Tyrosine kinase-independent inhibition of cyclic-AMP phosphodiesterase by genistein and tyrphostin 51.
The phosphodiesterase activity in the HT4.7 neural cell line was pharmacologically characterized, and phosphodiesterase isozyme 4 (PDE4) was found to be the predominant isozyme. The Km for cAMP was 1-2 microM, indicative of a "low Km" phosphodiesterase, and the activity was inhibited by PDE4-selective inhibitors rolipram and Ro20-1724, but not PDE3- or PDE2-selective inhibitors. Calcium, calmodulin, and cGMP, regulators of PDE1, PDE2, and PDE3, had no effect on cAMP hydrolysis. The protein tyrosine kinase inhibitor, genistein, inhibited HT4.7 cAMP phosphodiesterase activity by 85-95% with an IC50 of 4 microM; whereas daidzein, an inactive structural analog of genistein, had little effect on phosphodiesterase activity. This is a common pharmacological criterion used to implicate the regulation by a tyrosine kinase. However, genistein still inhibited phosphodiesterase activity with a mixed pattern of inhibition even when ion-exchange chromatography was used to partially purify phosphodiesterase away from the tyrosine kinase activity. Moreover, tyrphostin 51, another tyrosine kinase inhibitor, was found to also inhibit partially purified phosphodiesterase activity noncompetitively. These data suggest that HT4.7 phosphodiesterase activity is dominated by PDE4 and can be regulated by genistein and tyrphostin 51 by a tyrosine kinase-independent mechanism.
Nichols
M R
MR
Department of Chemistry, Purdue University, West Lafayette, Indiana, USA.
Morimoto
B H
BH
D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000378:metabolism; D000818:Animals; D054677:Cyclic Nucleotide Phosphodiesterases, Type 1; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D019833:Genistein / Q000494:pharmacology*; D007700:Kinetics; D051379:Mice; D009447:Neuroblastoma; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D014407:Tumor Cells, Cultured; D020032:Tyrphostins / Q000494:pharmacology*
D004791:Enzyme Inhibitors; D020032:Tyrphostins; C118515:tyrphostin 51; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D054677:Cyclic Nucleotide Phosphodiesterases, Type 1