Recent studies by our group and others have disclosed the presence of ceramides in mitochondria, and the activities of ceramide synthase and reverse ceramidase in mitochondria have also been reported. Since a possible contamination with the ER (endoplasmic reticulum)-related compartment MAM (mitochondria-associated membrane) could not be ruled out in previous studies, we have re-investigated the presence of the enzymes of ceramide metabolism in mitochondria and MAM highly purified from rat liver. In the present paper, we show that purified mitochondria as well as MAM are indeed able to generate ceramide in vitro through both ceramide synthase or reverse ceramidase, whereas the latter enzyme activity is barely detectable in microsomes. Moreover, ceramide synthase activities were recovered in outer mitochondrial membranes as well as in inner mitochondrial membranes. Using radiolabelled sphingosine as a substrate, mitochondria could generate ceramide and phytoceramide. However, the in vitro sensitivity of ceramide synthase toward FB1 (fumonisin B1) in mitochondria as well as in MAM was found to depend upon the sphingoid base: whereas dihydrosphingosine N-acyltransferase was inhibited by FB1 in a concentration-dependent manner, FB1 actually activated the ceramide synthase when using sphingosine as a substrate. Acylation of sphingosine 1-phosphate and dihydrosphingosine 1-phosphate, generating ceramide 1-phosphate, was also shown with both subcellular fractions. Moreover, the same difference in sensitivity towards FB1 for the ceramide synthase activities was seen between the two phosphorylated sphingoid bases, raising the possibility that distinct base-specific enzymes may be involved as ceramide synthases. Collectively, these results demonstrate the involvement of mitochondria in the metabolism of ceramides through different pathways, thereby supporting the hypothesis that topology of ceramide formation could determine its function.
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Research Article|
August 24 2004
Subcellular compartmentalization of ceramide metabolism: MAM (mitochondria-associated membrane) and/or mitochondria?
Clara BIONDA;
Clara BIONDA
*Laboratory of Radiobiology, UCBL EA-37-38, Lyon-Sud Medical School, B.P. 12, 69921 Oullins Cx, France
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Jacques PORTOUKALIAN;
Jacques PORTOUKALIAN
1
†INSERM U346 and UCBL EA-3732, Department of Dermatology, Edouard Herriot Hospital, 69437 Lyon Cx 03, France
1To whom correspondence should be addressed (email portoukalian@lyon.inserm.fr).
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Daniel SCHMITT;
Daniel SCHMITT
†INSERM U346 and UCBL EA-3732, Department of Dermatology, Edouard Herriot Hospital, 69437 Lyon Cx 03, France
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Claire RODRIGUEZ-LAFRASSE;
Claire RODRIGUEZ-LAFRASSE
*Laboratory of Radiobiology, UCBL EA-37-38, Lyon-Sud Medical School, B.P. 12, 69921 Oullins Cx, France
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Dominique ARDAIL
Dominique ARDAIL
*Laboratory of Radiobiology, UCBL EA-37-38, Lyon-Sud Medical School, B.P. 12, 69921 Oullins Cx, France
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Publisher: Portland Press Ltd
Received:
November 27 2003
Revision Received:
April 27 2004
Accepted:
May 14 2004
Accepted Manuscript online:
May 14 2004
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London
2004
Biochem J (2004) 382 (2): 527–533.
Article history
Received:
November 27 2003
Revision Received:
April 27 2004
Accepted:
May 14 2004
Accepted Manuscript online:
May 14 2004
Citation
Clara BIONDA, Jacques PORTOUKALIAN, Daniel SCHMITT, Claire RODRIGUEZ-LAFRASSE, Dominique ARDAIL; Subcellular compartmentalization of ceramide metabolism: MAM (mitochondria-associated membrane) and/or mitochondria?. Biochem J 1 September 2004; 382 (2): 527–533. doi: https://doi.org/10.1042/BJ20031819
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