Human chorionic gonadotropin (CG)- and phorbol ester-stimulated phosphorylation of the luteinizing hormone/CG receptor maps to serines 635, 639, 649, and 652 in the C-terminal cytoplasmic tail.
In a number of instances, binding of a ligand to its receptor results in receptor phosphorylation that mediates receptor uncoupling from its effector. Recently, we showed that human CG (hCG)- or phorbol ester- [phorbol 12-myristate-13-acetate (PMA)] stimulation of cells transfected with the LH/CG receptor induced rapid LH/CG receptor phosphorylation and a reduced cAMP response upon reexposure to hCG. The fact that hCG and PMA both phosphorylate and uncouple the LH/CG receptor suggests a common mechanism of action, namely the activation of protein kinase C. The studies presented here were designed to investigate the role of the C kinase in LH/CG receptor phosphorylation and to locate the phosphorylation site(s) within the receptor protein. The experiments presented here show that although hCG activates the C kinase in these cells, phosphorylation of the LH/CG receptor in response to hCG is maintained in C kinase-deficient cells. This suggests that activation of protein kinase C is not required for hCG-induced phosphorylation of its receptor. As a first step in locating the phosphorylation sites within the receptor polypeptide, we performed phosphoamino acid analysis of the phosphorylated LH/CG receptor. Only phosphoserine residues were detected. Based on the assumption that the phosphoserine(s) must be located within the intracellular regions of the receptor, we isolated cell lines expressing the wild type LH/CG receptor or receptors with cytoplasmic tails truncated at residue 653 or 631.(ABSTRACT TRUNCATED AT 250 WORDS)
Hipkin
R W
RW
Department of Pharmacology, University of Iowa College of Medicine, Iowa City 52242-1109, USA.
Wang
Z
Z
Ascoli
M
M
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D006063:Chorionic Gonadotropin / Q000494:pharmacology*; D000242:Cyclic AMP / Q000096:biosynthesis; D015536:Down-Regulation; D006801:Humans; D007295:Inositol Phosphates / Q000096:biosynthesis; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011493:Protein Kinase C / Q000502:physiology; D051381:Rats; D011974:Receptors, LH / Q000378:metabolism*; D012694:Serine / Q000378:metabolism*; D013268:Stimulation, Chemical; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014162:Transfection
D006063:Chorionic Gonadotropin; D007295:Inositol Phosphates; D011974:Receptors, LH; D012694:Serine; D000242:Cyclic AMP; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate