The sphingomyelin/ceramide pathway is involved in ERK1/2 phosphorylation, cell proliferation, and uPAR overexpression induced by tissue-type plasminogen activator.
Plasminogen activators (tPA and uPA) are serine proteases that convert the circulating zymogen plasminogen to active plasmin and mediate fibrin degradation. These multifunctional proteins trigger various biological events such as extracellular matrix degradation, cell adhesion, migration, and proliferation, through not yet fully characterized mechanisms. We report that, in smooth muscle cells and ECV-304 carcinoma cells, tPA and ATF (the N-terminal catalytically inactive fragment of tPA) elicited DNA synthesis that requires activation of the sphingomyelin/ceramide/sphingosine-1-phosphate (Spm/Cer/S1P), signaling pathway and was blocked by D-erythro-2-(N-myristoylamino)-1-phenyl-propanol (D-MAPP) and N-N'-dimethyl sphingosine (DMS), two classical inhibitors of sphingosine-1-phosphate biosynthesis. Binding of tPA to its receptor uPAR triggered the coordinated activation of two key enzymes of the Spm/Cer/S1P pathway, the neutral sphingomyelinase and the sphingosine kinase-1 that was mediated by a common pertussis toxin (PTX)-sensitive mechanism. The tPA-induced sphingosine kinase-1 activation was mediated by Src, since it was inhibited by herbimycin A and in SrcK- cells (overexpressing a dominant negative kinase defective form of Src) and by ERK1/2 (early phase peaking at 15 min). Sphingosine kinase-1 activation was followed by a second phase of ERK1/2 phosphorylation (peaking at 120 min) and subsequent DNA synthesis, which were inhibited by D-MAPP and DMS, by anti-EGD-1 antibodies and in SrcK- cells (in which the mitogenic signaling was rescued by sphingosine-1-phosphate). Altogether, these data underline a pivotal role for the Spm/Cer/S1P pathway in the tPA-induced mitogenic signaling.
Maupas-Schwalm
Françoise
F
Inserm U-466, CHU Rangueil, Toulouse, France.
Augé
Nathalie
N
Robinet
Catherine
C
Cambus
Jean-pierre
JP
Parsons
Sarah J
SJ
Salvayre
Robert
R
Nègre-Salvayre
Anne
A
D016227:Benzoquinones; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D002518:Ceramides / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D047029:Lactams, Macrocyclic; D008246:Lysophospholipids / Q000494:pharmacology; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008934:Mitogens / Q000494:pharmacology; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000494:pharmacology; D037342:Pertussis Toxin / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism; D011809:Quinones / Q000494:pharmacology; D017828:Rifabutin / Q000031:analogs & derivatives; D015398:Signal Transduction / Q000187:drug effects*; D013109:Sphingomyelins / Q000378:metabolism*; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D010959:Tissue Plasminogen Activator / Q000494:pharmacology*; D015854:Up-Regulation / Q000187:drug effects*; D014568:Urokinase-Type Plasminogen Activator / Q000737:chemistry / Q000235:genetics* / Q000494:pharmacology; D000099157:Sphingosine Kinase
D016227:Benzoquinones; D002518:Ceramides; D048049:Extracellular Signal-Regulated MAP Kinases; D047029:Lactams, Macrocyclic; D008246:Lysophospholipids; D008934:Mitogens; D010446:Peptide Fragments; D037342:Pertussis Toxin; D017853:Phosphotransferases (Alcohol Group Acceptor); D011809:Quinones; D017828:Rifabutin; D013109:Sphingomyelins; D013110:Sphingosine; D010959:Tissue Plasminogen Activator; D014568:Urokinase-Type Plasminogen Activator; D000099157:Sphingosine Kinase; C060506:sphingosine 1-phosphate; C020754:herbimycin