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Shiga toxin facilitates its retrograde transport by modifying microtubule dynamics.

ABSTRACT: 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.

SUBMITTER: Hehnly H

PROVIDER: S-EPMC1635369 | biostudies-literature | 2006 Oct

REPOSITORIES: biostudies-literature

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Shiga toxin facilitates its retrograde transport by modifying microtubule dynamics.

Hehnly Heidi H Sheff David D Stamnes Mark M

Molecular biology of the cell 20060802 10

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 ...[more]

PMID: 16885418

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Project description:Ricin is a member of the ubiquitous family of plant and bacterial AB toxins that gain entry into the cytosol of host cells through receptor-mediated endocytosis and retrograde traffic through the trans-Golgi network (TGN) and endoplasmic reticulum (ER). While a few ricin toxin-specific neutralizing monoclonal antibodies (MAbs) have been identified, the mechanisms by which these antibodies prevent toxin-induced cell death are largely unknown. Using immunofluorescence confocal microscopy and a TGN-specific sulfation assay, we demonstrate that 24B11, a MAb against ricin's binding subunit (RTB), associates with ricin in solution or when prebound to cell surfaces and then markedly enhances toxin uptake into host cells. Following endocytosis, however, toxin-antibody complexes failed to reach the TGN; instead, they were shunted to Rab7-positive late endosomes and LAMP-1-positive lysosomes. Monovalent 24B11 Fab fragments also interfered with toxin retrograde transport, indicating that neither cross-linking of membrane glycoproteins/glycolipids nor the recently identified intracellular Fc receptor is required to derail ricin en route to the TGN. Identification of the mechanism(s) by which antibodies like 24B11 neutralize ricin will advance our fundamental understanding of protein trafficking in mammalian cells and may lead to the discovery of new classes of toxin inhibitors and therapeutics for biodefense and emerging infectious diseases. IMPORTANCE Ricin is the prototypic member of the AB family of medically important plant and bacterial toxins that includes cholera and Shiga toxins. Ricin is also a category B biothreat agent. Despite ongoing efforts to develop vaccines and antibody-based therapeutics against ricin, very little is known about the mechanisms by which antibodies neutralize this toxin. In general, it is thought that antibodies simply prevent toxins from attaching to cell surface receptors or promote their clearance through Fc receptor (FcR)-mediated uptake. In this report, however, we describe a neutralizing monoclonal antibody (MAb) against ricin's binding subunit (RTB) that not only associates with ricin after the toxin has bound to the cell's surface but actually enhances toxin uptake into host cells. Following endocytosis, the antibody-toxin complexes are then routed for degradation. The results of this study are important because they reveal a previously unappreciated role for B-subunit-specific antibodies in intracellular neutralization of ricin toxin.

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Shiga toxin facilitates its retrograde transport by modifying microtubule dynamics.

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Shiga toxin facilitates its retrograde transport by modifying microtubule dynamics.

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