|ZFIN ID: ZDB-PUB-080825-17|
Identification of Mg2+-dependent neutral sphingomyelinase 1 as a mediator of heat stress-induced ceramide generation and apoptosis
Yabu, T., Imamura, S., Yamashita, M., and Okazaki, T.
|Source:||The Journal of biological chemistry 283(44): 29971-29982 (Journal)|
|Registered Authors:||Yamashita, Michiaki|
|PubMed:||18678863 Full text @ J. Biol. Chem.|
Yabu, T., Imamura, S., Yamashita, M., and Okazaki, T. (2008) Identification of Mg2+-dependent neutral sphingomyelinase 1 as a mediator of heat stress-induced ceramide generation and apoptosis. The Journal of biological chemistry. 283(44):29971-29982.
ABSTRACTA neutral sphingomyelinase (SMase) is involved in the induction of ceramide-mediated pro-apoptotic signaling under heat stress. Although ceramide is an important mediator of apoptosis, the neutral SMase that is activated under heat stress has not been identified. Our examination indicated that neutral SMase 1, which was isolated from zebrafish embryonic cultured ZE cells, was a mediator of stress-induced apoptosis. Mg2+-dependent neutral SMase was cloned from a cDNA library from ZE cells using an Escherichia coli expression vector by an SMase assay against the substrate C6-7-nitro-2-1, 3-benzoxadiazol-4-yl-sphingomyelin. The isolated cDNA clone encoded a polypeptide of 420 amino acids (putative molecular weight: 46.9 K) containing two predicted transmembrane domains in the C-terminal region. Bacterially expressed recombinant neutral SMase 1 hydrolyzed [choline-methyl-14C]sphingomyelin optimally at pH 7.5 in the presence of Mg2+ ion. In the endogenous SMase in ZE cells, the enzyme was localized in the microsomal fraction. The overexpressed FLAG-tagged SMase was co-localized with a Glogi cytostaining marker by cytochemical observation. The loss of function of neutral SMase 1 by anti-sense phosphorothioate oligonucleotides in ZE cells repressed ceramide generation, caspase-3 activation, and apoptotic cell death under heat stress. Thus, neutral SMase 1 participates in an inducible ceramide-mediating pro-apoptotic signaling pathway that operates heat-induced apoptosis.