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Biochem J. 2005 Aug 15;390(Pt 1):199-206.

Transforming growth factor-beta receptors localize to caveolae and regulate endothelial nitric oxide synthase in normal human endothelial cells.

Author information

1
Division of Cardiovascular Medicine, Stanford University School of Medicine, Falk Center, 300 Pasteur Dr., MC5406, Stanford, CA 94305-5406, U.S.A.

Abstract

Caveolae (sphingolipid- and cholesterol-rich, 100 nm flask-shaped invaginations of the cell membrane) serve as a nexus of cell signalling. In the present study caveolin-rich lipid raft domains were extracted from HUVEC (human umbilical-vein endothelial cells) using both density gradient and immunoprecipitation techniques, and demonstrated localization of the TGF-beta (transforming growth factor-beta) receptors TbetaRI and TbetaRII to the Cav-1 (caveolin-1)-enriched raft fractions of these normal, human endothelial cells. Immunoprecipitation demonstrated an association between TbetaRI and TbetaRII, as well as an association of the TbetaRs receptors with Cav-1 and eNOS (endothelial nitric oxide synthase), suggesting a mutual co-localization to caveolae; after treatment of HUVEC with 5 ng/ml TGF-beta1 for 15 min, however, co-precipitation of eNOS with TbetaRI, TbetaRII and Cav-1 was diminished. The loss of immunoprecipitable eNOS from Cav-1-enriched fractions was accompanied by a decrease both in phosphorylation of eNOS and in enzymatic activity (conversion of arginine into citrulline). No change in the localization of eNOS to morphologically distinct caveolae could be detected by electron microscopy after treatment of HUVEC with TGF-beta1 for 20 min. The results of these investigations provide evidence that TbetaRI interacts with eNOS in the caveolae of normal, human endothelial cells and has a regulatory function on basal eNOS enzymatic activity.

PMID:
15819614
PMCID:
PMC1184575
DOI:
10.1042/BJ20041182
[Indexed for MEDLINE]
Free PMC Article

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