Sphingomyelinase D, a novel probe for cellular sphingomyelin
Sphingomyelin (SM) and free cholesterol (FC) are concentrated in the plasma membranes of eukaryotes; however, the physiological significance of their association is unclear. A common tool for studying the role of membrane SM is digestion with bacterial sphingomyelinase (SMase) C, which hydrolyzes SM...
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| Format: | Article |
| Language: | English |
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Elsevier
2003-08-01
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| Series: | Journal of Lipid Research |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S0022227520337688 |
| _version_ | 1818931338561978368 |
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| author | Papasani V. Subbaiah Stephen J. Billington B. Helen Jost J. Glenn Songer Yvonne Lange |
| author_facet | Papasani V. Subbaiah Stephen J. Billington B. Helen Jost J. Glenn Songer Yvonne Lange |
| author_sort | Papasani V. Subbaiah |
| collection | DOAJ |
| description | Sphingomyelin (SM) and free cholesterol (FC) are concentrated in the plasma membranes of eukaryotes; however, the physiological significance of their association is unclear. A common tool for studying the role of membrane SM is digestion with bacterial sphingomyelinase (SMase) C, which hydrolyzes SM to ceramide. However, it is not known whether the observed effects of SMase C treatment are due to the loss of SM per se or to the signaling effects of ceramide. In this study, we tested SMase D from Corynebacterium pseudotuberculosis, which hydrolyzes SM to ceramide phosphate, as an alternative probe. This enzyme specifically hydrolyzed SM in fibroblasts without causing accumulation of ceramide. Treatment of fibroblasts with SMase D stimulated translocation of PM FC to intracellular sites by <20% of the rate observed after SMase C digestion. The cells regenerated SM nearly completely within 5 h after SMase C treatment. However, even after 20 h, no regeneration occurred following SMase D digestion.These findings suggest that the translocation of PM FC caused by SMase C digestion is due to the cellular effects of ceramide rather than the loss of SM. Since ceramide phosphate does not appear to have such effects, we suggest that SMase D is a useful probe of membrane SM. |
| first_indexed | 2024-12-20T04:15:00Z |
| format | Article |
| id | doaj.art-d6a7c4b7fd2844c6ac570d3640e25f6c |
| institution | Directory Open Access Journal |
| issn | 0022-2275 |
| language | English |
| last_indexed | 2024-12-20T04:15:00Z |
| publishDate | 2003-08-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Journal of Lipid Research |
| spelling | doaj.art-d6a7c4b7fd2844c6ac570d3640e25f6c2022-12-21T19:53:49ZengElsevierJournal of Lipid Research0022-22752003-08-0144815741580Sphingomyelinase D, a novel probe for cellular sphingomyelinPapasani V. Subbaiah0Stephen J. Billington1B. Helen Jost2J. Glenn Songer3Yvonne Lange4Department of Medicine, Rush Medical College, Chicago, IL 60612; Department of Pathology, Rush Medical College, Chicago, IL 60612; Department of Biochemistry, Rush Medical College, Chicago, IL 60612; Department of Veterinary Science and Microbiology, University of Arizona, Tucson, AZ 85721Department of Medicine, Rush Medical College, Chicago, IL 60612; Department of Pathology, Rush Medical College, Chicago, IL 60612; Department of Biochemistry, Rush Medical College, Chicago, IL 60612; Department of Veterinary Science and Microbiology, University of Arizona, Tucson, AZ 85721Department of Medicine, Rush Medical College, Chicago, IL 60612; Department of Pathology, Rush Medical College, Chicago, IL 60612; Department of Biochemistry, Rush Medical College, Chicago, IL 60612; Department of Veterinary Science and Microbiology, University of Arizona, Tucson, AZ 85721Department of Medicine, Rush Medical College, Chicago, IL 60612; Department of Pathology, Rush Medical College, Chicago, IL 60612; Department of Biochemistry, Rush Medical College, Chicago, IL 60612; Department of Veterinary Science and Microbiology, University of Arizona, Tucson, AZ 85721Department of Medicine, Rush Medical College, Chicago, IL 60612; Department of Pathology, Rush Medical College, Chicago, IL 60612; Department of Biochemistry, Rush Medical College, Chicago, IL 60612; Department of Veterinary Science and Microbiology, University of Arizona, Tucson, AZ 85721Sphingomyelin (SM) and free cholesterol (FC) are concentrated in the plasma membranes of eukaryotes; however, the physiological significance of their association is unclear. A common tool for studying the role of membrane SM is digestion with bacterial sphingomyelinase (SMase) C, which hydrolyzes SM to ceramide. However, it is not known whether the observed effects of SMase C treatment are due to the loss of SM per se or to the signaling effects of ceramide. In this study, we tested SMase D from Corynebacterium pseudotuberculosis, which hydrolyzes SM to ceramide phosphate, as an alternative probe. This enzyme specifically hydrolyzed SM in fibroblasts without causing accumulation of ceramide. Treatment of fibroblasts with SMase D stimulated translocation of PM FC to intracellular sites by <20% of the rate observed after SMase C digestion. The cells regenerated SM nearly completely within 5 h after SMase C treatment. However, even after 20 h, no regeneration occurred following SMase D digestion.These findings suggest that the translocation of PM FC caused by SMase C digestion is due to the cellular effects of ceramide rather than the loss of SM. Since ceramide phosphate does not appear to have such effects, we suggest that SMase D is a useful probe of membrane SM.http://www.sciencedirect.com/science/article/pii/S0022227520337688ceramideceramide phosphateacyl-CoA:cholesterol acyltransferasecholesterol effluxsphingomyelinsphingomyelinase |
| spellingShingle | Papasani V. Subbaiah Stephen J. Billington B. Helen Jost J. Glenn Songer Yvonne Lange Sphingomyelinase D, a novel probe for cellular sphingomyelin Journal of Lipid Research ceramide ceramide phosphate acyl-CoA:cholesterol acyltransferase cholesterol efflux sphingomyelin sphingomyelinase |
| title | Sphingomyelinase D, a novel probe for cellular sphingomyelin |
| title_full | Sphingomyelinase D, a novel probe for cellular sphingomyelin |
| title_fullStr | Sphingomyelinase D, a novel probe for cellular sphingomyelin |
| title_full_unstemmed | Sphingomyelinase D, a novel probe for cellular sphingomyelin |
| title_short | Sphingomyelinase D, a novel probe for cellular sphingomyelin |
| title_sort | sphingomyelinase d a novel probe for cellular sphingomyelin |
| topic | ceramide ceramide phosphate acyl-CoA:cholesterol acyltransferase cholesterol efflux sphingomyelin sphingomyelinase |
| url | http://www.sciencedirect.com/science/article/pii/S0022227520337688 |
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