Format

Send to

Choose Destination
Mol Biol Cell. 2017 Nov 7;28(23):3371-3382. doi: 10.1091/mbc.E17-03-0136. Epub 2017 Sep 27.

The balance between Gαi-Cdc42/Rac and Gα12/13-RhoA pathways determines endothelial barrier regulation by sphingosine-1-phosphate.

Author information

1
van Leeuwenhoek Centre for Advanced Microscopy, Molecular Cytology, Swammerdam Institute for Life Sciences, University of Amsterdam, 1098 XH Amsterdam, Netherlands.
2
Molecular Cell Biology and.
3
University of Amsterdam Academic Medical Centre-Landsteiner Laboratory, Sanquin Research, 1066 CX Amsterdam, Netherlands.
4
Center for Cell Analysis and Modeling, University of Connecticut Health Center, Farmington, CT 06030.
5
van Leeuwenhoek Centre for Advanced Microscopy, Molecular Cytology, Swammerdam Institute for Life Sciences, University of Amsterdam, 1098 XH Amsterdam, Netherlands p.hordijk@vumc.nl j.goedhart@uva.nl.
6
Department of Physiology, Free University Medical Center, 1081 HZ Amsterdam, Netherlands.

Abstract

The bioactive sphingosine-1-phosphatephosphate (S1P) is present in plasma, bound to carrier proteins, and involved in many physiological processes, including angiogenesis, inflammatory responses, and vascular stabilization. S1P can bind to several G-protein-coupled receptors (GPCRs) activating a number of different signaling networks. At present, the dynamics and relative importance of signaling events activated immediately downstream of GPCR activation are unclear. To examine these, we used a set of fluorescence resonance energy transfer-based biosensors for different RhoGTPases (Rac1, RhoA/B/C, and Cdc42) as well as for heterotrimeric G-proteins in a series of live-cell imaging experiments in primary human endothelial cells. These experiments were accompanied by biochemical GTPase activity assays and transendothelial resistance measurements. We show that S1P promotes cell spreading and endothelial barrier function through S1PR1-Gαi-Rac1 and S1PR1-Gαi-Cdc42 pathways. In parallel, a S1PR2-Gα12/13-RhoA pathway is activated that can induce cell contraction and loss of barrier function, but only if Gαi-mediated signaling is suppressed. Our results suggest that Gαq activity is not involved in S1P-mediated regulation of barrier integrity. Moreover, we show that early activation of RhoA by S1P inactivates Rac1 but not Cdc42, and vice versa. Together, our data show that the rapid S1P-induced increase in endothelial integrity is mediated by a S1PR1-Gαi-Cdc42 pathway.

PMID:
28954861
PMCID:
PMC5687037
DOI:
10.1091/mbc.E17-03-0136
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Atypon Icon for PubMed Central
Loading ...
Support Center