Arrestin-mediated ERK activation by gonadotropin-releasing hormone receptors - Receptor-specific activation mechanisms and compartmentalization

Christopher J Caunt, A R Finch, K R Sedgley, L Oakley, L M Luttrell, C A McArdle

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Abstract

Activation of seven-transmembrane region receptors typically causes their phosphorylation with consequent arrestin binding and desensitization. Arrestins also act as scaffolds, mediating signaling to Raf and ERK and, for some receptors, inhibiting nuclear translocation of ERK. GnRH receptors (GnRHRs) act via G(q/11) to stimulate the phospholipase C/Ca2+/protein kinase C (PKC) cascade and the Raf/MEK/ERK cassette. Uniquely, type I mammalian GnRHRs lack the C-tails that are found in other seven-\transmembrane region receptors ( including nonmammalian GnRHRs) and are implicated in arrestin binding. Here we have compared ERK signaling by human GnRHRs (hGnRHRs) and Xenopus GnRHRs (XGnRHRs). In HeLa cells, XGnRHRs underwent rapid and arrestin-dependent internalization and caused arrestin/green fluorescent protein (GFP) translocation to the membrane and endosomes, whereas hGnRHRs did not. Internalized XGnRHRs were co-localized with arrestin-GFP, whereas hGnRHRs were not. Both receptors mediated transient ERK phosphorylation and nuclear translocation (revealed by immunohistochemistry or by imaging of co-transfected ERK2-GFP), and for both, ERK phosphorylation was reduced by PKC inhibition but not by inhibiting epidermal growth factor receptor autophosphorylation. In the presence of PKC inhibitor, Delta arrestin-(319 - 418) blocked XGnRHR-mediated, but not hGnRHR- mediated, ERK phosphorylation. When receptor number was varied, hGnRHRs activated phospholipase C and ERK more efficiently than XGnRHRs but were less efficient at causing ERK2-GFP translocation. At high receptor number, XGnRHRs and hGnRHRs both caused ERK2-GFP translocation to the nucleus, but at low receptor number, XGnRHRs caused ERK2-GFP translocation, whereas hGnRHRs did not. Thus, experiments with XGnRHRs have revealed the first direct evidence of arrestin-mediated (probably G protein-independent) GnRHR signaling, whereas those with hGnRHRs imply that scaffolds other than arrestins can determine GnRHR effects on ERK compartmentalization.
LanguageEnglish
Pages2701-2710
Number of pages10
JournalJournal of Biological Chemistry
Volume281
Issue number5
DOIs
StatusPublished - 2006

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LHRH Receptors
Arrestin
Xenopus
Green Fluorescent Proteins
Chemical activation
Phosphorylation
Protein Transport
Arrestins
Protein Kinase C
Type C Phospholipases
Scaffolds
Protein Kinase C-delta
Mitogen-Activated Protein Kinase Kinases
Protein C Inhibitor
Endosomes
GTP-Binding Proteins
Epidermal Growth Factor Receptor
Protein Kinase Inhibitors
Cytoplasmic and Nuclear Receptors
HeLa Cells

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Arrestin-mediated ERK activation by gonadotropin-releasing hormone receptors - Receptor-specific activation mechanisms and compartmentalization. / Caunt, Christopher J; Finch, A R; Sedgley, K R; Oakley, L; Luttrell, L M; McArdle, C A.

In: Journal of Biological Chemistry, Vol. 281, No. 5, 2006, p. 2701-2710.

Research output: Contribution to journalArticle

Caunt, Christopher J ; Finch, A R ; Sedgley, K R ; Oakley, L ; Luttrell, L M ; McArdle, C A. / Arrestin-mediated ERK activation by gonadotropin-releasing hormone receptors - Receptor-specific activation mechanisms and compartmentalization. In: Journal of Biological Chemistry. 2006 ; Vol. 281, No. 5. pp. 2701-2710
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AU - Caunt,Christopher J

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AU - Oakley,L

AU - Luttrell,L M

AU - McArdle,C A

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N2 - Activation of seven-transmembrane region receptors typically causes their phosphorylation with consequent arrestin binding and desensitization. Arrestins also act as scaffolds, mediating signaling to Raf and ERK and, for some receptors, inhibiting nuclear translocation of ERK. GnRH receptors (GnRHRs) act via G(q/11) to stimulate the phospholipase C/Ca2+/protein kinase C (PKC) cascade and the Raf/MEK/ERK cassette. Uniquely, type I mammalian GnRHRs lack the C-tails that are found in other seven-\transmembrane region receptors ( including nonmammalian GnRHRs) and are implicated in arrestin binding. Here we have compared ERK signaling by human GnRHRs (hGnRHRs) and Xenopus GnRHRs (XGnRHRs). In HeLa cells, XGnRHRs underwent rapid and arrestin-dependent internalization and caused arrestin/green fluorescent protein (GFP) translocation to the membrane and endosomes, whereas hGnRHRs did not. Internalized XGnRHRs were co-localized with arrestin-GFP, whereas hGnRHRs were not. Both receptors mediated transient ERK phosphorylation and nuclear translocation (revealed by immunohistochemistry or by imaging of co-transfected ERK2-GFP), and for both, ERK phosphorylation was reduced by PKC inhibition but not by inhibiting epidermal growth factor receptor autophosphorylation. In the presence of PKC inhibitor, Delta arrestin-(319 - 418) blocked XGnRHR-mediated, but not hGnRHR- mediated, ERK phosphorylation. When receptor number was varied, hGnRHRs activated phospholipase C and ERK more efficiently than XGnRHRs but were less efficient at causing ERK2-GFP translocation. At high receptor number, XGnRHRs and hGnRHRs both caused ERK2-GFP translocation to the nucleus, but at low receptor number, XGnRHRs caused ERK2-GFP translocation, whereas hGnRHRs did not. Thus, experiments with XGnRHRs have revealed the first direct evidence of arrestin-mediated (probably G protein-independent) GnRHR signaling, whereas those with hGnRHRs imply that scaffolds other than arrestins can determine GnRHR effects on ERK compartmentalization.

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