Sphingolipid receptor signaling and function in human bladder carcinoma cells: inhibition of LPA- but enhancement of thrombin-stimulated cell motility

Abstract

Sphingosine-1-phosphate (SPP) induces a variety of cellular responses, including Ca2+ signaling, proliferation, and inhibition of motility, apparently by acting at specific G protein coupled receptors. Here, the expression, signaling, and motile responses of sphingolipid receptors were examined in human bladder carcinoma (J82) cells, for which lysophosphatidic acid (LPA) and thrombin act as potent agonists. SPP potently and rapidly mobilized Ca2+, stimulated phospholipases C and D, and inhibited cAMP accumulation, without affecting growth of J82 cells, which express the recently identified SPP receptors, Edg-1 and Edg-3. The effects of SPP were mimicked by sphingosylphosphorylcholine (SPPC) and strongly attenuated by pertussis toxin (PTX). SPP and SPPC by themselves induced a small, PTX-sensitive motile response. However, stimulation of cell motility by LPA, which by itself was also PTX-sensitive, was blocked by SPP and SPPC. In contrast, motility stimulation by thrombin, which by itself was PTX-insensitive, was strongly augmented by the sphingolipids in a PTX-sensitive manner. The bidirectional regulation of LPA- and thrombin-stimulated motility was not due to selective alterations in the activation of Rho GTPases which control cell motility. In fact, RhoA activation and Rho-dependent actin stress fiber formation induced by LPA and thrombin were mimicked, but not altered by SPP and SPPC. We conclude that J82 cells express sphingolipid receptors, coupled via G proteins to several signaling pathways. Most importantly, these sphingolipid receptors potently regulate thrombin- and LPA-stimulated motility, but in opposite directions, suggesting that migration of these human bladder carcinoma cells is controlled by a complex network of interacting extracellular ligands.

G-Protein-Coupled Receptors, Cell Motility and Proliferation

“…SPP inhibited J82 cell migration induced by LPA, which was virtually abolished in the presence of 1 M SPP. In complete contrast, stimulation of J82 cell migration induced by thrombin was strongly increased by SPP, resulting in almost doubling of the thrombin response in the presence of 1 mM SPP [25]. Thus SPP is not only a potent inhibitor of cell motility, as reported before by various other cell types [26][27][28][29][30] but, as demonstrated for J82 cells, this sphingolipid can also potently promote cell migration.…”

“…In PTXtreated cells, the stimulatory effect of SPP was completely abolished [25]. The most striking effect of the sphingolipid on J82 cell migration was observed when the weakly stimulatory agonist SPP was combined with the potent agonists, LPA and thrombin.…”

“…Thus SPP is not only a potent inhibitor of cell motility, as reported before by various other cell types [26][27][28][29][30] but, as demonstrated for J82 cells, this sphingolipid can also potently promote cell migration. These data indicate that cell motility of Lümmen/Rübben these human bladder carcinoma cells is tightly controlled by a complex network of interacting extracellular ligands [25].…”

“…The mechanism by which PTX prevented the development of high-grade tumor is not really clear. Besides cell motility LPA and other agonists of G-protein coupled receptors stimulates the growth of transitional cell carcinoma cells [25]. Several studies have shown that the ␣-subunits as well as ␤␥-subunits of these G proteins regulate several critical signalling pathways involved in cell proliferation, differentiation and apoptosis [47].…”

Tumor Cell Motility as a Novel Target in Cancer – Experimental and Clinical Results

“…Aim of this study was to examine the expression and the effects of G protein-coupled, calcium-mobilizing receptors on motility of J82 cells, a cell line originating from a human transitional-cell carcinoma. These cells express a variety of such receptors, including those for lysophosphatidic acid (LPA), thrombin, bradykinin, bombesin, and histamine [11]. Agonist activation of these receptors resulted in rapid and transient increase in [Ca 2+ ]i and inositol phosphate formation.…”