PUBLICATION
Identification and characterization of murine mitochondrial-associated neutral sphingomyelinase (ma-nsmase), the mammalian sphingomyelin phosphodiesterase 5
- Authors
- Wu, B.X., Rajagopalan, V., Roddy, P.L., Clarke, C.J., and Hannun, Y.A.
- ID
- ZDB-PUB-100420-6
- Date
- 2010
- Source
- The Journal of biological chemistry 285(23): 17993-18002 (Journal)
- Registered Authors
- Keywords
- Enzyme kinetics, Lipid, Manganese, Mitochondria, Sphingolipid, MA-nSMase, ceramide, neutral sphingomyelinase
- MeSH Terms
-
- Cyclic Nucleotide Phosphodiesterases, Type 5/genetics*
- DNA, Complementary/metabolism
- Animals
- Mitochondria/metabolism*
- Mice
- Reverse Transcriptase Polymerase Chain Reaction
- RNA, Messenger/metabolism
- Sphingomyelins/metabolism*
- Sequence Homology, Amino Acid
- Cell Line, Tumor
- Amino Acid Sequence
- Molecular Sequence Data
- Cell Line
- Expressed Sequence Tags
- Humans
- Models, Biological
- PubMed
- 20378533 Full text @ J. Biol. Chem.
Citation
Wu, B.X., Rajagopalan, V., Roddy, P.L., Clarke, C.J., and Hannun, Y.A. (2010) Identification and characterization of murine mitochondrial-associated neutral sphingomyelinase (ma-nsmase), the mammalian sphingomyelin phosphodiesterase 5. The Journal of biological chemistry. 285(23):17993-18002.
Abstract
Sphingolipids play important roles in regulating cellular responses. Although mitochondria contain sphingolipids, direct regulation of their levels in mitochondria or mitochondria-associated membranes (MAMs) is mostly unclear. Neutral SMase (N-SMase) isoforms catalyze hydrolysis of sphingomyelin (SM) to ceramide and phosphocholine, have been found in the mitochondria of yeast and zebrafish, yet their existence in mammalian mitochondria remains unknown. Here, we have identified and cloned a cDNA based on nSMase homologous sequences. This cDNA encodes a novel protein of 483 amino acids that displays significant homology to nSMase2 and possesses the same catalytic core residues as members of the extended N-SMase family. A transiently expressed V5-tagged protein co-localized with both mitochondria and endoplasmic reticulum markers in MCF7 and HEK293 cells; accordingly, the enzyme is referred to as mitochondrial-associated nSMase (MA-nSMase). MA-nSMase was highly expressed in testis, pancreas, epididymis, and brain. MA-nSMase had an absolute requirement for cations such as Mg2+ and Mn2+, and activation by the anionic phospholipids, especially phosphatidylserine (PS) and the mitochondrial cardiolipin (CL). Importantly, overexpression of MA-nSMase in HEK293 cells significantly increased in vitro N-SMase activity and also modulated the levels of sphingomyelin and ceramide, indicating that the identified cDNA encodes a functional SMase. Thus, these studies identify and characterize for the first time, a mammalian MA-nSMase. The characterization of MA-nSMase described here will contribute to our understanding of pathways regulated by sphingolipid metabolites, particularly with reference to the mitochondria and associated organelles.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping