title: Modulation by sphingolipids of calcium signals evoked by epidermal growth factor.
abstract: Receptor-activated breakdown of complex sphingolipids has been proposed as a mechanism for generating sphingoid base-containing putative second messenger molecules whose actions may modulate responses to extracellular signals. In human epidermoid carcinoma A431 cells, sphingosine (1-10 microM) by itself had no effect on intracellular free calcium concentrations ([Ca2+]i), yet within seconds, markedly enhanced the epidermal growth factor (EGF)-evoked Ca2+ influx (by up to 2-fold), but failed to alter Ca2+ release from the intracellular stores. Ca2+ signals evoked by serum were not affected by sphingosine. The response to sphingosine was dose-dependent and saturable, exhibiting an EC50 of 2.3 microM. In contrast, a ceramide, N-acetylsphingosine (10 microM), sphingosine 1-phosphate (10 microM), and sphingosylphosphorylcholine (10 microM) inhibited EGF-evoked elevations in [Ca2+]i. The latter two compounds by themselves transiently increased [Ca2+]i. N-Octanoylsphingosine, N,N-dimethylsphingosine, sphingomyelin, and stearylamine were inactive. The potentiation of calcium signals by sphingosine occurred at all concentrations of EGF tested (0.15-15 nM) and did not alter the EGF receptor protein kinase activity as determined by antiphosphotyrosine immunoblotting. Antiphosphoserine immunoblotting revealed that sphingosine (10 microM for 3 min) increased the phosphoserine content of two proteins with approximate molecular masses of 40 and 70 kDa. Serine hyperphosphorylation of the 40-kDa protein was also observed in cells treated with EGF alone, whereas the intensity of the 70-kDa band was highest in cells treated with both sphingosine and EGF. The modulation of growth factor receptor-regulated signaling, including changes in [Ca2+]i, may constitute a mechanism by which elevations in cellular levels of specific sphingolipids, which occur transiently upon activation of certain receptors and chronically in sphingolipid storage diseases, exert their physiological and pathophysiological effects.
authors.affiliation: Department of Pathology, Boston University School of Medicine, Massachusetts 02118.
authors.forename: P L
authors.initials: PL
authors.lastname: Hudson
authors.forename: W A
authors.initials: WA
authors.lastname: Pedersen
authors.forename: W S
authors.initials: WS
authors.lastname: Saltsman
authors.forename: M
authors.initials: M
authors.lastname: Liscovitch
authors.forename: D T
authors.initials: DT
authors.lastname: MacLaughlin
authors.forename: P K
authors.initials: PK
authors.lastname: Donahoe
authors.forename: J K
authors.initials: JK
authors.lastname: Blusztajn
mesh_terms: D002118:Calcium / Q000378:metabolism*; D002294:Carcinoma, Squamous Cell / Q000378:metabolism*; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D006801:Humans; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D015290:Second Messenger Systems / Q000187:drug effects*; D013107:Sphingolipids / Q000494:pharmacology*; D013110:Sphingosine / Q000494:pharmacology; D014407:Tumor Cells, Cultured
chemical_list: D013107:Sphingolipids; D004815:Epidermal Growth Factor; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D013110:Sphingosine; D002118:Calcium