Inhibition of Receptor Internalization by Monodansylcadaverine Selectively Blocks p55 Tumor Necrosis Factor Receptor Death Domain Signaling
1999; Elsevier BV; Volume: 274; Issue: 15 Linguagem: Inglês
10.1074/jbc.274.15.10203
ISSN1083-351X
AutoresStefan Schütze, Thomas Machleidt, Dieter Adam, Ralf Schwandner, Katja Wiegmann, Marie‐Luise Kruse, Michael Heinrich, Marc Wickel, Martin Krönke,
Tópico(s)Immune Response and Inflammation
ResumoThe 55-kDa receptor for tumor necrosis factor (TR55) triggers multiple signaling cascades initiated by adapter proteins like TRADD and FAN. By use of the primary amine monodansylcadaverine (MDC), we addressed the functional role of tumor necrosis factor (TNF) receptor internalization for intracellular signal distribution. We show that MDC does not prevent the interaction of the p55 TNF receptor (TR55) with FAN and TRADD. Furthermore, the activation of plasmamembrane-associated neutral sphingomyelinase activation as well as the stimulation of proline-directed protein kinases were not affected in MDC-treated cells. In contrast, activation of signaling enzymes that are linked to the “death domain” of TR55, like acid sphingomyelinase and c-Jun-N-terminal protein kinase as well as TNF signaling of apoptosis in U937 and L929 cells, are blocked in the presence of MDC. The results of our study suggest a role of TR55 internalization for the activation of select TR55 death domain signaling pathways including those leading to apoptosis. The 55-kDa receptor for tumor necrosis factor (TR55) triggers multiple signaling cascades initiated by adapter proteins like TRADD and FAN. By use of the primary amine monodansylcadaverine (MDC), we addressed the functional role of tumor necrosis factor (TNF) receptor internalization for intracellular signal distribution. We show that MDC does not prevent the interaction of the p55 TNF receptor (TR55) with FAN and TRADD. Furthermore, the activation of plasmamembrane-associated neutral sphingomyelinase activation as well as the stimulation of proline-directed protein kinases were not affected in MDC-treated cells. In contrast, activation of signaling enzymes that are linked to the “death domain” of TR55, like acid sphingomyelinase and c-Jun-N-terminal protein kinase as well as TNF signaling of apoptosis in U937 and L929 cells, are blocked in the presence of MDC. The results of our study suggest a role of TR55 internalization for the activation of select TR55 death domain signaling pathways including those leading to apoptosis. tumor necrosis factor neutral sphingomyelinase acid SMase N-hexanoylsphingosine c-Jun N-terminal kinase monodansylcadaverine 1-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide proline-directed protein kinase p55 TNF receptor gluthathione S-transferase phosphate-buffered saline fluorescein isothiocyanate interleukin Tumor necrosis factor (TNF),1 originally defined by its antitumoral activity, is now recognized as a pleiotropic cytokine exerting a wide variety of immunoregulatory activities (for review, see Refs. 1Goeddel D.V. Aggarwal B.B. Gray P.W. Leung D.W. Nedwin G.E. Palladino M.A. Patton J.S. Pennica D. Shepard H.M. Sugarman B.J. Wong G.H.W. Cold Spring Harbor Symp. Quant. Biol. 1986; 51: 597-609Crossref PubMed Google Scholar, 2Beutler B. Cerami A. Nature. 1986; 320: 584-588Crossref PubMed Scopus (1293) Google Scholar, 3Fiers W. FEBS Lett. 1991; 285: 199-212Crossref PubMed Scopus (677) Google Scholar). TNF action is mediated by two types of cell surface receptors of 55 kDa (TR55) and 75 kDa (TR75) molecular masses, respectively. Both receptors mediate distinct TNF responses (4Tartaglia L.A. Goeddel D.V. Reynolds C. Figari I.S. Weber R.F. Fendly B.M. Palladino Jr., M.A. J. Immunol. 1993; 151: 4637-4641PubMed Google Scholar, 5Gehr F. Gentz R. Brockhaus M. Loetscher H. Lesslauer W. J. Immunol. 1992; 149: 911-917PubMed Google Scholar, 6Grell M. Zimmermann G. Hulser D. Pfizenmaier K. Scheurich P. J. Immunol. 1994; 153: 1963-1972PubMed Google Scholar). The majority of activities of soluble TNF appears to be mediated by TR55 (7Tartaglia L.A. Rothe M. Hu Y.F. Goeddel D.V. Cell. 1993; 73: 213-216Abstract Full Text PDF PubMed Scopus (301) Google Scholar, 8Wiegmann K. Schütze S. Kampen E. Himmler A. Machleidt T. Krönke M. J. Biol. Chem. 1992; 267: 17997-18001Abstract Full Text PDF PubMed Google Scholar, 9Wiegmann K. Schütze S. Machleidt T. Witte D. Krönke M. Cell. 1994; 78: 1005-1015Abstract Full Text PDF PubMed Scopus (673) Google Scholar, 10Pfeffer K. Matsuyama T. Kündig T.M. Wakeham A. Kishihara K. Shahinian A. Wiegmann K. Ohasi P.S. Krönke M. Mak T.W. Cell. 1993; 73: 457-467Abstract Full Text PDF PubMed Scopus (1532) Google Scholar, 11Aggarwal B.B Natarajan K. Eur. Cytokine Netw. 1996; 7: 93-124PubMed Google Scholar). Like other cytokine receptors, the cytoplasmic domain of both TNF receptors lacks intrinsic enzymatic activities. The activation of intracellular signaling enzyme systems is initiated by a selective interplay between the cytoplasmic domain of the TNF receptors and a number of recently identified TNF receptor-associated proteins (see Ref.12Darnay B.G. Aggarwal B.B. J. Leukocyte Biol. 1997; 61: 559-566Crossref PubMed Scopus (168) Google Scholar for review). The adapter protein TRADD associates with the so-called death domain of TR55, recruiting FADD, RIP, and TRAF-2 (13Chinnaiyan A.M. Tepper C.G. Seldin M.F. O'Rourke K. Kischkel F.C. Hellbardt S. Krammer P.H. Peter M.E. Dixit V.M. J. Biol. Chem. 1996; 271: 4961-4965Abstract Full Text Full Text PDF PubMed Scopus (705) Google Scholar, 14Stanger B.Z. Leder P. Lee T.H. Kim E. Seed B. Cell. 1995; 81: 513-523Abstract Full Text PDF PubMed Scopus (860) Google Scholar, 15Hsu H. Huang J. Shu H.B. Baichwal V. Goeddel D.V. Immunity. 1996; 4: 387-396Abstract Full Text Full Text PDF PubMed Scopus (972) Google Scholar, 16Rothe M. Sarma V. Dixit V.M. Goeddel D.V. Science. 1995; 269: 1424-1427Crossref PubMed Scopus (972) Google Scholar). FADD mediates the activation of a protease termed FLICE/MACH (17Boldin M.P. Goncharov T.M. Goltsev Y.V. Wallach D. Cell. 1996; 85: 803-815Abstract Full Text Full Text PDF PubMed Scopus (2100) Google Scholar, 18Muzino M. Chinnaiyan A.M. Kischkel F.C. O'Rourke K. Shevchenko A. Ni J. Scaffidi C. Bretz J.D. Zhang M. Gentz R. Mann M. Krammer P.H. Peter M.E. Dixit V.M. Cell. 1996; 85: 817-827Abstract Full Text Full Text PDF PubMed Scopus (2723) Google Scholar), representing one of the first steps in induction of the apoptotic pathway. RIP mediates activation of the transcription factor NF-κB associated with anti-apoptotic regulatory functions (19Beg A.A. Baltimore D. Science. 1996; 274: 782-784Crossref PubMed Scopus (2926) Google Scholar). TRAF-2 links TR55 to activation of the N-terminal c-Jun kinase (JNK) cascade (20Liu Z.G. Hsu H. Goeddel D.V. Karin M. Cell. 1996; 87: 565-576Abstract Full Text Full Text PDF PubMed Scopus (1778) Google Scholar). The TR55-associated protein FAN (21Adam-Klages S. Adam D. Wiegmann K. Struve S. Kolanus W. Schneider-Mergener J. Krönke M. Cell. 1996; 86: 937-947Abstract Full Text Full Text PDF PubMed Scopus (356) Google Scholar) binds to a domain designated neutral sphingomyelinase activation domain (NSD) that is N-terminally adjacent to the death domain (22Adam D. Wiegmann K. Adam-Klages S. Ruff A. Krönke M. J. Biol. Chem. 1996; 271: 14617-14622Abstract Full Text Full Text PDF PubMed Scopus (135) Google Scholar). FAN mediates TNF-induced activation of neutral sphingomyelinase (N-SMase) (21Adam-Klages S. Adam D. Wiegmann K. Struve S. Kolanus W. Schneider-Mergener J. Krönke M. Cell. 1996; 86: 937-947Abstract Full Text Full Text PDF PubMed Scopus (356) Google Scholar).Results from numerous studies have revealed that TNF signaling further involves activation of downstream enzyme systems at multiple subcellular compartments such as the plasmamembrane, endosomes, mitochondria, the cytosol, and the nucleus (for review, see Refs. 11Aggarwal B.B Natarajan K. Eur. Cytokine Netw. 1996; 7: 93-124PubMed Google Scholar,23Heller R.A. Krönke M. J. Cell Biol. 1994; 126: 5-9Crossref PubMed Scopus (401) Google Scholar, and 24Beyaert R. Fiers W. FEBS Lett. 1994; 340: 9-16Crossref PubMed Scopus (243) Google Scholar). Membrane-associated enzyme systems transmitting TR55 signals include plasmamembrane-bound phospholipases such as phosphatidylcholine-specific phospholipase C (25Schütze S. Berkovic D. Tomsing O. Unger C. Krönke M. J. Exp. Med. 1991; 174: 975-988Crossref PubMed Scopus (181) Google Scholar), which generates the lipid second messenger molecule 1,2-diacylglycerol and a N-SMase (9Wiegmann K. Schütze S. Machleidt T. Witte D. Krönke M. Cell. 1994; 78: 1005-1015Abstract Full Text PDF PubMed Scopus (673) Google Scholar), producing ceramide by sphingomyelin hydrolysis. Ceramide generated at the plasmamembrane triggers activation of a 97-kDa ceramide-activated protein kinase (26Liu J. Mathias S. Yang Z. Kolesnick R.N. J. Biol. Chem. 1994; 269: 3047-3052Abstract Full Text PDF PubMed Google Scholar), recently suggested to be identical with the “kinase suppressor of ras” (27Zhang Y. Yao B. Delikat S. Bayoumy S. Lin X.-H. Basu S. McGinley M. Chan-Hui P.-Y. Lichenstein H. Kolesnick R. Cell. 1997; 89: 63-72Abstract Full Text Full Text PDF PubMed Scopus (390) Google Scholar). Ceramide-activated protein kinase belongs to a family of proline-directed protein kinases (PDPK) (9Wiegmann K. Schütze S. Machleidt T. Witte D. Krönke M. Cell. 1994; 78: 1005-1015Abstract Full Text PDF PubMed Scopus (673) Google Scholar), including members of the mitogen-activated protein kinases (28Vietor I. Schwenger P. Li W. Schlessinger J. Vilcek J. J. Biol. Chem. 1993; 268: 18994-18999Abstract Full Text PDF PubMed Google Scholar). TNF signaling further involves intracellular membrane compartments like caveolae and endosomes harboring an acid SMase (A-SMase) (9Wiegmann K. Schütze S. Machleidt T. Witte D. Krönke M. Cell. 1994; 78: 1005-1015Abstract Full Text PDF PubMed Scopus (673) Google Scholar, 29Schütze S. Potthoff K. Machleidt T. Berkovic D. Wiegmann K. Krönke M. Cell. 1992; 71: 765-776Abstract Full Text PDF PubMed Scopus (968) Google Scholar, 30Liu P. Anderson R.G. J. Biol. Chem. 1995; 270: 27179-27185Abstract Full Text Full Text PDF PubMed Scopus (301) Google Scholar). In mitochondria, TNF induces reactive oxygen species that are generated at the level of the oxidative phosphorylation complex III (31Schulze Osthoff K. Beyaert R. Vandevoorde V. Haegeman G. Fiers W. EMBO J. 1993; 12: 3095-3104Crossref PubMed Scopus (548) Google Scholar). The induction of the mitochondrial permeability transition has been linked to the TNF cytotoxic pathway (32Pastorino J.G. Simbula G. Yamamoto K. Glascott P.A. Rothman R.J. Farber J.L. J. Biol. Chem. 1996; 271: 29792-29798Abstract Full Text Full Text PDF PubMed Scopus (254) Google Scholar). In addition, cytosolic protein kinase cascades including PKC and JNK have been identified to transmit TNF signals (33Schütze S. Nottrott S. Pfizenmaier K. Krönke M. J. Immunol. 1990; 144: 2604-2608PubMed Google Scholar, 34Sluss H.K. Barrett T. Derijard B. Davis R.J. Mol. Cell. Biol. 1994; 14: 8376-8384Crossref PubMed Scopus (444) Google Scholar, 35Adam D. Ruff A. Strelow A. Wiegmann K. Krönke M. Biochem. J. 1998; 333: 343-350Crossref PubMed Scopus (23) Google Scholar).The question of how TNF signals are targeted to the different intracellular compartments is of fundamental biological significance. Intriguingly, rapidly diffusible ions like Ca2+ seem not to be involved in TNF signaling. Rather, the two lipid second messenger molecules 1,2-diacylglycerol and ceramide are likely to reside within membranes because of their hydrophobic nature. Thus the mechanisms of intracellular TNF signal trafficking remain obscure.In the present study we investigated the effects of inhibition of TNF receptor endocytosis on TNF signal transduction. Because structural motifs within the TNF receptor required for endocytosis are currently unknown, genetic approaches to block TNF receptor internalization are not available at present. We here employed the primary amine monodansylcadaverine (MDC) or K+ depletion to inhibit TNF receptor internalization. MDC is an inhibitor of transglutaminase, a membrane-bound enzyme that actively participates in internalization of various receptor systems (36Folk J.E. Chung S.H. Adv. Enzymol. 1973; 38: 109-191PubMed Google Scholar, 37Davies P. Davis D. Levitzki F. Maxfield P. Milhaud M. Willingham C. Pastan I. Nature. 1980; 283: 162-167Crossref PubMed Scopus (409) Google Scholar, 38Chow J.C. Condorelli G. Smith R.J. J. Biol. Chem. 1998; 273: 4672-4680Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar, 39Resink T.J. Scott Burden T. Boulanger C. Weber E. Buhler F.R. Mol. Pharmacol. 1990; 38: 244-252PubMed Google Scholar, 40Samanta A.K. Oppenheim J.J. Matsushima K. J. Biol. Chem. 1990; 265: 183-189Abstract Full Text PDF PubMed Google Scholar, 41Bradley J.R. Johnson D.R. Pober J.S. J. Immunol. 1993; 12: 5544-5555Google Scholar, 42Peavy D.E. Edmondson J.W. Duckworth W.C. Endocrinology. 1984; 114: 753-760Crossref PubMed Scopus (54) Google Scholar, 43Ray E. Samanta A.K. FEBS Lett. 1996; 378: 235-239Crossref PubMed Scopus (35) Google Scholar, 44Ray E. Samanta A.K. Cytokine. 1997; 9: 587-596Crossref PubMed Scopus (32) Google Scholar).We show here that MDC as well as K+-depletion block TNF endocytosis, which is related to inhibition of TR55 death domain signaling like TNF-dependent activation of endosomal A-SMase, JNK, and TNF-mediated apoptosis. In contrast, TNF-dependent stimulation of plasmamembrane-associated N-SMase and PDP kinase are not affected by MDC or K+ depletion.DISCUSSIONIn the present paper the role of receptor internalization for TNF signaling was investigated. Intriguingly, inhibition of TR55 internalization by MDC or K+ depletion abolished select signaling events while leaving others unaffected. In particular, TNF-induced activation of A-SMase, JNK, and the induction of apoptosis were sensitive to MDC. In contrast, MDC did not inhibit TNF-induced activation of N-SMase and PDP kinases. Similar results were obtained with K+-depletion, which inhibited TNF receptor internalization and, like MDC, selectively blocked TNF-induced activation of A-SMase and JNK but not N-SMase or PDP kinases.MDC has been extensively used to block endocytosis and trafficking of various ligand-receptor systems. MDC is known as a potent competitive inhibitor of transglutaminase (36Folk J.E. Chung S.H. Adv. Enzymol. 1973; 38: 109-191PubMed Google Scholar, 37Davies P. Davis D. Levitzki F. Maxfield P. Milhaud M. Willingham C. Pastan I. Nature. 1980; 283: 162-167Crossref PubMed Scopus (409) Google Scholar, 58Haigler H.T. Maxfield F.R. Willingham M.C. Pastan I. J. Biol. Chem. 1980; 255: 1239-1241Abstract Full Text PDF PubMed Google Scholar, 59Rikihisa Y. Zhang Y. Park J. Infect. Immun. 1994; 62: 5126-5132Crossref PubMed Google Scholar). The enzymatic action of transglutaminase involves cross-linking of proteins by forming an isopeptide bond between a lysine residue of one protein and a glutamine residue of another protein during coated pit formation. MDC has been shown to block endocytosis of α2-macroglobulin and many polypeptide hormones and cytokines like epidermal growth factor, hepatocyte growth factor, insulin-like growth factor-I, and IL-8 (37Davies P. Davis D. Levitzki F. Maxfield P. Milhaud M. Willingham C. Pastan I. Nature. 1980; 283: 162-167Crossref PubMed Scopus (409) Google Scholar,38Chow J.C. Condorelli G. Smith R.J. J. Biol. Chem. 1998; 273: 4672-4680Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar, 40Samanta A.K. Oppenheim J.J. Matsushima K. J. Biol. Chem. 1990; 265: 183-189Abstract Full Text PDF PubMed Google Scholar, 43Ray E. Samanta A.K. FEBS Lett. 1996; 378: 235-239Crossref PubMed Scopus (35) Google Scholar, 44Ray E. Samanta A.K. Cytokine. 1997; 9: 587-596Crossref PubMed Scopus (32) Google Scholar, 58Haigler H.T. Maxfield F.R. Willingham M.C. Pastan I. J. Biol. Chem. 1980; 255: 1239-1241Abstract Full Text PDF PubMed Google Scholar, 60Maxfield F.R. Willingham M.C. Schlessinger J. Davies P.J.A. Pastan I. Nature. 1979; 277: 661-663Crossref PubMed Scopus (168) Google Scholar). MDC-mediated inhibition of transglutaminase has also been implicated in the blocking of endocytosis of vesicular stomatitis virus, Semliki Forest virus, and other types of endocytosis occurring through clathrin-coated vesicles (61Marsh M. Helenius A. J. Cell Biol. 1980; 142: 439-454Google Scholar, 62Schlagal R. Dickson R.B. Willingham M.C. Pastan I.H. Proc. Natl. Acad. Sci. U. S. A. 1982; 79: 2291-2295Crossref PubMed Scopus (129) Google Scholar). MDC does not affect the number or affinity of TNF receptors expressed on the cell surface (41Bradley J.R. Johnson D.R. Pober J.S. J. Immunol. 1993; 12: 5544-5555Google Scholar). 2S. Schütze, unpublished observations. MDC blocked TNF receptor internalization at concentrations similar to the concentrations required for inhibition of transglutaminase in lysates from Chinese hamster ovary cells (37Davies P. Davis D. Levitzki F. Maxfield P. Milhaud M. Willingham C. Pastan I. Nature. 1980; 283: 162-167Crossref PubMed Scopus (409) Google Scholar), suggesting that the inhibition of this enzyme is related to the MDC effects on TNF receptor endocytosis. MDC concentrations exceeding 100 μm did not further increase the inhibitory effect on TNF receptor internalization in U937 cells. These findings are consistent with a previous study by Davies and co-workers, reporting on MDC-mediated inhibition of α2-macroglobulin receptor clustering (37Davies P. Davis D. Levitzki F. Maxfield P. Milhaud M. Willingham C. Pastan I. Nature. 1980; 283: 162-167Crossref PubMed Scopus (409) Google Scholar).The differential requirement of TR55 internalization for the activation of A-SMase and N-SMase are in concordance with a previous report by Hofmeister et al. in which the authors used an independent approach to study the functional consequences of IL-1 receptor internalization (63Hofmeister R. Wiegmann C. Korherr C. Bernado K. Krönke M. Falk W. J. Biol. Chem. 1997; 272: 27730-27736Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar). An IL-1 receptor type I-positive EL4 thymoma cell line was employed, which is defective in the IL-1R accessory protein (IL-1RAcP) required for IL-1 receptor internalization. In this IL-1R internalization-defective cell line, IL-1 induction of N-SMase appeared normal, whereas the activation of A-SMase was completely impaired. Transfection with IL-1RAcP cDNA restored IL-1 receptor internalization as well as IL-1-induced A-SMase stimulation, indicating the requirement of IL-1R internalization for A-SMase activation.Our findings on the differential requirement of receptor internalization for activation of A- and N-SMase may help to explain previous data by Andrieu et al. (51Andrieu N. Salvayre R. Jaffrezou J.P. Levade T. J. Biol. Chem. 1995; 270: 24518-24524Abstract Full Text Full Text PDF PubMed Scopus (46) Google Scholar). These authors could not detect a requirement of TNF or IL-1 receptor internalization for sphingomyelin turnover. Here we show that it is only N-SMase that is insensitive to MDC or K+ depletion, whereas activation of A-SMase depends on TNF receptor internalization. Because the overall enzymatic activity of A-SMase exceeds that of N-SMase by a factors of 3–10, depending on the cell line investigated (Fig. 4 and Refs. 9Wiegmann K. Schütze S. Machleidt T. Witte D. Krönke M. Cell. 1994; 78: 1005-1015Abstract Full Text PDF PubMed Scopus (673) Google Scholar and65Boucher L.-M. Wiegmann K. Fütterer A. Pfeffer K. Machleidt T. Schütze S. Mak T.W. Krönke M. J. Exp. Med. 1995; 181: 2059-2068Crossref PubMed Scopus (190) Google Scholar), the remaining N-SMase activity might have escaped the authors' detection system.A role for receptor clustering and internalization for activation of JNK has been also suggested by Rosette and Karin (64Rosette C. Karin M. Science. 1996; 274: 1194-1197Crossref PubMed Scopus (941) Google Scholar), who report that exposure of HeLa cells to UV light or osmotic shock induced clustering and internalization of cell surface receptors for TNF, epidermal growth factor, and IL-1 in the absence of the respective ligands. UV or osmotic shock-induced activation of JNK could be inhibited by blocking receptor clustering at 10 °C or after receptor down-regulation, suggesting that multimerization and clustering of the cytokine receptors is the initial signaling event that activates the JNK cascade.Intriguingly, MDC blocked TR55-induced apoptosis. The doses of MDC required to inhibit TNF receptor endocytosis correlated well with those needed for the inhibition of TNF-induced cytotoxicity, consistent with the idea that TR55 internalization is required for apoptotic signaling. It is worth mentioning that MDC does not prevent apoptosis in a nonspecific manner. C6-ceramide, which is taken up through pinocytosis, triggered programmed cell death in the presence of MDC. In addition, CD95-mediated apoptosis of U937 and Jurkat cells was not prevented by MDC, suggesting that CD95-induced cell death is regulated by a mechanism distinct from clathrin-coated pit-mediated endocytosis.Because the effects of MDC inhibition are presumably based on a blockade of protein cross-linking mediated by transglutaminase, the question is raised of whether MDC might disrupt the interaction of TRADD with the TR55 death domain. This would result in an uncoupling of death domain signaling independent of TR55 internalization. Our results, however, clearly show that association of TRADD to the death domain of TR55 was not impaired by MDC, suggesting that inhibition of A-SMase, JNK activation, and apoptosis by MDC does not result from disrupting the TRADD/TR55 signaling complex.The importance of TNF receptor internalization for signaling cell death is controversial. A functional role of TNF receptor internalization for mediating TNF cytotoxicity has been suggested previously by Kull and Cuatrecasas (55Kull F.C. Cuatrecasas P. Cancer Res. 1981; 41: 4885-4890PubMed Google Scholar), Watanabe et al. (56Watanabe N. Kuriyama H. Sone H. Neda H. Yamauchi N. Maeda M. Niitsu Y. J. Biol. Chem. 1988; 263: 10262-10266Abstract Full Text PDF PubMed Google Scholar), and Pastorinoet al. (32Pastorino J.G. Simbula G. Yamamoto K. Glascott P.A. Rothman R.J. Farber J.L. J. Biol. Chem. 1996; 271: 29792-29798Abstract Full Text Full Text PDF PubMed Scopus (254) Google Scholar). It should be emphasized that a membrane-bound form (26 kDa) of TNF triggers cell death by direct cell-to-cell contact and, indeed, is apparently independent of TNF receptor internalization (57Grell M. Douni E. Wajant H. Lohden M. Clauss M. Maxeiner B. Georgopoulos S. Lesslauer W. Kollias G. Pfizenmaier K. Scheurich P. Cell. 1995; 83: 793-802Abstract Full Text PDF PubMed Scopus (1145) Google Scholar, 66Lazdins J.K. Grell M. Walker M.R. Woods-Cook K. Scheurich P. Pfizenmaier K. J. Exp. Med. 1997; 185: 81-90Crossref PubMed Scopus (62) Google Scholar). It is important to note, however, that membrane-bound TNF has been shown to preferentially trigger the p75 TNF receptor (TR75) (57Grell M. Douni E. Wajant H. Lohden M. Clauss M. Maxeiner B. Georgopoulos S. Lesslauer W. Kollias G. Pfizenmaier K. Scheurich P. Cell. 1995; 83: 793-802Abstract Full Text PDF PubMed Scopus (1145) Google Scholar). TNF binding to TR75 does not result in receptor internalization but rather triggers shedding of the TR75 (49Higuchi M. Aggarwal B.B. J. Immunol. 1994; 152: 3550-3558PubMed Google Scholar, 50Porteu F. Hieblot C. J. Biol. Chem. 1994; 269: 2834-2840Abstract Full Text PDF PubMed Google Scholar). Clearly, the mechanism by which TR75 signals cytotoxicity is different from the TR55 signaling pathway. Unlike membrane-bound TNF, soluble TNF binds and triggers TR55 that is readily internalized following ligand binding. Thus, the conflicting results can be explained by the engagement of distinct TNF receptors by membrane-bound and -soluble TNF, respectively.Our study implies that the functional consequences of TR55 endocytosis go beyond receptor degradation to terminate ligand-induced signaling. The results of our study indicate that TR55 internalization is crucially important for targeting and coupling of the TR55 adapter protein complexes to specific signaling systems like A-SMase, JNK, and a proapoptotic signaling cascade. TNF receptor endocytosis may play a previously unrecognized role in relaying TNF signals to intracellular compartments, where TNF receptor bearing membranes can be sensed as “signaling rafts.” Tumor necrosis factor (TNF),1 originally defined by its antitumoral activity, is now recognized as a pleiotropic cytokine exerting a wide variety of immunoregulatory activities (for review, see Refs. 1Goeddel D.V. Aggarwal B.B. Gray P.W. Leung D.W. Nedwin G.E. Palladino M.A. Patton J.S. Pennica D. Shepard H.M. Sugarman B.J. Wong G.H.W. Cold Spring Harbor Symp. Quant. Biol. 1986; 51: 597-609Crossref PubMed Google Scholar, 2Beutler B. Cerami A. Nature. 1986; 320: 584-588Crossref PubMed Scopus (1293) Google Scholar, 3Fiers W. FEBS Lett. 1991; 285: 199-212Crossref PubMed Scopus (677) Google Scholar). TNF action is mediated by two types of cell surface receptors of 55 kDa (TR55) and 75 kDa (TR75) molecular masses, respectively. Both receptors mediate distinct TNF responses (4Tartaglia L.A. Goeddel D.V. Reynolds C. Figari I.S. Weber R.F. Fendly B.M. Palladino Jr., M.A. J. Immunol. 1993; 151: 4637-4641PubMed Google Scholar, 5Gehr F. Gentz R. Brockhaus M. Loetscher H. Lesslauer W. J. Immunol. 1992; 149: 911-917PubMed Google Scholar, 6Grell M. Zimmermann G. Hulser D. Pfizenmaier K. Scheurich P. J. Immunol. 1994; 153: 1963-1972PubMed Google Scholar). The majority of activities of soluble TNF appears to be mediated by TR55 (7Tartaglia L.A. Rothe M. Hu Y.F. Goeddel D.V. Cell. 1993; 73: 213-216Abstract Full Text PDF PubMed Scopus (301) Google Scholar, 8Wiegmann K. Schütze S. Kampen E. Himmler A. Machleidt T. Krönke M. J. Biol. Chem. 1992; 267: 17997-18001Abstract Full Text PDF PubMed Google Scholar, 9Wiegmann K. Schütze S. Machleidt T. Witte D. Krönke M. Cell. 1994; 78: 1005-1015Abstract Full Text PDF PubMed Scopus (673) Google Scholar, 10Pfeffer K. Matsuyama T. Kündig T.M. Wakeham A. Kishihara K. Shahinian A. Wiegmann K. Ohasi P.S. Krönke M. Mak T.W. Cell. 1993; 73: 457-467Abstract Full Text PDF PubMed Scopus (1532) Google Scholar, 11Aggarwal B.B Natarajan K. Eur. Cytokine Netw. 1996; 7: 93-124PubMed Google Scholar). Like other cytokine receptors, the cytoplasmic domain of both TNF receptors lacks intrinsic enzymatic activities. The activation of intracellular signaling enzyme systems is initiated by a selective interplay between the cytoplasmic domain of the TNF receptors and a number of recently identified TNF receptor-associated proteins (see Ref.12Darnay B.G. Aggarwal B.B. J. Leukocyte Biol. 1997; 61: 559-566Crossref PubMed Scopus (168) Google Scholar for review). The adapter protein TRADD associates with the so-called death domain of TR55, recruiting FADD, RIP, and TRAF-2 (13Chinnaiyan A.M. Tepper C.G. Seldin M.F. O'Rourke K. Kischkel F.C. Hellbardt S. Krammer P.H. Peter M.E. Dixit V.M. J. Biol. Chem. 1996; 271: 4961-4965Abstract Full Text Full Text PDF PubMed Scopus (705) Google Scholar, 14Stanger B.Z. Leder P. Lee T.H. Kim E. Seed B. Cell. 1995; 81: 513-523Abstract Full Text PDF PubMed Scopus (860) Google Scholar, 15Hsu H. Huang J. Shu H.B. Baichwal V. Goeddel D.V. Immunity. 1996; 4: 387-396Abstract Full Text Full Text PDF PubMed Scopus (972) Google Scholar, 16Rothe M. Sarma V. Dixit V.M. Goeddel D.V. Science. 1995; 269: 1424-1427Crossref PubMed Scopus (972) Google Scholar). FADD mediates the activation of a protease termed FLICE/MACH (17Boldin M.P. Goncharov T.M. Goltsev Y.V. Wallach D. Cell. 1996; 85: 803-815Abstract Full Text Full Text PDF PubMed Scopus (2100) Google Scholar, 18Muzino M. Chinnaiyan A.M. Kischkel F.C. O'Rourke K. Shevchenko A. Ni J. Scaffidi C. Bretz J.D. Zhang M. Gentz R. Mann M. Krammer P.H. Peter M.E. Dixit V.M. Cell. 1996; 85: 817-827Abstract Full Text Full Text PDF PubMed Scopus (2723) Google Scholar), representing one of the first steps in induction of the apoptotic pathway. RIP mediates activation of the transcription factor NF-κB associated with anti-apoptotic regulatory functions (19Beg A.A. Baltimore D. Science. 1996; 274: 782-784Crossref PubMed Scopus (2926) Google Scholar). TRAF-2 links TR55 to activation of the N-terminal c-Jun kinase (JNK) cascade (20Liu Z.G. Hsu H. Goeddel D.V. Karin M. Cell. 1996; 87: 565-576Abstract Full Text Full Text PDF PubMed Scopus (1778) Google Scholar). The TR55-associated protein FAN (21Adam-Klages S. Adam D. Wiegmann K. Struve S. Kolanus W. Schneider-Mergener J. Krönke M. Cell. 1996; 86: 937-947Abstract Full Text Full Text PDF PubMed Scopus (356) Google Scholar) binds to a domain designated neutral sphingomyelinase activation domain (NSD) that is N-terminally adjacent to the death domain (22Adam D. Wiegmann K. Adam-Klages S. Ruff A. Krönke M. J. Biol. Chem. 1996; 271: 14617-14622Abstract Full Text Full Text PDF PubMed Scopus (135) Google Scholar). FAN mediates TNF-induced activation of neutral sphingomyelinase (N-SMase) (21Adam-Klages S. Adam D. Wiegmann K. Struve S. Kolanus W. Schneider-Mergener J. Krönke M. Cell. 1996; 86: 937-947Abstract Full Text Full Text PDF PubMed Scopus (356) Google Scholar). Results from numerous studies have revealed that TNF signaling further involves activation of downstream enzyme systems at multiple subcellular compartments such as the plasmamembrane, endosomes, mitochondria, the cytosol, and the nucleus (for review, see Refs. 11Aggarwal B.B Natarajan K. Eur. Cytokine Netw. 1996; 7: 93-124PubMed Google Scholar,23Heller R.A. Krönke M. J. Cell Biol. 1994; 126: 5-9Crossref PubMed Scopus (401) Google Scholar, and 24Beyaert R. Fiers W. FEBS Lett. 1994; 340: 9-16Crossref PubMed Scopus (243) Google Scholar). Membrane-associated enzyme systems transmitting TR55 signals include plasmamembrane-bound phospholipases such as phosphatidylcholine-specific phospholipase C (25Schütze S. Berkovic D. Tomsing O. Unger C. Krönke M. J. Exp. Med. 1991; 174: 975-988Crossref PubMed Scopus (181) Google Scholar), which
Referência(s)