title: Regulation by a beta-adrenergic receptor of a Ca2+-independent adenosine 3',5'-(cyclic)monophosphate phosphodiesterase in C6 glioma cells.
abstract: The hormonal control of cyclic nucleotide phosphodiesterase (EC 3.1.4.17) activity has been studied by using as a model the isoproterenol stimulation of cyclic AMP phosphodiesterase activity in C6 glioma cells. A 2-fold increase in cyclic AMP phosphodiesterase specific activity was observed in homogenates of isoproterenol-treated cells relative to control. This increase reached a maximum 3 h after addition of isoproterenol, was selective for cyclic AMP hydrolysis, was reproduced by incubation with 8-Br cycl AMP but not with 8-Br cyclic GMP and was limited to the soluble enzyme activity. The presence of 0.1 mM EGTA did not alter the magnitude of the increase in phosphodiesterase activity. Moreover, the calmodulin content in the cell extracts was not changed after isoproterenol. DEAE-Sephacel chromatography of the 100000 X g supernatant resolved two peaks of phosphodiesterase activity. The first peak hydrolyzed both cyclic nucleotides and was activated by Ca2+ an purified calmodulin. The second peak was specific for cyclic AMP but it was Ca2+- and calmodulin-insensitive. Isoproterenol selectively increased the specific activity of the second peak. Kinetic analysis of the cyclic AMP hydrolysis by the induced enzyme revealed a non-linear Hofstee plot with apparent Km values of 2-5 microM. Cyclic GMP was not hydrolyzed by this enzyme in the absence or presence of calmodulin and failed to affect the kinetics of the hydrolysis of cyclic AMP. Gel filtration chromatography of the induced DEAE-Sephacel peak resolved a single peak of enzyme activity with an apparent molecular weight of 54000.
mesh_terms: D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000378:metabolism*; D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000378:metabolism; D015124:8-Bromo Cyclic Adenosine Monophosphate; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000494:pharmacology; D002460:Cell Line; D002852:Chromatography, Ion Exchange; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000494:pharmacology; D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000494:pharmacology; D005910:Glioma / Q000201:enzymology*; D007527:Isoenzymes / Q000302:isolation & purification; D007545:Isoproterenol / Q000494:pharmacology; D007700:Kinetics; D051381:Rats; D011941:Receptors, Adrenergic / Q000378:metabolism*; D011943:Receptors, Adrenergic, beta / Q000378:metabolism*; D013347:Subcellular Fractions / Q000201:enzymology
chemical_list: D002147:Calmodulin; D007527:Isoenzymes; D011941:Receptors, Adrenergic; D011943:Receptors, Adrenergic, beta; D015124:8-Bromo Cyclic Adenosine Monophosphate; C016276:8-bromocyclic GMP; D000242:Cyclic AMP; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D006152:Cyclic GMP; D007545:Isoproterenol; D002118:Calcium