Activation of mTOR and RhoA is a major mechanism by which Ceramide 1-phosphate stimulates macrophage proliferation.
This study tested the hypothesis that Ceramide 1-phosphate (C1P) stimulates macrophage proliferation through activation of the mammalian target of rapamycin (mTOR). We first reported that C1P is mitogenic for fibroblasts and macrophages, but the mechanisms whereby it stimulates cell proliferation are incompletely understood. Here we demonstrate that C1P causes phosphorylation of mTOR in primary (bone marrow-derived) macrophages. Activation of this kinase was tested my measuring the phosphorylation state of its downstream target p70S6K after treatment with C1P. These actions were dependent upon prior activation of phosphoinositide 3 kinase (PI3-K), as selective inhibition of this kinase blocked mTOR phosphorylation and activation. In addition, C1P caused phosphorylation of PRAS40, a component of the mTOR complex 1 (mTORC1) that is absent in mTORC2. Furthermore, inhibition of the small G protein Ras homolog enriched in brain (Rheb), which is also a specific component of mTORC1, with FTI277, completely blocked C1P-stimulated mTOR phosphorylation, DNA synthesis and macrophage growth. In addition, C1P caused phosphorylation of another Ras homolog gene family member, RhoA, which is also involved in cell proliferation. Interestingly, inhibition of the RhoA downstream effector RhoA-associated kinase (ROCK) also blocked C1P-stimulated mTOR and cell proliferation. It can be concluded that mTORC1, and RhoA/ROCK are essential components of the mechanism whereby C1P stimulates macrophage proliferation.
Gangoiti
Patricia
P
Department of Biochemistry and Molecular Biology, Faculty of Science and Technology, University of the Basque Country, Bilbao, Spain.
Arana
Lide
L
Ouro
Alberto
A
Granado
Maria H
MH
Trueba
Miguel
M
Gómez-Muñoz
Antonio
A
D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D049109:Cell Proliferation; D002478:Cells, Cultured; D002518:Ceramides / Q000494:pharmacology*; D005260:Female; D008264:Macrophages / Q000166:cytology* / Q000201:enzymology* / Q000276:immunology; D000076222:Mechanistic Target of Rapamycin Complex 1; D051379:Mice; D020559:Monomeric GTP-Binding Proteins / Q000378:metabolism; D046912:Multiprotein Complexes; D009479:Neuropeptides / Q000378:metabolism; D037342:Pertussis Toxin / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D011506:Proteins / Q000378:metabolism; D000076205:Ras Homolog Enriched in Brain Protein; D015398:Signal Transduction; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*; D054460:rho-Associated Kinases / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
D000900:Anti-Bacterial Agents; D002518:Ceramides; D000076222:Mechanistic Target of Rapamycin Complex 1; D020559:Monomeric GTP-Binding Proteins; D046912:Multiprotein Complexes; D009479:Neuropeptides; D037342:Pertussis Toxin; D019869:Phosphatidylinositol 3-Kinases; D011506:Proteins; D000076205:Ras Homolog Enriched in Brain Protein; D020123:Sirolimus; D058570:TOR Serine-Threonine Kinases; D054460:rho-Associated Kinases; D020742:rhoA GTP-Binding Protein; C490213:Rheb protein, mouse; C065576:ceramide 1-phosphate