Expression of Mutant Dynamin Inhibits Toxicity and Transport of Endocytosed Ricin to the Golgi Apparatus

Llorente, Alicia; Rapak, Andrzej; Schmid, Sandra L.; Deurs, Bo van; Sandvig, Kirsten

doi:10.1083/jcb.140.3.553

Cited by 118 publications

(110 citation statements)

References 81 publications

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“…5). The latter finding is in accordance with recent reports on lysosomal degradation of proteins being unaffected in dynamin K44A -overexpressing cells (49,50). Recently, association of dynamin-2 with late endosomes has been reported; this results in defective trafficking from late endosomes to the trans-Golgi network (50).…”

Section: Discussionsupporting

confidence: 81%

Inhibition of Clathrin-dependent Endocytosis Has Multiple Effects on Human Rhinovirus Serotype 2 Cell Entry

Bayer¹,

Schober²,

Hüttinger³

et al. 2001

Journal of Biological Chemistry

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Physiol. 131, 14 -22.) To resolve this apparent contradiction, we investigated the binding, internalization, conformational changes, and productive uncoating of HRV2 in HeLa cells subjected to hypotonic shock and potassium depletion. This treatment led to an increase in HRV2 binding, with internalization being barely affected. The generation of C-antigenic particles requiring pH <5.6 was strongly reduced due to an elevation of the pH in endosomal compartments. However, K ؉ depletion only slightly affected de novo viral protein synthesis, suggesting that productivity of viral RNA in the cytoplasm is enhanced and thus compensates for the reduction in C-antigenic particles. The distinct steps in the entry pathway of HRV2 are thus differently influenced by potassium depletion. Viral internalization under conditions of inhibited clathrin-dependent endocytosis without the need to disturb the ionic milieu was confirmed in HeLa cells overexpressing the nonfunctional dynamin-1 mutant K44A. Unexpectedly, overexpression of dynamin-1 K44A resulted in elevated endosomal pH compared with overexpression of wild-type dynamin.

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Section: Discussionsupporting

confidence: 81%

Inhibition of Clathrin-dependent Endocytosis Has Multiple Effects on Human Rhinovirus Serotype 2 Cell Entry

Bayer¹,

Schober²,

Hüttinger³

et al. 2001

Journal of Biological Chemistry

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“…A variety of evidence has accumulated to suggest the importance of traffic to the TGN and ER for cell killing by ricin, including the ability of brefeldin A and ilimaquinone to block ricin intoxication of cells in which these compounds cause a disruption of Golgi morphology [6,7]. In addition, expression of mutant dynamin, which blocks endocytosis and traffic of ricin to the TGN [8], inhibits ricin intoxication. Moreover, addition of the ER retrieval motif KDEL to the C-terminus of the A chain results in an engineered polypeptide with an increased cytotoxicity of 10-100-fold, dependent on cell type [9].…”

Section: Introductionmentioning

confidence: 99%

Cell polarization is required for ricin sensitivity in a Caco-2 cell line selected for ricin resistance

Jackman

Ellis

Gray

et al. 1999

Biochemical Journal

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It has been proposed that killing of mammalian cells by ricin requires efficient endocytic delivery to the trans-Golgi network (TGN) prior to retrograde transport to the endoplasmic reticulum and entry to the cytosol. In polarized epithelial cells, an efficient membrane-traffic pathway to the TGN is present from the basolateral but not the apical plasma-membrane domain. Thus one can hypothesize that a ricin-resistant phenotype might be demonstrated by polarized cells that fail to differentiate and thus fail to develop an efficient membrane-traffic pathway from the basolateral plasma membrane to the TGN. We have isolated and studied a ricin-resistant Caco-2 cell clone (Caco-2-RCAr clone 2) which, when grown on plastic, was deficient in differentiation, measured by the development of polarized-cell-surface marker enzymes. The deficiency in differentiation was partially reversed, and ricin sensitivity was restored, when the cells were grown on filter supports. Our data provide the first evidence of a ricin-resistant cell line where resistance is due to the lack of development of polarized cell surfaces. The observed ricin resistance is consistent with the requirement that ricin is delivered to the TGN before its A chain enters the cytosol to mediate cell killing.

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“…Dynamin is most likely part of the machinery involved in vesicles formation that is located on endosomes and required for ricin to be transported to the TGN (Llorente et al, 1998). However, it is not yet clear whether RT is transferred directly from early endosomes to the TGN or if it transits by late endosomes (Simpson et al, 1995).…”

Section: Discussionmentioning

confidence: 99%

“…Transfer of RT from early endosomes to the TGN and subsequent toxicity are blocked by expression of dyn-K44A (Llorente et al, 1998). Dynamin is most likely part of the machinery involved in vesicles formation that is located on endosomes and required for ricin to be transported to the TGN (Llorente et al, 1998).…”

Section: Discussionmentioning

confidence: 99%

“…An alternative explanation is that a minor amount of ricin molecules is taken up by the dynamindependent mechanism from early endosomes and transferred to late endosomes, whereas CNF1 will not follow this pathway but rather the degradative pathway to proceed to late endosomes. It has been shown that ligands going to late endosomes for degradation into lysosomes are not sensitive to dyn-K44A effects (Damke et al, 1994;Llorente et al, 1998).…”

Section: Discussionmentioning

confidence: 99%

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The p21 Rho-activating Toxin Cytotoxic Necrotizing Factor 1 Is Endocytosed by a Clathrin-independent Mechanism and Enters the Cytosol by an Acidic-dependent Membrane Translocation Step

Contamin

Galmiche

Doye

et al. 2000

MBoC

100

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Cytotoxic necrotizing factor 1 (CNF1), a protein produced by pathogenic strains of Escherichia coli, activates the p21 Rho-GTP-binding protein, inducing a profound reorganization of the actin cytoskeleton. CNF1 binds to its cell surface receptor on HEp-2 cells with high affinity (K d ϭ 20 pM). In HEp-2 cells the action of CNF1 is not blocked in the presence of filipin, a drug described to reduce cholera toxin internalization by the caveolae-like mechanism. Moreover, HEp-2 cells, which express a dominant negative form of proteins that impair the formation of clathrin coated-vesicles and internalization of transferrin (Eps15, dynamin or intersectin-Src homology 3), are still sensitive to CNF1. In this respect, the endocytosis of CNF1 is similar to the plant toxin ricin. However, unlike ricin toxin, CNF1 does not cross the Golgi apparatus and requires an acidic cell compartment to transfer its enzymatic activity into the cytosol in a manner similar to that required by diphtheria toxin. As shown for diphtheria toxin, the pH-dependent membrane translocation step of CNF1 could be mimicked at the level of the plasma membrane by a brief exposure to a pH of Յ5.2. CNF1 is the first bacterial toxin described that uses both a clathrinindependent endocytic mechanism and an acidic-dependent membrane translocation step in its delivery of the catalytic domain to the cell cytosol. INTRODUCTIONBacterial protein toxins are among the most powerful virulence factors produced by pathogenic microorganisms. In general, these toxins are divided into three groups according to their mechanism of action (Boquet and Gill, 1991). Group I toxins such as the heat-stable Escherichia coli enterotoxin (reviewed by Sears and Kaper, 1996) act on cell surface where they induce transmembrane signaling. Group II toxins or "pore-forming toxins" (e.g., perfringolysin O, Staphylococcus aureus alpha toxin, and aerolysin [reviewed by Bhakdi et al., 1996]) act by disrupting the integrity of the plasma membrane. Finally, group III toxins, such as diphtheria, cholera, or tetanus toxins, transfer an enzymatically active domain into the cytosol and modify a eukaryotic target, which gives rise to toxicity (reviewed by Montecucco et al., 1994). The latter group is clearly the most potent, because it is believed that one or a few copies of the toxin enzymatic domain, for instance of the diphtheria toxin (DT) fragment A, introduced into the cytosol can kill a cell within 2 d (Yamaizumi et al., 1978). Group III toxins are also known as A-B toxins on the basis of their structure-function relationships, which involve a catalytic domain (the A subunit) and cell binding and membrane translocation domains (the B subunit).Translocation of the catalytic domain of the type III toxins into the eukaryotic cytosol appears to be accomplished by either one or two mechanisms (reviewed by Johannes and Goud, 1998;Lord and Roberts, 1998;Montecucco, 1998). In the first instance, for which the paradigm is DT (Sandvig and Olsnes, 1991), the toxin first binds to its cell surface and is ...

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Expression of Mutant Dynamin Inhibits Toxicity and Transport of Endocytosed Ricin to the Golgi Apparatus

Cited by 118 publications

References 81 publications

Inhibition of Clathrin-dependent Endocytosis Has Multiple Effects on Human Rhinovirus Serotype 2 Cell Entry

Inhibition of Clathrin-dependent Endocytosis Has Multiple Effects on Human Rhinovirus Serotype 2 Cell Entry

Cell polarization is required for ricin sensitivity in a Caco-2 cell line selected for ricin resistance

The p21 Rho-activating Toxin Cytotoxic Necrotizing Factor 1 Is Endocytosed by a Clathrin-independent Mechanism and Enters the Cytosol by an Acidic-dependent Membrane Translocation Step

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