Abstract
Tumor necrosis factor-α (TNF-α) is one of the most pleiotropic of cytokines, acting as a host defense factor in myriad immunological and inflammatory responses and antitumor activity (1–3). The cytotoxic effects of TNF-α are primarily mediated through TNF-R1 and the receptor-associated proteins, TNF-R1-associated death domain protein (TRADD) and Fas-associated death domain (FADD/MORT1) (3–5). Ceramide generation and caspase activation represent potential regulation points of apoptotic signaling by TNF-α (6,7). Increased ceramide formation via sphingomyelinase represents an early event in the apoptotic cascade of TNF-α (6,8,9). In MCF-7 breast cancer cells, ceramide is one of the essential molecules in TNF-α-induced apoptosis (10–12). Increased ceramide generation induced by the P-glycoprotein blocker PSC 833, restores TNF-α-induced apoptosis in KG1a leukemia cells (13), whereas loss of ceramide production is a cause of cellular resistance to apoptosis induced by TNF-α (14). Glucosylceramide synthase (GCS), converting ceramide into glucosylceramide (15), exerts strong influence over cellular ceramide levels and is thus a major modulator of programmed cell death (16). Introducing the GCS gene into TNF-α-sensitive MCF-7 cells greatly diminishes cellular response to TNF-α cytotoxicity (12).
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Aggarwal, B. B. and Natarajan, K. (1996) Tumor necrosis factors: developments during the last decade. Eur. Cytokine Netw. 7, 93–124.
Fiers, W. (1991) Tumor necrosis factor. Characterization at the molecular, cellular and in vivo level. FEBS Lett. 285, 199–212.
Larrick, J. W. and Wright, S. C. (1990) Cytotoxic mechanism of tumor necrosis factor-α. FASEB J. 4, 3215–3223.
Vandenabeele, P., Declercq, W., Beyaert, R., and Fiers, T. (1995) Two tumor necrosis factor receptors: structure and function. Trends Cell Biol. 5, 392–399.
Nagata, S. (1997) Apoptosis by death factor. Cell 88, 355–365.
Hannun, Y. A. (1996) Function of ceramide in coordinating cellular responses to stress. Science, 274, 1855–1859.
Nicholson, D. W. and Thormberry, N. A. (1997) Caspase: killer proteases. Trends Biochem. Sci. 22, 299–306.
Pena, L. A., Fuks, Z., and Kolesnick, R. (1997) Stress-induced apoptosis and the sphingomyelin pathways. Biochem. Pharmacol. 53, 615–621.
Wiegmann, K., Schutze, S., Machchleidt, T., Witte, D., and Kronke, M. (1994) Functional dichotomy of neutral and acidic sphingomyelinase in tumor necrosis factor signaling. Cell 78, 1005–1015.
Cai, Z., Bettaieb, A., Mahdani, N. E., et al. (1997) Alteration of the sphingomyelin/ceramide pathway is associated with resistance of human breast carcinoma MCF7 cells to tumor necrosis factor-α-mediated cytotoxicity. J. Biol. Chem. 272, 6918–6926.
Burow, M. E., Weldon, C. B., Tang, Y., et al. (1998) Differences in susceptibility to tumor necrosis factor-a induced apoptosis among MCF-7 breast cancer cell variants. Cancer Res. 58, 4940–4946.
Liu, Y. Y., Han, T. Y., Giuliano, A. E., Ichikawa, S., Hirabayashi, Y., and Cabot, M. C. (1999) Glycosylation of ceramide potentiates cellular resistance to tumor necrosis factor-α-induced apoptosis. Exp. Cell Res. 252, 464–470.
Bezombes, C., Maestre, N., Laurent, G., Levade, T., Bettaieb, A., and Jaffrezou J. P. (1998) Restoration of TNF-α-induced ceramide generation and apoptosis in resistant human leukemia KG1a cells by the P-glycoprotein blocker PSC 833. FASEB J. 12, 101–109.
Zyad A., Benard J., Tursz T., Clarke R., and Chouaib S. (1994) Resistance to TNF-α and adriamycin in the human breast cancer MCF-7 cell line: relationship to MDR1, MnSOD and TNF gene expression. Cancer Res. 54, 825–831.
Basu, S., Kaufeman, B., and Roseman, S. (1968) Enzymatic synthesis of ceramide-glucose and ceramide-lactose by glycosyltransferases from embryonic chicken brain. J. Biol. Chem. 243, 5802–5804.
Senchenkov, A. and Cabot, M. C. (2001) Targeting ceramide metabolism-a strategy for overcoming drug resistance. J. Natl. Cancer Inst. 93, 347–357.
Goseen, M., Freundlieb, S., Bender, G., Muller, G., Hillen, W., and Bujard, H. (1995) Transcriptional activation by tetracyclines in mammalian cells. Science 268, 1766–1769.
Resnitzky, D., Gossen, M., Bujard, H., and Reed, S. I. (1994) Acceleration of the G1/S phase transition by expression of cyclins D1 and E with an inducible system. Mol. Cell Biol. 14, 1669–1679.
Gossen, M. and Bujard, H. (1992) Tight control of gene expression in mammalian cells by tetracycline-responsive promoters. Proc. Natl. Acad. Sci. USA 89, 5547–5551.
Yin, D. X., Zhu, L., and Schimke, R. T. (1996) Tetracycline-controlled gene expression system achieves high-level and quantitative control of gene expression. Anal. Biochem. 235, 195–201.
Faris, M., Kokot, N., Lee, L., and Nel, A. (1996) Regulation of interleukin-2 transcription by inducible stable expression of dominant negative and dominant active mitogen-activated protein kinase kinase kinase in Jurkat T cells. Evidence for the importance of Ras in a pathway that is controlled by dual receptor stimulation. J. Biol. Chem. 271, 27,366–27,373.
Liu, Y. Y., Han, T. Y., Giuliano, A. E., and Cabot, M. C. (1999) Expression of glucosylceramide synthase, converting ceramide to glucosylceramide confers adriamycin resistance in human breast cancer cells. J. Biol. Chem. 274, 1140–1146.
Ichikawa, S., Sakiyama, H., Suzuki, G., Hidari, K. I. P., and Hirabayashi, Y. (1996) Expression cloning of a cDNA for human ceramide glucosyltransferase that catalyzes the first glycosylation step of glycosphingolipid synthesis. Proc. Natl. Acad. Sci. USA. 93, 4638–4643.
Liu, Y. Y., Han, T. Y., Giuliano, A. E., Hansen, R., and Cabot, M. C. (2000) Uncoupling ceramide glycosylation by transfection of glucosylceramide synthase antisense reverses adriamycin resistance. J. Biol. Chem. 275, 7138–7143.
Zhu, H. J., Iaria, J., and Sizeland, A. M. (1999) Smad7 differentially regulates transforming growth factor b-mediated signaling pathways. J. Biol. Chem. 274, 32,258–32,264.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2004 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Liu, YY., Cabot, M.C. (2004). Development of a Mammalian Tet-On Expression Cell Line. In: De Ley, M. (eds) Cytokine Protocols. Methods in Molecular Biology, vol 249. Humana Press. https://doi.org/10.1385/1-59259-667-3:177
Download citation
DOI: https://doi.org/10.1385/1-59259-667-3:177
Publisher Name: Humana Press
Print ISBN: 978-0-89603-948-3
Online ISBN: 978-1-59259-667-6
eBook Packages: Springer Protocols