Receptor-mediated endocytosis of diphtheria toxin by cells in culture.

Keen, James H.; Maxfield, Frederick R.; Hardegree, M. C.; Habig, William H.

doi:10.1073/pnas.79.9.2912

Cited by 86 publications

(47 citation statements)

References 31 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Therefore, we could not show any direct evidence on location of the inhibitor. However, Keen et al (16) observed that fluorescently labeled DT binds to and is internalized into mouse cells when the toxin is added at relatively high concentrations, indicating that binding sites for DT are present on the surface of DT-insensitive cells. Weiss and Mayhew (23,47) reported that RNA is a structural component of the cell surface membrane based on observations made on the mobility of cells after RNase treatment and electrophoresis.…”

Section: Discussionmentioning

confidence: 99%

“…The nicked form of CRM197 was prepared by treatment with trypsin (7). Fragment A of diphtheria toxin was purified from the culture fluid of the C7 (15)(16)(17)(18)(19)(20)(21)(22) strain (45).…”

Section: Dt and Related Proteinsmentioning

confidence: 99%

“…However, the specific binding of labeled DT to isolated membrane has not been observed as yet. When binding studies were performed with labeled toxin and an isolated membrane fraction (2) or with intact cells at relative high toxin concentrations (16), a significant association of toxin with DT-insensitive cells was observed, and the amount of toxin associated with cells was similar for DT-insensitive and DT-sensitive cells. These results have led to the interpretation that DT-insensitive cells also bear DT receptors.…”

mentioning

confidence: 99%

See 2 more Smart Citations

Identification of diphtheria toxin receptor and a nonproteinous diphtheria toxin-binding molecule in Vero cell membrane.

Mekada

Okada

Uchida

1988

The Journal of Cell Biology

View full text Add to dashboard Cite

Abstract. Two substances possessing the ability to bind to diphtheria toxin (DT) were found to be present in a membrane fraction from DT-sensitive Vero cells. One of these substances was found on the basis of its ability to bind DT and inhibit its cytotoxic effect. This inhibitory substance competitively inhibited the binding of DT to Veto cells. However this inhibitor could not bind to CRM197, the product of a missense mutation in the DT gene, and did not inhibit the binding of CRM197 to Vero cells. Moreover, similar levels of the inhibitory activity were observed in membrane fractions from DT-insensitive mouse cells, suggesting the inhibitor is not the DT receptor which is specifically present in DT-sensitive cells. The second DT-binding substance was found in the same Vero cell membrane preparation by assaying the binding of ~25I-labeled CRM197. Such DT-binding activity could not be observed in membrane preparation from mouse L cells. From competition studies using labeled DT and CRM proteins, we conclude that this binding activity is due to the surface receptor for DT. Treatment of these substances with several enzymes revealed that the inhibitor was sensitive to certain RNases but resistant to proteases, whereas the DT receptor was resistant to RNase but sensitive to proteases. The receptor was solubilized and partially purified by chromatography on CM-Sepharose column. Immunoprecipitation and Western blotting analysis of the partially purified receptor revealed that a 14.5-kD protein is the DT receptor, or at least a component of it.D PHTHERIA toxin (DT) ~ is a cytotoxic protein which inhibits cellular protein synthesis (4, 40) in eukaryotes by catalyzing the ADP-ribosylation of EF-2, which results in its inactivation (9, 12). The first step of intoxication by DT is binding of the toxin to a susceptible cell. A specific receptor for DT is believed to be involved in this step (13,43). Cells from a number of mammals including humans and monkeys are sensitive to DT, but mouse and rat cells are insensitive (29). Several lines of evidence show that the difference in sensitivity to DT between species is primarily determined by the presence or absence of a cell surface receptor (18,28,31,48). However, this receptor has not been isolated. The toxins bound to cell surface are then internalized by endocytosis (33), and the toxins, or at least their A fragments, enter the cytoplasm to exert its effects. Like in the case of Semliki Forest virus (SFV) (11), intravesicular low pH is required for the penetration of the toxin into cytoplasm (6,17,20,27,36).Some information has been obtained on the biochemical properties of DT receptor. The treatment of DT-sensitive cells with some proteases or phospholipase C reduces the sensitivity to DT (32), whereas treatment with neuraminidase increases the sensitivity (26). Although these studies 1. Abbreviations used in this paper: CRMs, cross reacting materials related to diphtheria toxin; DT, diphtheria toxin.give a clue to the chemical nature of DT receptor, the possibility that ...

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Dt and Related Proteinsmentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

Identification of diphtheria toxin receptor and a nonproteinous diphtheria toxin-binding molecule in Vero cell membrane.

Mekada

Okada

Uchida

1988

The Journal of Cell Biology

View full text Add to dashboard Cite

show abstract

“…All are resistant due to some defect in the process of internalization of diphtheria toxin, and a purpose of our investigation was to ascertain the nature of these defects, i.e., whether they were related to reduced capacity to bind the toxin or to alterations at other steps in the entry of toxin. In the case of cultured mouse L cells, resistance has been attributed either to the lack of high-affinity toxin receptors (33,41,42) or to the lack of an efficient mechanism for transport of diphtherial fragment A into the cytosol (5,6,18,23 Radiolabeling of toxin. Diphtheria toxin was labeled with radioactive iodine by a modification of the method of Marchalonis (27).…”

mentioning

confidence: 99%

Binding and uptake of diphtheria toxin by toxin-resistant Chinese hamster ovary and mouse cells.

Didsbury¹,

Moehring²,

Moehring³

1983

Mol. Cell. Biol.

View full text Add to dashboard Cite

We investigated two phenotypically distinct types of diphtheria toxin-resistant mutants of Chinese hamster cells and compared their resistance with that of naturally resistant mouse cells. All are resistant due to a defect in the process of internalization and delivery of toxin to its target in the cytosol, elongation factor 2. By cell hybridization studies, analysis of cross-resistance, and determination of specific binding sites for 125I-labeled diphtheria toxin, we showed that these cell strains fall into two distinct complementation groups. The Dipr group encompasses Chinese hamster strains that are resistant only to diphtheria toxin, as well as mouse LM cells. These strains possess a normal complement of high-affinity binding sites for diphtheria toxin, but these receptors are unable to deliver active toxin fragment A to the cytosol. Cells of the DPVr group have a broader spectrum of resistance, including Pseudomonas exotoxin A and several enveloped viruses as well as diphtheria toxin. In these studies, which investigate the resistance of these cells to diphtheria toxin, we demonstrate that they possess a reduced number of specific binding sites for this toxin and behave, phenotypically, like cells treated with the proton ionophore monensin. Their resistance is related to a defect in a mechanism required for release of active toxin from the endocytic vesicle.

show abstract

“…There are reports from resistant mouse and rat cells, that these cells bind the toxin. [28][29][30] Irrespective of the molecular mechanism involved DTx-induced transient podocyte malfunction and proteinuria might be a supplemental new and convenient experimental model, which can also be applied to the great variety of genetically engineered C57BL/6 mice. Of note, BALB/c mice get sick as well but the magnitude of proteinuria is lower than in C57BL/6 mice (Supplementary Figure 1D).…”

Section: Discussionmentioning

confidence: 99%

Impairment of podocyte function by diphtheria toxin—a new reversible proteinuria model in mice

Goldwich

Steinkasserer

Gessner

et al. 2012

Laboratory Investigation

View full text Add to dashboard Cite

Diphtheria toxin (DTx) receptor (DTR)-mediated conditional cell ablation in transgenic mice is a powerful tool to analyze cell function in vivo. Transgenic mice with cell-specific expression of the human DTR have been developed that allow conditional depletion of these cells in vivo through administration of the toxin. We have performed a careful analysis of mice after DTx injection and found an unexpected side effect. Treatment of wild-type C57BL/6 mice with DTx leads to a marked transient and completely reversible proteinuria, as a consequence of podocyte dysfunction that is morphologically characterized by foot process fusion and detachment from the glomerular basal membrane. In vitro analysis displayed that DTx-treated podocytes show diminished attachment to basal membrane proteins. Five to 9 days after DTx application the mice recover completely. Glomerular proteinuria is a hallmark of glomerular disease due to dysfunction of the filtration barrier. Rodents have been extensively used experimentally to better define the mechanisms of disease induction and progression. However, nongenetic mouse models of proteinuric glomerular damage are limited and display various shortcomings. We suggest DTx-induced transient kidney dysfunction as a new reversible model of experimental podocyte injury, which could be used as an additional approach to complement studies in human.

show abstract

Receptor-mediated endocytosis of diphtheria toxin by cells in culture.

Cited by 86 publications

References 31 publications

Identification of diphtheria toxin receptor and a nonproteinous diphtheria toxin-binding molecule in Vero cell membrane.

Identification of diphtheria toxin receptor and a nonproteinous diphtheria toxin-binding molecule in Vero cell membrane.

Binding and uptake of diphtheria toxin by toxin-resistant Chinese hamster ovary and mouse cells.

Impairment of podocyte function by diphtheria toxin—a new reversible proteinuria model in mice

Contact Info

Product

Resources

About