Endocytosis, intracellular transport, and cytotoxic action of Shiga toxin and ricin

1996 ◽  
Vol 76 (4) ◽  
pp. 949-966 ◽  
Author(s):  
K. Sandvig ◽  
B. van Deurs
Keyword(s):  
Shiga Toxin ◽  
Intracellular Transport ◽  
Retrograde Transport ◽  
Cytotoxic Action ◽  
Coated Pits ◽  
Polarized Epithelia ◽  
Protein Toxins ◽  
A Cell ◽  
Membrane Bound ◽  
Golgi Network

Protein toxins such as ricin and Shiga toxin with intracellular targets have to be endocytosed and translocated to the cytosol to inhibit the protein synthesis and thereby kill the cell. Ricin is internalized by both clathrin-dependent and -independent endocytic mechanisms, whereas Shiga toxin seems to be taken up exclusively from clathrin-coated pits. After endocytosis, internalized membrane and content are delivered to endosomes, where sorting for further routing in the cell takes place. Toxins that remain membrane bound at low endosomal pH can be recycled to the cell surface or transcytosed in polarized epithelia. A large proportion of internalized toxin is transported to lysosomes for degradation. Most importantly, a fraction of the internalized ricin and Shiga toxin molecules is delivered to the trans-Golgi network (TGN). Shiga toxin can, in some very sensitive cells, be transported retrogradely through the Golgi cisterns all the way back to the endoplasmic reticulum (ER), and it is possible that also ricin is transported retrogradely to the ER. In this review, a cell biological overview of these intracellular transport steps is presented, and evidence is provided that the delivery to the TGN and the subsequent retrograde transport to the ER are required for optimal intoxication. Moreover, it is argued that knowledge of this transport is important for targeted drug delivery such as the application of immunotoxins in cancer therapy.

scholarly journals Syntaxin 5-Dependent Retrograde Transport to thetrans-Golgi Network Is Required for Adeno-Associated Virus Transduction

2014 ◽  
Vol 89 (3) ◽  
pp. 1673-1687 ◽  
Author(s):  
Mathieu E. Nonnenmacher ◽  
Jean-Christophe Cintrat ◽  
Daniel Gillet ◽  
Thomas Weber
Keyword(s):  
Gene Delivery ◽  
Golgi Apparatus ◽  
Intracellular Transport ◽  
Brefeldin A ◽  
Retrograde Transport ◽  
Transport Pathway ◽  
Adeno Associated Virus ◽  
Viral Attachment ◽  
Golgi Network ◽  
Endosomal System

ABSTRACTIntracellular transport of recombinant adeno-associated virus (AAV) is still incompletely understood. In particular, the trafficking steps preceding the release of incoming AAV particles from the endosomal system into the cytoplasm, allowing subsequent nuclear import and the initiation of gene expression, remain to be elucidated fully. Others and we previously showed that a significant proportion of viral particles are transported to the Golgi apparatus and that Golgi apparatus disruption caused by the drug brefeldin A efficiently blocks AAV serotype 2 (AAV2) transduction. However, because brefeldin A is known to exert pleiotropic effects on the entire endosomal system, the functional relevance of transport to the Golgi apparatus for AAV transduction remains to be established definitively. Here, we show that AAV2 trafficking toward thetrans-Golgi network (TGN) and the Golgi apparatus correlates with transduction efficiency and relies on a nonclassical retrograde transport pathway that is independent of the retromer complex, late endosomes, and recycling endosomes. AAV2 transduction is unaffected by the knockdown of syntaxins 6 and 16, which are two major effectors in the retrograde transport of both exogenous and endogenous cargo. On the other hand, inhibition of syntaxin 5 function by small interfering RNA silencing or treatment with cyclized Retro-2 strongly decreases AAV2 transduction and transport to the Golgi apparatus. This inhibition of transduction is observed with several AAV serotypes and a number of primary and immortalized cells. Together, our data strongly suggest that syntaxin 5-mediated retrograde transport to the Golgi apparatus is a broadly conserved feature of AAV trafficking that appears to be independent of the identity of the receptors used for viral attachment.IMPORTANCEGene therapy constitutes a promising approach for the treatment of life-threatening conditions refractory to any other form of remedy. Adeno-associated virus (AAV) vectors are currently being evaluated for the treatment of diseases such as Duchenne muscular dystrophy, hemophilia, heart failure, Parkinson's disease, and others. Despite their promise as gene delivery vehicles, a better understanding of the biology of AAV-based vectors is necessary to improve further their efficacy. AAV vectors must reach the nucleus in order to deliver their genome, and their intracellular transport is not fully understood. Here, we dissect an important step of the intracellular journey of AAV by showing that retrograde transport of capsids to thetrans-Golgi network is necessary for gene delivery. We show that the AAV trafficking route differs from that of known Golgi apparatus-targeted cargos, and we raise the possibility that this nonclassical pathway is shared by most AAV variants, regardless of their attachment receptors.

scholarly journals Shiga Toxin Facilitates Its Retrograde Transport by Modifying Microtubule Dynamics

2006 ◽  
Vol 17 (10) ◽  
pp. 4379-4389 ◽  
Author(s):  
Heidi Hehnly ◽  
David Sheff ◽  
Mark Stamnes
Keyword(s):  
Shiga Toxin ◽  
Intracellular Transport ◽  
Secretory Pathway ◽  
Retrograde Transport ◽  
Host Cells ◽  
Mammalian Host ◽  
Golgi Stack ◽  
Toxin Binding ◽  
Cytoskeletal Dynamics ◽  
Induced Changes

The bacterial exotoxin Shiga toxin is endocytosed by mammalian host cells and transported retrogradely through the secretory pathway before entering the cytosol. Shiga toxin also increases the levels of microfilaments and microtubules (MTs) upon binding to the cell surface. The purpose for this alteration in cytoskeletal dynamics is unknown. We have investigated whether Shiga toxin-induced changes in MT levels facilitate its intracellular transport. We have tested the effects of the Shiga toxin B subunit (STB) on MT-dependent and -independent transport steps. STB increases the rate of MT-dependent Golgi stack repositioning after nocodazole treatment. It also enhances the MT-dependent accumulation of transferrin in a perinuclear recycling compartment. By contrast, the rate of MT-independent transferrin recycling is not significantly different when STB is present. We found that STB normally requires MTs and dynein for its retrograde transport to the juxtanuclear Golgi complex and that STB increases MT assembly. Furthermore, we find that MT polymerization is limiting for STB transport in cells. These results show that STB-induced changes in cytoskeletal dynamics influence intracellular transport. We conclude that the increased rate of MT assembly upon Shiga toxin binding facilitates the retrograde transport of the toxin through the secretory pathway.

scholarly journals The Protein Toxins Ricin and Shiga Toxin as Tools to Explore Cellular Mechanisms of Internalization and Intracellular Transport

Toxins ◽  
2021 ◽  
Vol 13 (6) ◽  
pp. 377
Author(s):  
Kirsten Sandvig ◽  
Simona Kavaliauskiene ◽  
Tore Skotland
Keyword(s):  
Shiga Toxin ◽  
Intracellular Transport ◽  
Lipid Classes ◽  
Mass Spectrometry Analysis ◽  
Biological Research ◽  
Cellular Mechanisms ◽  
Spectrometry Analysis ◽  
Lipid Species ◽  
Protein Toxins

Protein toxins secreted by bacteria and found in plants can be threats to human health. However, their extreme toxicity can also be exploited in different ways, e.g., to produce hybrid toxins directed against cancer cells and to study transport mechanisms in cells. Investigations during the last decades have shown how powerful these molecules are as tools in cell biological research. Here, we first present a partly historical overview, with emphasis on Shiga toxin and ricin, of how such toxins have been used to characterize processes and proteins of importance for their trafficking. In the second half of the article, we describe how one can now use toxins to investigate the role of lipid classes for intracellular transport. In recent years, it has become possible to quantify hundreds of lipid species using mass spectrometry analysis. Thus, it is also now possible to explore the importance of lipid species in intracellular transport. The detailed analyses of changes in lipids seen under conditions of inhibited toxin transport reveal previously unknown connections between syntheses of lipid classes and demonstrate the ability of cells to compensate under given conditions.

scholarly journals Retrograde transport from the Golgi complex to the ER of both Shiga toxin and the nontoxic Shiga B-fragment is regulated by butyric acid and cAMP.

1994 ◽  
Vol 126 (1) ◽  
pp. 53-64 ◽  
Author(s):  
K Sandvig ◽  
M Ryd ◽  
O Garred ◽  
E Schweda ◽  
P K Holm ◽  
...  
Keyword(s):  
Fatty Acid Composition ◽  
Fatty Acid ◽  
Golgi Apparatus ◽  
Butyric Acid ◽  
Golgi Complex ◽  
Shiga Toxin ◽  
Acid Treatment ◽  
Retrograde Transport ◽  
A431 Cells ◽  
Golgi Network

Endocytosed Shiga toxin is transported from the Golgi complex to the endoplasmic reticulum in butyric acid-treated A431 cells. We here examine the extent of this retrograde transport and its regulation. The short B fragment of Shiga toxin is sufficient for transport to the ER. The B fragment of cholera toxin, which also binds to glycolipids, is transported to all the Golgi cisterns, but cannot be localized in the ER even after butyric acid treatment. Under all conditions the toxic protein ricin was found predominantly in the trans-Golgi network. There is no transport of endocytosed fluid to the Golgi apparatus or to the ER even after butyric acid treatment and in the presence of Shiga toxin, indicating that transport to the ER, through the trans-Golgi network and the cisterns of the Golgi apparatus, involves several sorting stations. Since Shiga toxin receptors (Gb3) in butyric acid-treated A431 cells seem to have a ceramide moiety with longer fatty acids than in untreated cells, the possibility exists that fatty acid composition of the receptor is important for sorting to the ER. Both retrograde transport and intoxication with Shiga toxin can also be induced by cAMP, supporting the idea that retrograde transport from the Golgi to the ER is required for intoxication. The data suggest that transport to the ER in cells in situ may depend on fatty acid composition and is regulated by physiological signals.

I. Shiga toxin B-subunit system: retrograde transport, intracellular vectorization, and more

2002 ◽  
Vol 283 (1) ◽  
pp. G1-G7 ◽  
Author(s):  
Ludger Johannes
Keyword(s):  
Endoplasmic Reticulum ◽  
Shiga Toxin ◽  
Intracellular Transport ◽  
Molecular Mechanisms ◽  
Physiological Function ◽  
Retrograde Transport ◽  
Toxin B ◽  
B Subunit ◽  
New Methods ◽  
Therapeutic Compounds

Many intracellular transport routes are still little explored. This is particularly true for retrograde transport between the plasma membrane and the endoplasmic reticulum. Shiga toxin B subunit has become a powerful tool to study this pathway, and recent advances on the molecular mechanisms of transport in the retrograde route and on its physiological function(s) are summarized. Furthermore, it is discussed how the study of retrograde transport of Shiga toxin B subunit allows one to design new methods for the intracellular delivery of therapeutic compounds.

μ1A deficiency induces a profound increase in MPR300/IGF-II receptor internalization rate

2001 ◽  
Vol 114 (24) ◽  
pp. 4469-4476 ◽  
Author(s):  
Christoph Meyer ◽  
Eeva-Liisa Eskelinen ◽  
Medigeshi Ramarao Guruprasad ◽  
Kurt von Figura ◽  
Peter Schu
Keyword(s):  
Plasma Membrane ◽  
Fold Increase ◽  
Retrograde Transport ◽  
Receptor Internalization ◽  
Recycling Rate ◽  
Coated Pits ◽  
Trans Golgi Network ◽  
Internalization Rate ◽  
Clathrin Adaptor ◽  
Golgi Network

The mannose-6-phosphate/IGF-II receptor MPR300 mediates sorting of lysosomal enzymes from the trans-Golgi network to endosomes and endocytosis of hormones, for example, of IGF-II. We analyzed transport of MPR300 in μ1A-adaptin-deficient fibroblasts, which lack a functional AP-1 clathrin adaptor complex. In μ1A-adaptin-deficient fibroblasts, the homologous MPR46 accumulates in endosomes due to a block in retrograde transport to the trans-Golgi network. The MPR300-mediated endocytosis is markedly enhanced. We demonstrate that the seven-fold increase in endocytosis is not associated with an increased steady-state concentration of receptors at the plasma membrane, but with an increased internalization rate of MPR300. Internalization of other receptors that are also endocytosed by AP-2 is not affected. More MPR300 receptors are found in clathrin-coated pits of the plasma membrane, whereas outside coated-areas, more MPR300 are concentrated in clusters and all intracellular receptors reside in endosomes, which are in equilibrium with the plasma membrane. Thus AP-1-mediated transport of MPR300 from endosomes to the TGN controls indirectly the recycling rate of the receptor between the plasma membrane and endosomes.

scholarly journals Human Papillomavirus Entry: Hiding in a Bubble

2016 ◽  
Vol 90 (18) ◽  
pp. 8032-8035 ◽  
Author(s):  
Stephen DiGiuseppe ◽  
Malgorzata Bienkowska-Haba ◽  
Martin Sapp
Keyword(s):  
Human Papillomavirus ◽  
Nuclear Envelope ◽  
Viral Dna ◽  
Nuclear Envelope Breakdown ◽  
Transient Nature ◽  
Transport Vesicle ◽  
A Cell ◽  
Membrane Bound ◽  
Spindle Microtubules ◽  
Golgi Network

Incoming human papillomavirus (HPV) utilize vesicular transport to traffic from the plasma membrane to the trans-Golgi network. Following nuclear envelope breakdown during mitosis, the viral DNA associates with condensed chromosomes utilizing spindle microtubules for delivery. Most intriguingly, the viral DNA resides in a transport vesicle until mitosis is completed and the nuclear envelope has reformed. This finding provides support for the transient existence of nuclear membrane-bound vesicles. Due to their transient nature, it also points to the existence of a cell pathway for the disposal of vesicles ending up fortuitously or purposefully in the nucleus.

scholarly journals Endocytosis and intracellular transport of the glycolipid-binding ligand Shiga toxin in polarized MDCK cells.

1991 ◽  
Vol 113 (3) ◽  
pp. 553-562 ◽  
Author(s):  
K Sandvig ◽  
K Prydz ◽  
M Ryd ◽  
B van Deurs
Keyword(s):  
Golgi Apparatus ◽  
Shiga Toxin ◽  
Intracellular Transport ◽  
Mdck Cells ◽  
Short Chain Fatty Acids ◽  
Subcellular Fractionation ◽  
Shigella Dysenteriae ◽  
Tumor Promoter ◽  
Coated Pits ◽  
Cell Monolayers

The glycolipid-binding cytotoxin produced by Shigella dysenteriae 1, Shiga toxin, binds to MDCK cells (strain 1) only after treatment with short-chain fatty acids like butyric acid or with the tumor promoter 12-O-tetradecanoylphorbol 13-acetate. The induced binding sites were found to be functional with respect to endocytosis and translocation of toxin to the cytosol. Glycolipids that bind Shiga toxin appeared at both the apical and the basolateral surface of polarized MDCK cells grown on filters, and Shiga toxin was found to be endocytosed from both sides of the cells. This was demonstrated by EM of cells incubated with Shiga-HRP and by subcellular fractionation of cells incubated with 125I-labeled Shiga toxin. The data indicated that toxin molecules are endocytosed from coated pits, and that some internalized Shiga toxin is transported to the Golgi apparatus. Fractionation of polarized cells incubated with 125I-Shiga toxin showed that the transport of toxin to the Golgi apparatus was equally efficient from both poles of the cells. After 1-h incubation at 37 degrees C approximately 10% of the internalized toxin was found in the Golgi fractions. The results thus suggest that glycolipids can be efficiently transported to the Golgi apparatus from both sides of polarized MDCK cell monolayers.

Targeting of Shiga Toxin B-Subunit to Retrograde Transport Route in Association with Detergent-resistant Membranes

2001 ◽  
Vol 12 (8) ◽  
pp. 2453-2468 ◽  
Author(s):  
Thomas Falguières ◽  
Frédéric Mallard ◽  
Carole Baron ◽  
Daniel Hanau ◽  
Clifford Lingwood ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Hela Cells ◽  
Shiga Toxin ◽  
Secretory Pathway ◽  
Retrograde Transport ◽  
Toxin B ◽  
B Subunit ◽  
Early Endosomes ◽  
Golgi Network ◽  
Triton X

In HeLa cells, Shiga toxin B-subunit is transported from the plasma membrane to the endoplasmic reticulum, via early endosomes and the Golgi apparatus, circumventing the late endocytic pathway. We describe here that in cells derived from human monocytes, i.e., macrophages and dendritic cells, the B-subunit was internalized in a receptor-dependent manner, but retrograde transport to the biosynthetic/secretory pathway did not occur and part of the internalized protein was degraded in lysosomes. These differences correlated with the observation that the B-subunit associated with Triton X-100-resistant membranes in HeLa cells, but not in monocyte-derived cells, suggesting that retrograde targeting to the biosynthetic/secretory pathway required association with specialized microdomains of biological membranes. In agreement with this hypothesis we found that in HeLa cells, the B-subunit resisted extraction by Triton X-100 until its arrival in the target compartments of the retrograde pathway, i.e., the Golgi apparatus and the endoplasmic reticulum. Furthermore, destabilization of Triton X-100-resistant membranes by cholesterol extraction potently inhibited B-subunit transport from early endosomes to thetrans-Golgi network, whereas under the same conditions, recycling of transferrin was not affected. Our data thus provide first evidence for a role of lipid asymmetry in membrane sorting at the interface between early endosomes and the trans-Golgi network.

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