Norepinephrine- and phorbol ester-induced phosphorylation of alpha(1a)-adrenergic receptors. Functional aspects.
Maximal adrenergic responses in Rat-1 fibroblasts expressing alpha(1a)-adrenergic receptors are not blocked by activation of protein kinase C. In contrast, activation of protein kinase C induces the phosphorylation of alpha(1b)-adrenoreceptors and blocks their actions. The effect of norepinephrine and phorbol esters on alpha(1a)-adrenoreceptor phosphorylation and coupling to G proteins were studied. Both stimuli lead to dose-dependent receptor phosphorylation. Interestingly, protein kinase C activation affected to a much lesser extent the actions of alpha(1a)-adrenergic receptors than those of the alpha(1b) subtype (norepinephrine elicited increases in calcium in whole cells and [(35)S]GTPgammaS binding to membranes). Basal phosphorylation of alpha(1a)-adrenergic receptors was much less than that observed with the alpha(1b) subtype. The carboxyl terminus seems to be the main domain for receptor phosphorylation. Therefore, chimeric receptors, where the carboxyl-terminal tails of alpha(1a) and alpha(1b) adrenergic receptors were exchanged, were constructed and expressed. alpha(1a)-Adrenoreceptors wearing the carboxyl tail of the alpha(1b) subtype had a high basal phosphorylation and displayed a strong phosphorylation in response to norepinephrine and phorbol esters. Our results demonstrate that stimulation of alpha(1a)-adrenergic receptor, or activation of protein kinase C, leads to alpha(1a)-adrenergic receptor phosphorylation. alpha(1a)-Adrenoreceptors are affected to a much lesser extent than alpha(1b)-adrenoreceptors by protein kinase C activation.
Vázquez-Prado
J
J
Departamento de Biología Celular, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Apartado Postal 70-248, México City 04510, México.
Medina
L C
LC
Romero-Avila
M T
MT
González-Espinosa
C
C
García-Sáinz
J A
JA
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D007700:Kinetics; D009638:Norepinephrine / Q000494:pharmacology*; D010766:Phosphorylation; D019920:Photoaffinity Labels; D011233:Precipitin Tests; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D018340:Receptors, Adrenergic, alpha-1 / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
C547715:Adra1a protein, rat; C547718:Adra1b protein, rat; D004791:Enzyme Inhibitors; D019920:Photoaffinity Labels; D018340:Receptors, Adrenergic, alpha-1; D011993:Recombinant Fusion Proteins; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; D009638:Norepinephrine