Modulation of cyclic AMP levels in a clonal neural cell line by inhibitors of tyrosine phosphorylation.
The convergence of tyrosine kinase and cyclic AMP (cAMP) signal transduction pathways was investigated in the HT4.7 neural cell line with inhibitors of tyrosine kinases and tyrosine phosphatases. The protein tyrosine kinase inhibitor genistein inhibited isoproterenol-stimulated cAMP production by 40-60% in whole cells, with no effect on basal cAMP levels. In both whole cells and membranes, genistein also inhibited cAMP produced in response to direct stimulation of adenylyl cyclase with forskolin. However, in the absence of phosphodiesterase inhibitors, genistein presentation resulted in an increase in cAMP levels. Genistein inhibited phosphodiesterase activity by 80-85%, indicating that tyrosine phosphorylation stimulates both cAMP synthesis and degradation. The decrease in cAMP levels by genistein was not merely competitive inhibition of adenylyl cyclase with respect to ATP, since the Km of adenylyl cyclase for ATP remained essentially the same in either the presence or the absence of genistein. Another tyrosine kinase inhibitor, herbimycin A, which inhibits by a different mechanism than genistein, also decreased forskolin-stimulated cAMP in whole cells. As would be expected for the involvement of tyrosine phosphorylation in the control of cAMP production, inhibition of tyrosine phosphatases by vandate increased forskolin-stimulated cAMP production. These results suggest that cAMP production can be regulated by tyrosine phosphorylation, and the simultaneous activation of both cAMP synthesis and degradation may serve to alter the duration of cAMP elevation.
Stringfield
T M
TM
Department of Chemistry, Purdue University, West Lafayette, IN 47907-1393, U.S.A.
Morimoto
B H
BH
D000818:Animals; D016227:Benzoquinones; D002460:Cell Line; D003198:Computer Simulation; D000242:Cyclic AMP / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D019833:Genistein; D007529:Isoflavones / Q000494:pharmacology; D047029:Lactams, Macrocyclic; D009474:Neurons / Q000187:drug effects*; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011809:Quinones / Q000494:pharmacology; D017828:Rifabutin / Q000031:analogs & derivatives; D014638:Vanadates / Q000494:pharmacology
D016227:Benzoquinones; D004791:Enzyme Inhibitors; D007529:Isoflavones; D047029:Lactams, Macrocyclic; D010726:Phosphodiesterase Inhibitors; D011809:Quinones; D017828:Rifabutin; D014638:Vanadates; C020754:herbimycin; D019833:Genistein; D000242:Cyclic AMP; D011505:Protein-Tyrosine Kinases