-bromo-cAMP, can mimic the effect of insulin on ovalbumin gene expression and that the phosphodiesterase inhibitor isobutylmethylxanthine can potentiate this response to 8-bromo-cAMP. Activation of oviduct adenylate cyclase with either forskolin or cholera toxin also allows the estrogen-mediated induction of mRNA, and both of these agents produce a dramatic rise in intracellular cAMP. Although these results suggest that the effect of insulin on gene regulation might depend on a cAMP-mediated mechanism, we are unable to detect any change in cellular cAMP levels in response to insulin. We conclude that insulin and cAMP are producing their effects on activation of the ovalbumin gene via convergent pathways. The involvement of an activator of protein kinase, cAMP, strongly suggests that protein phosphorylation events play a regulatory role in the expression of the ovalbumin gene.
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000818:Animals; D002645:Chickens; D002772:Cholera Toxin / Q000494:pharmacology; D005576:Colforsin; D046508:Culture Techniques; D000242:Cyclic AMP / Q000494:pharmacology*; D004224:Diterpenes / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004958:Estradiol / Q000494:pharmacology*; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects*; D007328:Insulin / Q000378:metabolism / Q000494:pharmacology*; D007335:Insulin-Like Growth Factor II; D010047:Ovalbumin / Q000235:genetics*; D010057:Oviducts / Q000378:metabolism; D010455:Peptides / Q000494:pharmacology; D011384:Proinsulin / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis; D011972:Receptor, Insulin / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects
D004224:Diterpenes; D007328:Insulin; D010455:Peptides; D012333:RNA, Messenger; D005576:Colforsin; D015124:8-Bromo Cyclic Adenosine Monophosphate; D004958:Estradiol; D007335:Insulin-Like Growth Factor II; D010047:Ovalbumin; D002772:Cholera Toxin; D011384:Proinsulin; D000242:Cyclic AMP; D011972:Receptor, Insulin