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Intracellular S1P generation is essential for S1P-induced motility of human lung endothelial cells: Role of sphingosine kinase 1 and S1P lyase

Berdyshev, EV and Gorshkova, I and Usatyuk, P and Kalari, S and Zhao, Y and Pyne, NJ and Pyne, S and Sabbadini, RA and Garcia, JGN and Natarajan, V (2011) Intracellular S1P generation is essential for S1P-induced motility of human lung endothelial cells: Role of sphingosine kinase 1 and S1P lyase. PLoS ONE, 6 (1).

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Background: Earlier we have shown that extracellular sphingosine-1-phosphate (S1P) induces migration of human pulmonary artery endothelial cells (HPAECs) through the activation of S1P1 receptor, PKCε, and PLD2-PKCζ-Rac1 signaling cascade. As endothelial cells generate intracellular S1P, here we have investigated the role of sphingosine kinases (SphKs) and S1P lyase (S1PL), that regulate intracellular S1P accumulation, in HPAEC motility. Methodology/Principal Findings: Inhibition of SphK activity with a SphK inhibitor 2-(p-Hydroxyanilino)-4-(p-Chlorophenyl) Thiazole or down-regulation of Sphk1, but not SphK2, with siRNA decreased S1Pint, and attenuated S1Pext or serum-induced motility of HPAECs. On the contrary, inhibition of S1PL with 4-deoxypyridoxine or knockdown of S1PL with siRNA increased S1Pint and potentiated motility of HPAECs to S1Pext or serum. S1Pext mediates cell motility through activation of Rac1 and IQGAP1 signal transduction in HPAECs. Silencing of SphK1 by siRNA attenuated Rac1 and IQGAP1 translocation to the cell periphery; however, knockdown of S1PL with siRNA or 4-deoxypyridoxine augmented activated Rac1 and stimulated Rac1 and IQGAP1 translocation to cell periphery. The increased cell motility mediated by down-regulation was S1PL was pertussis toxin sensitive suggesting "inside-out" signaling of intracellularly generated S1P. Although S1P did not accumulate significantly in media under basal or S1PL knockdown conditions, addition of sodium vanadate increased S1P levels in the medium and inside the cells most likely by blocking phosphatases including lipid phosphate phosphatases (LPPs). Furthermore, addition of anti-S1P mAb to the incubation medium blocked S1Pext or 4-deoxypyridoxine-dependent endothelial cell motility. Conclusions/Significance: These results suggest S1Pext mediated endothelial cell motility is dependent on intracellular S1P production, which is regulated, in part, by SphK1 and S1PL. © 2011 Berdyshev et al.


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Item Type: Article
Status: Published
CreatorsEmailPitt UsernameORCID
Berdyshev, EV
Gorshkova, I
Usatyuk, P
Kalari, S
Zhao, Yyuz42@pitt.eduYUZ42
Pyne, NJ
Pyne, S
Sabbadini, RA
Garcia, JGN
Natarajan, V
Date: 9 February 2011
Date Type: Publication
Journal or Publication Title: PLoS ONE
Volume: 6
Number: 1
DOI or Unique Handle: 10.1371/journal.pone.0016571
Refereed: Yes
MeSH Headings: Aldehyde-Lyases--physiology; Cell Movement; Endothelial Cells--cytology; Humans; Lung--cytology; Lysophospholipids--biosynthesis; Lysophospholipids--physiology; Phosphotransferases (Alcohol Group Acceptor)--physiology; RNA, Small Interfering--pharmacology; Sphingosine--analogs & derivatives; Sphingosine--biosynthesis; Sphingosine--physiology
Other ID: NLM PMC3031585
PubMed Central ID: PMC3031585
PubMed ID: 21304987
Date Deposited: 03 Aug 2012 19:01
Last Modified: 19 Jun 2021 10:55


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