TSH action on cAMP binding to the regulatory subunits of cAMP-dependent protein kinases in pig thyroid cell cultures.
This study examines the mechanism of TSH action on the cAMP-dependent protein kinases (PKA) by measuring the catalytic activity of the two PKA isozymes (PKA I and PKA II) and their capacity to bind cAMP to the regulatory subunits (RI and RII) in thyroid cell cultures exposed for two days to different doses of TSH. In TSH-treated cell cultures a selective down regulation (up to 60%) of the catalytic activity was found; the PKA I was down regulated at lower TSH doses (0.1 mU/ml and even 0.05 mU/ml) than was the PKA II (1.0 mU/ml TSH). At the dose of 1.0 mU/ml the loss of the catalytic activity in PKA I and PKA II was respectively 60% and 40%. No free catalytic activity was found either in control or in TSH-treated cells. Binding of cAMP to regulatory subunits (R) measured under exchange conditions at 37 degrees C, showed that no change in total regulatory subunit protein content occurs in TSH-treated cells. Binding of cAMP to R subunits at 4 degrees C (when only free cAMP binding sites are measured) revealed an important endogenous occupancy of cAMP binding sites of RI and RII isoreceptors under basal conditions (40%) and a significantly increased occupancy after exposure of cells to TSH (60%). Pools of regulatory subunits with more than 50% of sites occupied, which were devoid of enzyme activity, were found both, in control and TSH-exposed cells. They were identified as RI subunits which represented a mixed population of native and partly degraded molecules.(ABSTRACT TRUNCATED AT 250 WORDS)
Ben Abdelkhalek
M
M
Laboratoire de Physiologie Animale, Faculté des Sciences, Rabat, Marocco.
Breton
M F
MF
Feliers
D
D
Haye
B
B
Pavlovic-Hournac
M
M
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000495:Allosteric Site; D000818:Animals; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D007527:Isoenzymes / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D013552:Swine; D013961:Thyroid Gland / Q000378:metabolism*; D013972:Thyrotropin / Q000494:pharmacology*
D007527:Isoenzymes; D013972:Thyrotropin; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D015056:1-Methyl-3-isobutylxanthine