Regulation of ERK1/2 activity by ghrelin-activated growth hormone secretagogue receptor 1A involves a PLC/PKCvarepsilon pathway.
1. The growth hormone secretagogue receptor 1a (GHSR-1a) is a G-protein coupled receptor, involved in the biological actions of ghrelin by triggering inositol phosphates and calcium intracellular second messengers. It has also been reported that ghrelin could activate the 44- and 42-kDa extracellular signal-regulated protein kinases (ERK1/2) in different cell lines, but it is not clear whether this regulation is GHSR-1a dependent or not. 2. To provide direct evidence for the coupling of GHSR-1a to ERK1/2 activation, this pathway has been studied in a heterologous expression system. 3. Thus, in Chinese hamster ovary (CHO) cells we showed that ghrelin induced, via the human GHSR-1a, a transient and dose-dependent activation of ERK1/2 leading to activation of the transcriptional factor Elk1. 4. We then investigated the precise mechanisms involved in GHSR-1a-mediated ERK1/2 activation using various specific inhibitors and dominant-negative mutants and found that internalization of GHSR-1a was not necessary. Our results also indicate that phospholipase C (PLC) was involved in GHSR-1a-mediated ERK1/2 activation, however, pathways like tyrosine kinases, including Src, and phosphoinositide 3-kinases were not found to be involved. GHSR-1a-mediated ERK1/2 activation was abolished both by a general protein kinase C (PKC) inhibitor, Gö6983, and by PKC depletion using overnight pretreatment with phorbol ester. Moreover, the calcium chelator, BAPTA-AM, and the inhibitor of conventional PKCs, Gö6976, had no effect on the GHSR-1a-mediated ERK1/2 activation, suggesting the involvement of novel PKC isoforms (epsilon, delta), but not conventional or atypical PKCs. Further analyses suggest that PKCepsilon is required for the activation of ERK1/2. 5. Taken together, these data suggest that ghrelin, through GHSR-1a, activates the Elk1 transcriptional factor and ERK1/2 by a PLC- and PKCepsilon-dependent pathway.
Mousseaux
Delphine
D
Laboratoire des Aminoacides, Peptides et Protéines (LAPP), CNRS UMR-5810, UMI et UMII, UFR Pharmacie, 15 avenue Charles Flahault, Montpellier 34093, Cedex 5, France.
Le Gallic
Lionel
L
Ryan
Joanne
J
Oiry
Catherine
C
Gagne
Didier
D
Fehrentz
Jean-Alain
JA
Galleyrand
Jean-Claude
JC
Martinez
Jean
J
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D054439:Ghrelin; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D036361:Peptide Hormones / Q000502:physiology*; D051744:Protein Kinase C-epsilon / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D054440:Receptors, Ghrelin; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism; D050807:ets-Domain Protein Elk-1 / Q000378:metabolism
D054439:Ghrelin; D036361:Peptide Hormones; D043562:Receptors, G-Protein-Coupled; D054440:Receptors, Ghrelin; D050807:ets-Domain Protein Elk-1; D051744:Protein Kinase C-epsilon; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D010738:Type C Phospholipases