The general goal of this grant proposal is to define a signal transduction pathway for tumor necrosis factor (TNF)-alpha. This cytokine is a principal host defense against foreign antigen and may serve a variety of physiologic regulatory functions. Although a number of reports have suggested a role for one of the known signalling systems, no coherent picture has emerged for a second messenger pathway that may account for the entirety of the pleiotropic effects of TNF-alpha. This proposal is based on the recent observation that sphingomyelin degradation to ceramide is an early event in TNF-alpha action in human promyelocytic leukemia (HL-60) cells and that ceramide may substitute for TNF-alpha in monocytic differentiation. During the past few years, this laboratory has described a new metabolic pathway involving sphingomyelin and its derivatives. This pathway is initiated by hydrolysis of plasma membrane sphingomyelin to ceramide by the action of a sphingomyelinase. Subsequently, ceramide may be de-acylated to sphingosine, an inhibitor of protein kinase C, or phosphorylated to ceramide 1-phosphate, a compound newly discovered in this laboratory. Because of apparent similarities between this metabolic pathway and the phosphoinositide pathway, it was postulated that ceramide may serve second messenger function. Indeed, a cell-permeable ceramide analog induced selective phosphorylation of the epidermal growth factor receptor on threonine 669. Based on this observation, this laboratory has recently characterized a novel kinase that mediates this action of ceramide, termed ceramide-activated protein kinase, in membranes derived from HL-60 and A431 human epidermoid carcinoma cells. Elevation of ceramide levels by TNF-alpha also resulted in enhanced kinase activity in membranes derived from stimulated cells.
The specific aims of this grant are to: (1) demonstrate that this sphingomyelin pathway is tightly coupled to activation of the TNF receptor both in intact cells and in vitro: (2) show that this pathway is sufficient to mediate TNF-alpha-induced protein phosphorylation; and (3) generalize this pathway to various models of TNF- alpha action and in particular determine how it might relate to TNF-induced cytotoxicity and cytostasis. Hopefully, these studies will provide fundamental insights into the mechanism of TNF-alpha action and allow for eventual pharmacologic manipulation of this system.
| Kester, Mark; Kolesnick, Richard (2003) Sphingolipids as therapeutics. Pharmacol Res 47:365-71 |
| Kolesnick, R N; Kronke, M (1998) Regulation of ceramide production and apoptosis. Annu Rev Physiol 60:643-65 |
| Haimovitz-Friedman, A; Kolesnick, R N; Fuks, Z (1997) Ceramide signaling in apoptosis. Br Med Bull 53:539-53 |
| Haimovitz-Friedman, A; Kolesnick, R N; Fuks, Z (1997) Differential inhibition of radiation-induced apoptosis. Stem Cells 15 Suppl 2:43-7 |
| Santana, P; Pena, L A; Haimovitz-Friedman, A et al. (1996) Acid sphingomyelinase-deficient human lymphoblasts and mice are defective in radiation-induced apoptosis. Cell 86:189-99 |
| Bose, R; Verheij, M; Haimovitz-Friedman, A et al. (1995) Ceramide synthase mediates daunorubicin-induced apoptosis: an alternative mechanism for generating death signals. Cell 82:405-14 |
| Kolesnick, R; Fuks, Z (1995) Ceramide: a signal for apoptosis or mitogenesis? J Exp Med 181:1949-52 |
| Wright, S D; Kolesnick, R N (1995) Does endotoxin stimulate cells by mimicking ceramide? Immunol Today 16:297-302 |
| Joseph, C K; Wright, S D; Bornmann, W G et al. (1994) Bacterial lipopolysaccharide has structural similarity to ceramide and stimulates ceramide-activated protein kinase in myeloid cells. J Biol Chem 269:17606-10 |
| Jarvis, W D; Fornari Jr, F A; Browning, J L et al. (1994) Attenuation of ceramide-induced apoptosis by diglyceride in human myeloid leukemia cells. J Biol Chem 269:31685-92 |
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