Lack of constitutive activation or inactivation of the platelet-activating factor receptor by glutamate substitution of alanine 230

S A Carlson; T K Chatterjee; R A Fisher

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Lack of constitutive activation or inactivation of the platelet-activating factor receptor by glutamate substitution of alanine 230

Journal article

Peer reviewed

Lack of constitutive activation or inactivation of the platelet-activating factor receptor by glutamate substitution of alanine 230

S A Carlson, T K Chatterjee and R A Fisher

Receptors & signal transduction, Vol.6(2), pp.111-120

1996

PMID: 9015866

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Abstract

The platelet-activating factor (PAF) receptor (PAFR) is a G protein-coupled receptor (GPCR) that mediates a diverse array of biological responses to PAF. Recently, we provided evidence that the third intracellular domain (3i) of the rat PAFR (rPAFR) is a critical determinant in its coupling to phosphoinositide phospholipase C (PI PLC)-activating G proteins. In the present study, we assessed the potential role of a conserved alanine in the carboxyl-terminal region of 3i of the rPAFR in rPAFR signaling activity. Previous studies with the m5 muscarinic acetylcholine and human PAF receptors revealed that substitution of a carboxyl-terminal alanine was found to impair receptor-mediated PI PLC activation. Here we report the effects of the analogous nonconservative substitution of glutamate for alanine 230 of the rPAFR (rPAFR A230E) on receptor-mediated agonist binding and PI PLC activation following transient expression of the receptor cDNA. BHK cells transfected with a cDNA encoding the rPAFR A230E exhibited PAF-stimulated increases in inositol phosphate (IP) accumulation with no increase in basal levels of IPs. PAF-stimulated IP production in rPAFR transfectants was dependent on the amount of DNA transfected, although PAF provoked a larger increase in IPs in rPAFR transfectants than in rPAFRA230E transfectants in cells transfected with equal amounts of receptor cDNA. This latter finding apparently reflects differences in the transfection efficiency or expression of the wild-type and rPAFR A230E cDNAs because PAF produced indistinguishable effects on IP accumulation in rPAFR and rPAFR A230E transfectants expressing equivalent numbers of receptors. These results provide evidence for a nonconserved role of this conserved alanine in coupling of group I GPCRs to PI PLC-activating G proteins and also suggest that this residue has differential roles in regulating expression and signaling by rat and human PAFRs.

Recombinant Proteins - metabolism

Amino Acid Sequence

Mutagenesis, Site-Directed

Signal Transduction

Glutamic Acid - genetics

Receptors, G-Protein-Coupled

Rats

Platelet Membrane Glycoproteins - genetics

Receptors, Cell Surface

Phosphatidylinositol Diacylglycerol-Lyase

Phosphoinositide Phospholipase C

Dose-Response Relationship, Drug

Phosphoric Diester Hydrolases

Platelet Activating Factor - metabolism

Animals

Alanine - genetics

Platelet Membrane Glycoproteins - metabolism

Conserved Sequence

Enzyme Activation

Mutation

Inositol Phosphates - metabolism

GTP-Binding Proteins - metabolism

Details

Title: Subtitle: Lack of constitutive activation or inactivation of the platelet-activating factor receptor by glutamate substitution of alanine 230
Creators: S A Carlson - Department of Pharmacology, University of Iowa College of Medicine, Iowa City 52242, USA
T K Chatterjee
R A Fisher
Resource Type: Journal article
Publication Details: Receptors & signal transduction, Vol.6(2), pp.111-120
Publisher: United States
PMID: 9015866
ISSN: 1087-8475
Grant note: DK25295 / NIDDK NIH HHS HL41071 / NHLBI NIH HHS
Language: English
Date published: 1996
Academic Unit: Iowa Neuroscience Institute; Neuroscience and Pharmacology; Internal Medicine
Record Identifier: 9984040363602771

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