How Ricin and Shiga Toxin Reach the Cytosol of Target Cells: Retrotranslocation from the Endoplasmic Reticulum

Spooner, Robert A.; Lord, J. Michael

doi:10.1007/82_2011_154

Cited by 116 publications

(135 citation statements)

References 112 publications

(119 reference statements)

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“…Furthermore, pH-dependent conformation is a characteristic shared by several classes of bacterial toxins (47,48) as well as by viral proteins (49,50). Therefore, it is tempting to speculate that this scenario of increased immune response related to a pH-dependent heparin-binding capacity might apply to a variety of proteins from micro-organisms.…”

Section: Discussionmentioning

confidence: 99%

Acidosis Increases MHC Class II–Restricted Presentation of a Protein Endowed with a pH-Dependent Heparan Sulfate–Binding Ability

Knittel

Savatier

Upert

et al. 2015

The Journal of Immunology

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Heparan sulfate proteoglycans (HSPGs) are ubiquitously expressed molecules that participate in numerous biological processes. We previously showed that HSPGs expressed on the surface of APCs can serve as receptors for a hybrid protein containing an HS ligand and an Ag, which leads to more efficient stimulation of Th cells. To investigate whether such behavior is shared by proteins with inherent HS-binding ability, we looked for proteins endowed with this characteristic. We found that diphtheria toxin and its nontoxic mutant, called CRM197, can interact with HS. However, we observed that their binding ability is higher at pH 6 than at pH 7.4. Therefore, as extracellular acidosis occurs during infection by various micro-organisms, we assessed whether HS-binding capacity affects MHC class II–restricted presentation at different pHs. We first observed that pH decrease allows CRM197 binding to HSPG-expressing cells, including APCs. Then, we showed that this interaction enhances Ag uptake and presentation to Th cells. Lastly, we observed that pH decrease does not affect processing and presentation abilities of the APCs. Our findings show that acidic pH causes an HSPG-mediated uptake and an enhancement of T cell stimulation of Ags with the inherent ability to bind HSPGs pH-dependently. Furthermore, they suggest that proteins from micro-organisms with this binding characteristic might be supported more efficiently by the adaptive immune system when acidosis is triggered during infection.

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Section: Discussionmentioning

confidence: 99%

Acidosis Increases MHC Class II–Restricted Presentation of a Protein Endowed with a pH-Dependent Heparan Sulfate–Binding Ability

Knittel

Savatier

Upert

et al. 2015

The Journal of Immunology

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“…These are type II ribosome-inactivating proteins (RIPs) produced by Shigella dysenteriae and enterohemorrhagic strains of Escherichia coli, repectively (STx-1 is 99% identical in amino acid sequence to Shiga toxin). After binding to its cellular receptor, the Gb3 glycosphingolipid, the non-toxic B-subunit of Shiga toxin (STxB) transports the non-covalently linked catalytic A-subunit (STxA) along the retrograde route, via early endosomes, the trans-Golgi network (TGN) and Golgi cisternae, to the ER, from where STxA dislocates to the cytosol by exploiting the host ERassociated degradation (ERAD) machinery (Mallard et al, 1998;Spooner and Lord, 2012). STxA has N-glycosidase activity and inhibits protein biosynthesis by removing a specific adenine base from 28S rRNA of the 60S large ribosomal subunit.…”

Section: Introductionmentioning

confidence: 99%

Retrograde transport is not required for cytosolic translocation of the B-subunit of Shiga toxin

García-Castillo

Tran

Bobard

et al. 2015

Journal of Cell Science

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Antigen-presenting cells have the remarkable capacity to transfer exogenous antigens to the cytosol for processing by proteasomes and subsequent presentation on major histocompatibility complex class-I (MHC-I) molecules, a process termed cross-presentation. This is the target of biomedical approaches that aim to trigger a therapeutic immune response. The receptor-binding B-subunit of Shiga toxin (STxB) has been developed as an antigen delivery tool for such immunotherapy applications. In this study, we have analyzed pathways and trafficking factors that are involved in this process. A covalent conjugate between STxB and saporin was generated to quantitatively sample the membrane translocation step to the cytosol in differentiated monocyte-derived THP-1 cells. We have found that retrograde trafficking to the Golgi complex was not required for STxBsaporin translocation to the cytosol or for STxB-dependent antigen cross-presentation. Depletion of endosomal Rab7 inhibited, and lowering membrane cholesterol levels favored STxB-saporin translocation. Interestingly, experiments with reducible and nonreducible linker-arm-STxB conjugates led to the conclusion that after translocation, STxB remains associated with the cytosolic membrane leaflet. In summary, we report new facets of the endosomal escape process bearing relevance to antigen cross-presentation.

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“…Following endocytosis, RTB mediates the retrograde transport of ricin to the trans-Golgi network (TGN) and endoplasmic reticulum (ER). Once within the ER, the disulfide bond that links RTA to RTB is reduced by protein disulfide isomerase and RTA is retrotranslocated (dislocated) into the cell cytoplasm, where it triggers programmed cell death (7,8). Ricin is classified as a category B biothreat agent by the Centers for Disease Control and Prevention (CDC) because of its potential, when delivered via aerosol, to induce severe lung inflammation and tissue damage (9,10).…”

mentioning

confidence: 99%

High-Resolution Epitope Positioning of a Large Collection of Neutralizing and Nonneutralizing Single-Domain Antibodies on the Enzymatic and Binding Subunits of Ricin Toxin

Vance

Tremblay

Rong

et al. 2017

Clin Vaccine Immunol

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We previously produced a heavy-chain-only antibody (Ab) VH domain (V H H)-displayed phage library from two alpacas that had been immunized with ricin toxoid and nontoxic mixtures of the enzymatic ricin toxin A subunit (RTA) and binding ricin toxin B subunit (RTB) (D. J. Vance, J. M. Tremblay, N. J. Mantis, and C. B. Shoemaker, J Biol Chem 288:36538 -36547, 2013, https://doi.org/ 10.1074/jbc.M113.519207). Initial and subsequent screens of that library by direct enzyme-linked immunosorbent assay (ELISA) yielded more than two dozen unique RTAand RTB-specific V H Hs, including 10 whose structures were subsequently solved in complex with RTA. To generate a more complete antigenic map of ricin toxin and to define the epitopes associated with toxin-neutralizing activity, we subjected the V H Hdisplayed phage library to additional "pannings" on both receptor-bound ricin and antibody-captured ricin. We now report the full-length DNA sequences, binding affinities, and neutralizing activities of 68 unique V H Hs: 31 against RTA, 33 against RTB, and 4 against ricin holotoxin. Epitope positioning was achieved through cross-competition ELISAs performed with a panel of monoclonal antibodies (MAbs) and verified, in some instances, with hydrogen-deuterium exchange mass spectrometry. The 68 V H Hs grouped into more than 20 different competition bins. The RTA-specific V H Hs with strong toxin-neutralizing activities were confined to bins that overlapped two previously identified neutralizing hot spots, termed clusters I and II. The four RTB-specific V H Hs with potent toxin-neutralizing activity grouped within three adjacent bins situated at the RTA-RTB interface near cluster II. These results provide important insights into epitope interrelationships on the surface of ricin and delineate regions of vulnerability that can be exploited for the purpose of vaccine and therapeutic development.KEYWORDS antibody, biodefense, epitope, neutralizing, toxins, vaccines R icin is the prototype of the type II ribosome-inactivating protein (RIP) family of toxins (1). Ricin toxin A subunit (RTA) is an RNA N-glycosidase that catalyzes the hydrolysis of a conserved adenine residue within the sarcin/ricin loop (SRL) of 28S rRNA, resulting in ribosome arrest and apoptosis (2-4). Ricin toxin B subunit (RTB) is a galactose-and N-acetylgalactosamine (Gal/GalNAc)-specific lectin that promotes toxin entry into mammalian cells, including the epithelial cells that line the respiratory tract

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How Ricin and Shiga Toxin Reach the Cytosol of Target Cells: Retrotranslocation from the Endoplasmic Reticulum

Cited by 116 publications

References 112 publications

Acidosis Increases MHC Class II–Restricted Presentation of a Protein Endowed with a pH-Dependent Heparan Sulfate–Binding Ability

Acidosis Increases MHC Class II–Restricted Presentation of a Protein Endowed with a pH-Dependent Heparan Sulfate–Binding Ability

Retrograde transport is not required for cytosolic translocation of the B-subunit of Shiga toxin

High-Resolution Epitope Positioning of a Large Collection of Neutralizing and Nonneutralizing Single-Domain Antibodies on the Enzymatic and Binding Subunits of Ricin Toxin

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