Over-expression of the truncated ghrelin receptor polypeptide attenuates the constitutive activation of phosphatidylinositol-specific phospholipase C by ghrelin receptors but has no effect on ghrelin-stimulated extracellular signal-regulated kinase 1/2 activity.
In addition to regulating growth hormone release from the pituitary, ghrelin receptors also influence cell proliferation and apoptosis. By studying mitogen-activated protein kinase activity in human embryonic kidney 293 cells over-expressing ghrelin receptors, we aimed to identify the specific cell signalling pathways used by ghrelin receptors, and to determine if the truncated ghrelin receptor polypeptide had any influence on the functional activity of ghrelin receptors. We found that ghrelin activated extracellular signal-regulated kinases 1/2 with an EC50 value of 10 nM, and that this response was inhibited by the ghrelin receptor antagonists D-Lys3-GHRP-6 and [D-Arg1,D-Phe5,D-Trp(7,9),Leu11]-substance P. Ghrelin had little or no effect on the activity of c-Jun N-terminal kinase, p38 kinase or Akt. Ghrelin appeared to activate extracellular signal-regulated kinases 1/2 through a calcium-independent novel protein kinase C isoform which may utilize diacylglycerol derived from hydrolysis of phosphatidylcholine rather than from phosphatidylinositol. Ghrelin-stimulated extracellular signal-regulated kinases 1/2 activity was independent of transactivation of epidermal growth factor receptors, and even when ghrelin receptor internalization was blocked by concanavalin A or a beta-arrestin mutant, there was no decrease in phosphorylated extracellular signal-regulated kinases 1/2, suggesting this is a G protein-dependent process. The truncated ghrelin receptor polypeptide had no effect on ghrelin receptor signalling to extracellular signal-regulated kinases 1/2, but decreased the constitutive activation of phosphatidylinositol-specific phospholipase C by ghrelin receptors. In conclusion, our results suggest that any up-regulation of the truncated ghrelin receptor polypeptide might preferentially attenuate functional activity dependent on the constitutive activation of ghrelin receptors, while leaving ghrelin-dependent signalling unaffected.
Chu
Kit-Man
KM
Department of Pharmacology, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong, SAR, China.
Chow
Kevin B S
KB
Leung
Po-Ki
PK
Lau
Pui-Ngan
PN
Chan
Chi-Bun
CB
Cheng
Christopher H K
CH
Wise
Helen
H
D015153:Blotting, Western; D002460:Cell Line; D003208:Concanavalin A / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004705:Endocytosis / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D015870:Gene Expression; D054439:Ghrelin; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D009842:Oligopeptides / Q000494:pharmacology; D036361:Peptide Hormones / Q000494:pharmacology*; D043265:Phosphatidylinositol Diacylglycerol-Lyase / Q000378:metabolism*; D054801:Phosphoinositide Phospholipase C; D010766:Phosphorylation / Q000187:drug effects; D020033:Protein Isoforms / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D054440:Receptors, Ghrelin; D013373:Substance P / Q000031:analogs & derivatives / Q000494:pharmacology; D014162:Transfection; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D054439:Ghrelin; D009842:Oligopeptides; D036361:Peptide Hormones; D020033:Protein Isoforms; D043562:Receptors, G-Protein-Coupled; D054440:Receptors, Ghrelin; D003208:Concanavalin A; D013373:Substance P; C041048:growth hormone releasing hexapeptide; C045800:substance P, Phe(5)-Trp(7,9)-Leu(11)-; D051057:Proto-Oncogene Proteins c-akt; D048031:JNK Mitogen-Activated Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D048051:p38 Mitogen-Activated Protein Kinases; D054801:Phosphoinositide Phospholipase C; D043265:Phosphatidylinositol Diacylglycerol-Lyase