Stimulation of DNA synthesis by natural ceramide 1-phosphate.
We found that natural (long-chain) ceramide 1-phosphate can be dispersed into aqueous solution when dissolved in an appropriate mixture of methanol/dodecane (49:1, v/v). This solvent mixture facilitates the interaction of this phosphosphingolipid with cells. Under these conditions, incubation of EGFR T17 fibroblasts with natural ceramide 1-phosphate caused a potent stimulation of DNA synthesis. This effect was accompanied by an increase in the levels of proliferating-cell nuclear antigen. Concentrations of natural ceramide 1-phosphate that stimulated the synthesis of DNA did not inhibit adenylate cyclase activity, nor did they stimulate phospholipase D. Natural ceramide 1-phosphate did not alter the cellular phosphorylation state of tyrosine residues or of mitogen-activated protein kinase. Furthermore, natural ceramide 1-phosphate failed to induce the expression of the proto-oncogenes c-myc and c-fos. Both the stimulation of DNA synthesis and the induction of proliferating-cell nuclear antigen by natural ceramide 1-phosphate were inhibited by natural ceramides. This work suggests that the use of methanol and dodecane to deliver natural ceramide 1-phosphate to cells may be useful for elucidation of the biological function(s) and mechanism(s) of action of ceramide 1-phosphate.
Gomez-Muñoz
A
A
Instituto de Investigaciones Biomédicas, Consejo Superior de Investigaciones Científicas (CSIC), C/Arturo Duperier 4, 28029-Madrid, Spain.
Frago
L M
LM
Alvarez
L
L
Varela-Nieto
I
I
D016475:3T3 Cells; D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002518:Ceramides / Q000138:chemical synthesis / Q000378:metabolism / Q000494:pharmacology*; D004247:DNA / Q000096:biosynthesis*; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects / Q000235:genetics; D016762:Genes, fos / Q000235:genetics; D016259:Genes, myc / Q000235:genetics; D051379:Mice; D010766:Phosphorylation; D018809:Proliferating Cell Nuclear Antigen / Q000096:biosynthesis
D002518:Ceramides; D018809:Proliferating Cell Nuclear Antigen; C065576:ceramide 1-phosphate; D004815:Epidermal Growth Factor; D004247:DNA; D017871:Calcium-Calmodulin-Dependent Protein Kinases