Evidence for tight coupling of Gi protein-mediated lysophosphatidic acid receptor to stimulated cytokine production in ovarian cancer cell

Objective: Lysophosphatidic acid stimulates the proliferation of ovarian cancer cell through specific members of the guanosine triphosphate-binding protein-coupled receptor family. We attempted to identify the,guanosine triphosphate-binding protein subtypes that are linked to lysophosphatidic acid receptor-stimulated production of cytokine, which are involved potentially in ovarian cancer development. Study design: Cytokine assay kits were used to determine interleukin-6, interleukin-8, and tumor necrotic factor-a that were produced from Caov-3 and SK-OV3 ovarian cancer cell lines. The a-subunit of Gi was detected by pertussis toxin-catalyzed adenosine diphosphate ribosylation from nicotinamide adenine dinucleotide in isolated plasma membrane. Results: Pertussis toxin, but not cholera toxin, brought about adenosine diphosphate ribosylation of Galphai of 41 kd in the plasma membrane. Incubation with lysophosphatidic acid and nonhydrolyzable guanosine triphosphate analog decreased the adenosine diphosphate-ribosylation activity in a dose-dependent manner; a one half-maximal effect occurred with 10 mumol/L lysophosphatidic acid.. The apparent inhibition by lysophosphatidic acid of the adenosine diphosphate ribosylation demonstrated that lysophosphatidic acid resolved the a-subunit of the Gi to guanosine triphosphate-bound form in the membranes. Pretreatment of the ovarian cancer cells with the pertussis toxin completely inhibited lysophosphatidic acid-stimulated production of interleukin-6, interleukin-8 and tumor necrosis factor-alpha. The lysophosphatidic acid-stimulated cytokine production was dose-dependent with a one half-maximal effect at 10 mumol/L. Phosphatidic acid and ceramide 1-phosphate had no effect on the lysophosphatidic acid action on cytokine expression. Conclusion: These data demonstrate the coupling of lysophosphatidic acid receptor to Gi protein subfamily in ovarian cancer cell. The Gi that couples lysophosphatidic acid receptor to the effector may define the differences in the signaling pathways of lysophosphatidic acid-activated cytokine expression and proliferation. (C) 2004 Elsevier Inc. All rights reserved.