title: Role of Sphingosine Kinase 1 and S1P Transporter Spns2 in HGF-mediated Lamellipodia Formation in Lung Endothelium.
abstract: Hepatocyte growth factor (HGF) signaling via c-Met is known to promote endothelial cell motility and angiogenesis. We have previously reported that HGF stimulates lamellipodia formation and motility of human lung microvascular endothelial cells (HLMVECs) via PI3K/Akt signal transduction and reactive oxygen species generation. Here, we report a role for HGF-induced intracellular sphingosine-1-phosphate (S1P) generation catalyzed by sphingosine kinase 1 (SphK1), S1P transporter, spinster homolog 2 (Spns2), and S1P receptor, S1P1, in lamellipodia formation and perhaps motility of HLMVECs. HGF stimulated SphK1 phosphorylation and enhanced intracellular S1P levels in HLMVECs, which was blocked by inhibition of SphK1. HGF enhanced co-localization of SphK1/p-SphK1 with actin/cortactin in lamellipodia and down-regulation or inhibition of SphK1 attenuated HGF-induced lamellipodia formation in HLMVECs. In addition, down-regulation of Spns2 also suppressed HGF-induced lamellipodia formation, suggesting a key role for inside-out S1P signaling. The HGF-mediated phosphorylation of SphK1 and its localization in lamellipodia was dependent on c-Met and ERK1/2 signaling, but not the PI3K/Akt pathway; however, blocking PI3K/Akt signaling attenuated HGF-mediated phosphorylation of Spns2. Down-regulation of S1P1, but not S1P2or S1P3, with specific siRNA attenuated HGF-induced lamellipodia formation. Further, HGF enhanced association of Spns2 with S1P1that was blocked by inhibiting SphK1 activity with PF-543. Moreover, HGF-induced migration of HLMVECs was attenuated by down-regulation of Spns2.Taken together, these results suggest that HGF/c-Met-mediated lamellipodia formation, and perhaps motility is dependent on intracellular generation of S1P via activation and localization of SphK1 to cell periphery and Spns2-mediated extracellular transportation of S1P and its inside-out signaling via S1P1.
authors.affiliation: From the Departments of Pharmacology.
authors.forename: Panfeng
authors.initials: P
authors.lastname: Fu
authors.affiliation: Biochemistry and Molecular Genetics.
authors.forename: David L
authors.initials: DL
authors.lastname: Ebenezer
authors.affiliation: the Department of Medicine, National Jewish Health, Denver, Colorado 80206.
authors.forename: Evgeny V
authors.initials: EV
authors.lastname: Berdyshev
authors.affiliation: the Department of Medicine, National Jewish Health, Denver, Colorado 80206.
authors.forename: Irina A
authors.initials: IA
authors.lastname: Bronova
authors.affiliation: From the Departments of Pharmacology.
authors.forename: Mark
authors.initials: M
authors.lastname: Shaaya
authors.affiliation: Pediatrics, and.
authors.forename: Anantha
authors.initials: A
authors.lastname: Harijith
authors.affiliation: From the Departments of Pharmacology, visnatar@uic.edu.|Medicine, University of Illinois, Chicago, Illinois 60612 and.
authors.forename: Viswanathan
authors.initials: V
authors.lastname: Natarajan
mesh_terms: D027321:Anion Transport Proteins / Q000378:metabolism*; D002465:Cell Movement; D002478:Cells, Cultured; D051356:Cortactin / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology* / Q000378:metabolism; D017228:Hepatocyte Growth Factor / Q000378:metabolism*; D006801:Humans; D008168:Lung / Q000166:cytology* / Q000378:metabolism; D008246:Lysophospholipids / Q000378:metabolism; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D019859:Proto-Oncogene Proteins c-met / Q000378:metabolism; D011554:Pseudopodia / Q000378:metabolism*; D049349:Receptors, Lysosphingolipid / Q000378:metabolism; D015398:Signal Transduction; D013110:Sphingosine / Q000031:analogs & derivatives / Q000378:metabolism; D000099157:Sphingosine Kinase
chemical_list: D027321:Anion Transport Proteins; D051356:Cortactin; D017228:Hepatocyte Growth Factor; D008246:Lysophospholipids; D017853:Phosphotransferases (Alcohol Group Acceptor); D019859:Proto-Oncogene Proteins c-met; D049349:Receptors, Lysosphingolipid; D013110:Sphingosine; D000099157:Sphingosine Kinase; C495868:CTTN protein, human; C066855:HGF protein, human; C000598727:Spns2 protein, human; C060506:sphingosine 1-phosphate; C491743:MET protein, human
keywords: Sphingosine kinase (SphK); Spns2; cell signaling; hepatocyte growth factor/scatter factor (HGF/SF); lipid signaling; sphingosine-1-phosphate (S1P); trans