Ceramide-binding and activation defines protein kinase c-Raf as a ceramide-activated protein kinase.
Interleukin 1 is the prototype of an inflammatory cytokine, and evidence suggests that it uses the sphingomyelin pathway and ceramide production to trigger mitogen-activated protein kinase (MAPK) activation and subsequent gene expression required for acute inflammatory processes. To identify downstream signaling targets of ceramide, a radioiodinated photoaffinity labeling analog of ceramide ([125I] 3-trifluoromethyl-3-(m-iodophenyl)diazirine-ceramide) was employed. It is observed that ceramide specifically binds to and activates protein kinase c-Raf, leading to a subsequent activation of the MAPK cascade. Ceramide does not bind to any other member of the MAPK module nor does it bind to protein kinase C-zeta. These data identify protein kinase c-Raf as a specific molecular target for interleukin 1 beta-stimulated ceramide formation and demonstrate that ceramide is a lipid cofactor participating in regulation of c-Raf activity.
Huwiler
A
A
Department of Pharmacology, Biozentrum, University of Basel, Switzerland.
Brunner
J
J
Hummel
R
R
Vervoordeldonk
M
M
Stabel
S
S
van den Bosch
H
H
Pfeilschifter
J
J
D000345:Affinity Labels; D000818:Animals; D001389:Azirines / Q000302:isolation & purification / Q000378:metabolism*; D001665:Binding Sites; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002478:Cells, Cultured; D002518:Ceramides / Q000302:isolation & purification / Q000378:metabolism* / Q000494:pharmacology*; D004789:Enzyme Activation; D005920:Glomerular Mesangium / Q000378:metabolism*; D007375:Interleukin-1 / Q000494:pharmacology; D007700:Kinetics; D017346:Protein Serine-Threonine Kinases / Q000302:isolation & purification / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000302:isolation & purification / Q000378:metabolism*; D019908:Proto-Oncogene Proteins c-raf; D051381:Rats
C100592:3-trifluoromethyl-3-(3-iodophenyl)diazirineceramide; D000345:Affinity Labels; D001389:Azirines; D002518:Ceramides; D007375:Interleukin-1; D011518:Proto-Oncogene Proteins; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf; D017871:Calcium-Calmodulin-Dependent Protein Kinases