Regulation of sphingosine 1-phosphate-induced endothelial cytoskeletal rearrangement and barrier enhancement by S1P1 receptor, PI3 kinase, Tiam1/Rac1, and alpha-actinin.

TitleRegulation of sphingosine 1-phosphate-induced endothelial cytoskeletal rearrangement and barrier enhancement by S1P1 receptor, PI3 kinase, Tiam1/Rac1, and alpha-actinin.
Publication TypeJournal Article
Year of Publication2005
AuthorsSingleton PA, Dudek SM, Chiang ET, Garcia JGN
JournalFASEB J
Volume19
Issue12
Pagination1646-56
Date Published2005 Oct
ISSN Number1530-6860
KeywordsActinin, Catalytic Domain, Caveolin 1, Cells, Cultured, Cholesterol, Chromones, Cytoskeleton, Electrophoresis, Polyacrylamide Gel, Endothelium, Vascular, Enzyme Inhibitors, Gene Expression Regulation, Enzymologic, Guanine Nucleotide Exchange Factors, Humans, Immunoblotting, Immunoprecipitation, Inflammation, Microfilament Proteins, Microscopy, Fluorescence, Models, Biological, Morpholines, Neoplasm Proteins, Phosphatidylinositol 3-Kinases, Protein Isoforms, Protein Structure, Tertiary, Pulmonary Artery, rac1 GTP-Binding Protein, Receptors, Lysosphingolipid, RNA, Small Interfering, Signal Transduction, T-Lymphoma Invasion and Metastasis-inducing Protein 1, Transfection
Abstract

<p>Endothelial cell (EC) barrier dysfunction results in increased vascular permeability observed in inflammation, tumor angiogenesis, and atherosclerosis. The platelet-derived phospholipid sphingosine-1-phosphate (S1P) decreases EC permeability in vitro and in vivo and thus has obvious therapeutic potential. We examined S1P-mediated human pulmonary artery EC signaling and barrier regulation in caveolin-enriched microdomains (CEM). Immunoblotting from S1P-treated EC revealed S1P-mediated rapid recruitment (1 microM, 5 min) to CEMs of the S1P receptors S1P1 and S1P3, p110 PI3 kinase alpha and beta catalytic subunits, the Rac1 GEF, Tiam1, and alpha-actinin isoforms 1 and 4. Immunoprecipitated p110 PI3 kinase catalytic subunits from S1P-treated EC exhibited PIP3 production in CEMs. Immunoprecipitation of S1P receptors from CEM fractions revealed complexes containing Tiam1 and S1P1. PI3 kinase inhibition (LY294002) attenuated S1P-induced Tiam1 association with S1P1, Tiam1/Rac1 activation, alpha-actinin-1/4 recruitment, and EC barrier enhancement. Silencing of either S1P1 or Tiam1 expression resulted in the loss of S1P-mediated Rac1 activation and alpha-actinin-1/4 recruitment to CEM. Finally, silencing S1P1, Tiam1, or both alpha-actinin isoforms 1/4 inhibits S1P-induced cortical F-actin rearrangement and S1P-mediated barrier enhancement. Taken together, these results suggest that S1P-induced recruitment of S1P1 to CEM fractions promotes PI3 kinase-mediated Tiam1/Rac1 activation required for alpha-actinin-1/4-regulated cortical actin rearrangement and EC barrier enhancement.</p>

DOI10.1096/fj.05-3928com
Alternate JournalFASEB J.
PubMed ID16195373