Comparison of the intoxication pathways of tumor necrosis factor and diphtheria toxin

Abstract

The mechanism by which tumor necrosis factor alpha (TNF) initiates tumor cell destruction is unknown.Having established that a brief drop in extracellular pH enhances the killing activity of TNF, our next objective was to explore whether TNF-induced cell death is dependent on endosomal acidification. Diphtheria toxin (DTx), a well-characterized acid-dependent cytotoxin, served as an indicator of the effectiveness of each treatment condition. Studies with lysosomotropic agents demonstrated that the cytotoxic pathway of TNF can operate independently of low pH exposure in contrast to the lethal pathway of DTx. When NH4CI-treated cells were exposed to TNF at low pH, the level of killing increased twoto threefold over that attained with cells maintained at neutral pH (either with or without NH4Cl). Furthermore, inhibition of metabolic processes by sodium azide in combination with 2-deoxyglucose severely reduced DTx killing but stimulated TNF killing. Despite these differences, TNF and DTx provoked extensive internucleosomal DNA cleavage in prelytic target cells. Inhibitors of nuclear poly(ADP-ribose) transferase also evoked similar levels of cellular resistance to both cytotoxins. Models for DTx-and TNF-induced cytolysis are discussed in view of these new discoveries.

Induction of α-Helix in the β-Sheet Protein Tumor Necrosis Factor-α: Acid-Induced Denaturation

“…This might be physiologically relevant, as a pH of about 3.6 has been reported in the space between macrophages and their substrates (Silver et al, 1988). The R-helical monomer might also be capable of passing through the cell membrane and into the cytosol itself, where the change in pH would result in a return to the β-sheet conformation, consistent with the model of Chang and Wisnieski (1990). It is also possible that upon interaction with membrane lipids the acidic molten globular structure may be altered to form a channel-like structure as observed by Baldwin et al (1996).…”

“…Under acid conditions, TNF-R has been shown to insert into membranes and liposomes and is capable of functioning as an ion channel (Yoshimura & Sone, 1987Baldwin et al, 1988Baldwin et al, , 1996. The formation of R-helix at this pH could facilitate some of these changes, as well as the change in surface hydrophobicity also reported (Chang & Wisnieski, 1990;Chang et al, 1988;Kazan et al, 1992). This might be physiologically relevant, as a pH of about 3.6 has been reported in the space between macrophages and their substrates (Silver et al, 1988).…”

“…Tumor necrosis factor-R (TNF-R), 1 first observed by Carswell et al (1975), plays an important role as a mediator of inflammation and the immune response (Ming et al, 1987;Sherry & Cerami, 1988). TNF-R and lymphotoxin, a related protein, have been shown to undergo an acid-induced change in conformation which results in an increase in surface hydrophobicity and a simultaneous increase in cytolytic activity, membrane insertion, and lipid vesicle binding (Yoshimura & Sone, 1987Baldwin et al, 1988;Chang & Wisnieski, 1990). The inserted proteins appear to be capable of functioning as an ion channel (Kazan et al, 1992;Baldwin et al, 1995).…”

“…The inserted proteins appear to be capable of functioning as an ion channel (Kazan et al, 1992;Baldwin et al, 1995). These data have led to a hypothesis that the increased cytolytic activity seen at low pH is due to an acid-induced conformational change which allows the TNF-R to insert into lipid membranes and function as a Na + channel; this new conformation may also allow the TNF-R to penetrate directly into the cell, where neutral pH in the cytosol restores the original structure and activity (Kazan et al, 1992;Chang & Wisnieski, 1990). TNF-R is a β-sheet protein which self-associates into a trimer Davis et al, 1987;Narhi et al, 1987;Lewitt-Bentley et al, 1988).…”

A novel hybrid aspirin-NO-releasing compound inhibits TNFalpha release from LPS-activated human monocytes and macrophages

“…TNFα is secreted by monocytes, macrophages and neutrophils following their stimulation by bacterial LPS. The various activities of TNFα include mediation of cell adhesion molecules [ 21 ], regulation of cell death in tumour cells [ 22 ], enhancement of neutrophil responsiveness [ 23 ], control of neutrophil adherence to the endothelium [ 21 ] and synthesis of IL-1 production by macrophages [ 24 ]. LPS is known to stimulate TNFα production via the activation of the transcription factor, NF-κB [ 25 - 27 ].…”