De-an Wang

Sphingosylphosphocholine is a naturally occurring lipid mediator in blood plasma: a possible role in regulating cardiac function via sphingolipid receptors.

Blood plasma and serum contain factors that activate inwardly rectifying GIRK1/GIRK4 K+ channels in atrial myocytes via one or more non-atropine-sensitive receptors coupled to pertussis-toxin-sensitive G-proteins. This channel is also the target of muscarinic M(2) receptors activated by the physiological release of acetylcholine from parasympathetic nerve endings. By using a combination of HPLC and TLC techniques with matrix-assisted laser desorption ionization-time-of-flight MS, we purified and identified sphingosine 1-phosphate (SPP) and sphingosylphosphocholine (SPC) as the plasma and serum factors responsible for activating the inwardly rectifying K+ channel (I(K)). With the use of MS the concentration of SPC was estimated at 50 nM in plasma and 130 nM in serum; those concentrations exceeded the 1.5 nM EC(50) measured in guinea-pig atrial myocytes. With the use of reverse-transcriptase-mediated PCR and/or Western blot analysis, we detected Edg1, Edg3, Edg5 and Edg8 as well as OGR1 sphingolipid receptor transcripts and/or proteins. In perfused guinea-pig hearts, SPC exerted a negative chronotropic effect with a threshold concentration of 1 microM. SPC was completely removed after perfusion through the coronary circulation at a concentration of 10 microM. On the basis of their constitutive presence in plasma, the expression of specific receptors, and a mechanism of ligand inactivation, we propose that SPP and SPC might have a physiologically relevant role in the regulation of the heart.

Molecular basis for lysophosphatidic acid receptor antagonist selectivity

Recent characterization of lysophosphatidic acid (LPA) receptors has made possible studies elucidating the structure-activity relationships (SAR) for agonist activity at individual receptors. Additionally, the availability of these receptors has allowed the identification of antagonists of LPA-induced effects. Two receptor-subtype selective LPA receptor antagonists, one selective for the LPA1/EDG2 receptor (a benzyl-4-oxybenzyl N-acyl ethanolamide phosphate, NAEPA, derivative) and the other selective for the LPA3/EDG7 receptor (diacylglycerol pyrophosphate, DGPP, 8:0), have recently been reported. The receptor SAR for both agonists and antagonists are reviewed, and the molecular basis for the difference between agonism and antagonism as well as for receptor-subtype antagonist selectivity identified by molecular modeling is described. The implications of the newly available receptor-subtype selective antagonists are also discussed.

Structural Features of EDG1 Receptor‐Ligand Complexes Revealed by Computational Modeling and Mutagenesis

ABBY L. PARRILL,a,b DANIEL L. BAKER,c DE-AN WANG,c DAVID J. FISCHER,c DEBRA L. BAUTISTA,b JAMES VAN BROCKLYN,d SARAH SPIEGEL,d AND GABOR TIGYIc bDepartment of Chemistry, University of Memphis, Memphis, Tennessee 38152, USA cDepartment of Physiology, University of Tennessee, Memphis, Tennessee 38163, USA dDepartment of Biochemistry and Molecular Biology, Georgetown University, Washington, District of Columbia 20007, USA

Inhibition of Ca2+ signalling by the sphingosine 1-phosphate receptor S1P1

The lysophospholipid, sphingosine 1-phosphate (S1P), regulates a multitude of cellular functions by activating specific G protein-coupled receptors (GPCRs) (S1P1–5, plus three newly identified S1P receptors). The Gi-coupled S1P1 receptor inhibits adenylyl cyclase, stimulates mitogen-activated protein kinases (MAP kinases) and cell migration, and is required for blood vessel maturation. Here, we report that S1P1 inhibits Ca2+ signalling in a number of cell types. In HEK-293 cells, which endogenously express S1P1–3, overexpression of S1P1 reduced intracellular free Ca2+ concentration ([Ca2+]i) increases induced by various receptor agonists as well as thapsigargin. The inhibitory Ca2+ signalling of S1P1 was blocked by pertussis toxin (PTX) and the protein kinase C (PKC) inhibitor, Go6976, and imitated by phorbol ester and overexpression of classical PKC isoforms. Activation of S1P1 stably expressed in RH7777 cells, which endogenously do not express S1P receptors, also inhibited Ca2+ signalling, without mediating Ca2+ mobilization on its own. It is concluded that the widely expressed S1P receptor S1P1 inhibits Ca2+ signalling, most likely via Gi proteins and classical PKC isoforms. Co-expression of S1P1 with S1P3, but not S1P2, reversed the inhibitory effect of S1P1, furthermore suggesting a specific interplay of S1P receptor subtypes usually found within a single cell type.