Pathways followed by protein toxins into cells

Sandvig, Kirsten; Spilsberg, Bjørn; Lauvrak, Silje Ugland; Torgersen, Maria Lyngaas; Iversen, Tore Geir; Deurs, Bo van

doi:10.1078/1438-4221-00294

Cited by 125 publications

(99 citation statements)

References 54 publications

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“…Further, in the case of anthrax lethal factor, the motif is positioned between residues 31-40 in the mature protein, a region that is structurally distinct from the protective antigen binding domain (32), and N-terminal deletion analysis of anthrax lethal factor has shown previously that this region is required for toxicity (32,33). In contrast, this motif was not detected in those protein toxins known to undergo retrograde transport and whose catalytic domains enter the cytosol from the endoplasmic reticulum (e.g., cholera toxin, Shiga toxin, ricin toxin, and Pseudomonas exotoxin A) (17,34).…”

Section: P]-labeled Adpr-ef-2 (B)mentioning

confidence: 95%

A conserved motif in transmembrane helix 1 of diphtheria toxin mediates catalytic domain delivery to the cytosol

Ratts

Trujillo

Bharti

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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A 10-aa motif in transmembrane helix 1 of diphtheria toxin that is conserved in anthrax edema factor, anthrax lethal factor, and botulinum neurotoxin serotypes A, C, and D was identified by BLAST, CLUSTAL W, and MEME computational analysis. Using the diphtheria toxin-related fusion protein toxin DAB 389IL-2, we demonstrate that introduction of the L221E mutation into a highly conserved residue within this motif results in a nontoxic catalytic domain translocation deficient phenotype. To further probe the function of this motif in the process by which the catalytic domain is delivered from the lumen of early endosomes to the cytosol, we constructed a gene encoding a portion of diphtheria toxin transmembrane helix 1, T1, which carries the motif and is expressed from a CMV promoter. We then isolated stable transfectants of Hut102͞6TG cells that express the T1 peptide, Hut102͞6TG-T1. In contrast to the parental cell line, Hut102͞6TG-T1 cells are ca. 10 4 -fold more resistant to the fusion protein toxin. This resistance is completely reversed by coexpression of small interfering RNA directed against the gene encoding the T1 peptide in Hut102͞ 6TG-T1 cells. We further demonstrate by GST-DT140-271 pull-down experiments in the presence and absence of synthetic T1 peptides the specific binding of coatomer protein complex subunit ␤ to this region of the diphtheria toxin transmembrane domain.early endosomes ͉ translocation ͉ ADP-ribosyltransferase ͉ coatomer protein complex subunit ␤ T he intoxication of eukaryotic cells by diphtheria toxin follows an ordered series of interactions between the toxin and cellular factors that lead to inhibition of protein synthesis and cell death (1). Biochemical, genetic, and x-ray crystallographic analysis of the toxin has shown the protein to be composed of three distinct domains: an N-terminal catalytic (C) domain, a central transmembrane (T) domain, and the C-terminal receptor-binding domain (2-5). The intoxication process is initiated by the binding of the toxin to its cell surface receptor, a heparin binding epidermal growth factor-like precursor, and CD9 (6, 7). Once bound to its receptor, the toxin is internalized by receptormediated endocytosis into an early endosomal compartment (8). Upon acidification of the endosomal lumen by vesicular ATPase, the T domain undergoes a conformational change and spontaneously inserts into the vesicular membrane, forming an 18-to 22-Å pore or channel (9, 10). It is widely believed that the C domain, in a fully denatured form, is specifically thread through this channel and released into the cytosol. Once the C domain is refolded into an active conformation, it catalyzes the NAD ϩ -dependent ADP ribosylation of elongation factor 2 (EF-2), causing irreversible inhibition of protein synthesis and death of the cell by apoptosis (11,12). Although the endosomal membrane translocation of the C domain is understood in broad terms, the precise molecular mechanism(s) of this step in the intoxication process has remained largely unknown.Two hypotheses for C ...

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Section: P]-labeled Adpr-ef-2 (B)mentioning

confidence: 95%

A conserved motif in transmembrane helix 1 of diphtheria toxin mediates catalytic domain delivery to the cytosol

Ratts

Trujillo

Bharti

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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“…Thus, clathrin-independent uptake of the IL2 receptor is dependent on lipid rafts, 25 whereas ricin and cholera toxin can be internalized under conditions where clathrin-dependent uptake is inhibited and cholesterol has been depleted from the rest of the membrane. 26,28 In the case of anthrax toxin, the uptake is dependent on lipid rafts for the initiation of processing (protective antigen (PA) heptamerization, cleavage of PA and subsequent binding of lethal factor (LF)), and then the toxin (the complex of PA and LF) is taken in by a clathrin-dependent process. 29 One danger in using protein toxins to study a given process is that the toxin might induce signalling that could affect the process.…”

Section: Introductionmentioning

confidence: 99%

Delivery into cells: lessons learned from plant and bacterial toxins

2005

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“…Many protein toxins from poisonous plants or from pathogenic bacteria are able to penetrate into the cytosol of their target cells where they exert their toxic effects. Some of these toxins exploit the endogenous cellular machinery of endocytosis and intracellular sorting to gain access to the cell cytosol, but others carry their own translocation apparatus (1)(2)(3)(4). These latter toxins provide a unique opportunity to analyze the molecular mechanisms and the physicochemical principles underlying polypeptide transport across biological membranes.…”

mentioning

confidence: 99%

Bordetella pertussis adenylate cyclase toxin translocation across a tethered lipid bilayer

Veneziano

Rossi

Chenal

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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Significance Many bacterial toxins can cross biological membranes to reach the cytosol of mammalian cells, although how they pass through a lipid bilayer remains largely unknown. Bordetella pertussis adenylate cyclase (CyaA) toxin delivers its catalytic domain directly across the cell membrane. To characterize this unique translocation process, we designed an in vitro assay based on a tethered lipid bilayer assembled over a biosensor surface derivatized with calmodulin, a natural activator of the toxin. CyaA activation by calmodulin provided a highly sensitive readout for toxin translocation across the bilayer. CyaA translocation was calcium- and membrane potential-dependent but independent of any additional eukaryotic protein. This biomimetic membrane will permit in vitro studies of protein translocation in precisely defined conditions.

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Pathways followed by protein toxins into cells

Cited by 125 publications

References 54 publications

A conserved motif in transmembrane helix 1 of diphtheria toxin mediates catalytic domain delivery to the cytosol

A conserved motif in transmembrane helix 1 of diphtheria toxin mediates catalytic domain delivery to the cytosol

Delivery into cells: lessons learned from plant and bacterial toxins

Bordetella pertussis adenylate cyclase toxin translocation across a tethered lipid bilayer

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