Key Points
-
The emergence of bioinformatics has changed how the study of microbial pathogenesis is carried out, which has facilitated the development of 'reverse proteomics' strategies. Researchers no longer need to clinically identify a pathogen before identifying putative virulence factors.
-
These analyses can be used to identify new bacterial ADP-ribosyltransferase toxins (bARTTs) on the basis of conserved function, even when the proteins have novel structural organizations. This strategy has been proven by the identification of putative bARTTs, followed by experimental confirmation of their ADP-ribosyltransferase activity. Several novel protein toxins have been identified and characterized in this manner.
-
New toxins, such as cholix toxin (ChxA), SpyA, HopU1, SpvB and the TcC proteins have been identified using this strategy of reverse proteomics and have been experimentally shown to have the predicted ADP-ribosyltransferase enzymatic activity.
-
However, this strategy still has limitations. Many of the novel toxins have unique substrates or structural or delivery properties that were not predicted by bioinformatic methods. Experimental confirmation of the importance of a putative virulence factor according to Falkow's molecular postulates also needs to be considered.
-
The continuing growth of the family of bacterial ADP-ribosylating toxins should enable greater ability to create 'rules' to properly identify substrates and the structural organization of novel toxins. Extrapolation of this technique to other toxins or families of proteins may greatly improve the field of microbiology.
Abstract
Bacterial ADP-ribosyltransferase toxins (bARTTs) transfer ADP-ribose to eukaryotic proteins to promote bacterial pathogenesis. In this Review, we use prototype bARTTs, such as diphtheria toxin and pertussis toxin, as references for the characterization of several new bARTTs from human, insect and plant pathogens, which were recently identified by bioinformatic analyses. Several of these toxins, including cholix toxin (ChxA) from Vibrio cholerae, SpyA from Streptococcus pyogenes, HopU1 from Pseudomonas syringae and the Tcc toxins from Photorhabdus luminescens, ADP-ribosylate novel substrates and have unique organizations, which distinguish them from the reference toxins. The characterization of these toxins increases our appreciation of the range of structural and functional properties that are possessed by bARTTs and their roles in bacterial pathogenesis.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 12 print issues and online access
$209.00 per year
only $17.42 per issue
Buy this article
- Purchase on Springer Link
- Instant access to full article PDF
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Aktories, K., Braun, U., Rösener, S., Just, I. & Hall, A. The rho gene product expressed in E. coli is a substrate of botulinum ADP-ribosyltransferase C3. Biochem. Biophys. Res. Commun. 158, 209–213 (1989).
Braun, U., Habermann, B., Just, I., Aktories, K. & Vandekerckhove, J. Purification of the 22 kDa protein substrate of botulinum ADP-ribosyltransferase C3 from porcine brain cytosol and its characterization as a GTP-binding protein highly homologous to the rho gene product. FEBS Lett. 243, 70–76 (1989).
Cassel, D. & Selinger, Z. Mechanism of adenylate cyclase activation by cholera toxin: inhibition of GTP hydrolysis at the regulatory site. Proc. Natl Acad. Sci. USA 74, 3307–3311 (1977).
Gill, D. M. & Meren, R. ADP-ribosylation of membrane proteins catalyzed by cholera toxin: basis of the activation of adenylate cyclase. Proc. Natl Acad. Sci. USA 75, 3050–3054 (1978).
West, R. E., Moss, J., Vaughan, M., Liu, T. & Liu, T. Y. Pertussis toxin-catalyzed ADP-ribosylation of transducin. Cysteine 347 is the ADP-ribose acceptor site. J. Biol. Chem. 260, 14428–14430 (1985).
Aktories, K. et al. Botulinum C2 toxin ADP-ribosylates actin. Nature 322, 390–392 (1986).
Holbourn, K. P., Shone, C. C. & Acharya, K. R. A family of killer toxins. FEBS J. 273, 4579–4593 (2006).
Hottiger, M. O., Hassa, P. O., Lüscher, B., Schüler, H. & Koch-Nolte, F. Toward a unified nomenclature for mammalian ADP-ribosyltransferases. Trends Biochem. Sci. 35, 208–219 (2010).
Tsuge, H. et al. Structural basis of actin recognition and arginine ADP-ribosylation by Clostridium perfringens ι-toxin. Proc. Natl Acad. Sci. USA 105, 7399–7404 (2008).
Tsurumura, T. et al. Arginine ADP-ribosylation mechanism based on structural snapshots of iota-toxin and actin complex. Proc. Natl Acad. Sci. USA 110, 4267–4272 (2013).
Jank, T. & Aktories, K. Strain-alleviation model of ADP-ribosylation. Proc. Natl Acad. Sci. USA 110, 4163–4164 (2013).
Barth, H. & Aktories, K. New insights into the mode of action of the actin ADP-ribosylating virulence factors Salmonella enterica SpvB and Clostridium botulinum C2 toxin. Eur. J. Cell Biol. 90, 944–950 (2011).
Barth, H., Aktories, K., Popoff, M. R. & Stiles, B. G. Binary bacterial toxins: biochemistry, biology, and applications of common Clostridium and Bacillus proteins. Microbiol. Mol. Biol. Rev. 68, 373–402 (2004).
Vogelsgesang, M., Pautsch, A. & Aktories, K. C3 exoenzymes, novel insights into structure and action of Rho-ADP-ribosylating toxins. Naunyn Schmiedebergs Arch. Pharmacol. 374, 347–360 (2007).
Dueholm, M. S., Albertsen, M., Otzen, D. & Nielsen, P. H. Curli functional amyloid systems are phylogenetically widespread and display large diversity in operon and protein structure. PLoS ONE 7, e51274 (2012).
Doxey, A. C. & McConkey, B. J. Prediction of molecular mimicry candidates in human pathogenic bacteria. Virulence 4, 453–466 (2013).
Priest, N. K. et al. From genotype to phenotype: can systems biology be used to predict Staphylococcus aureus virulence? Nature Rev. Microbiol. 10, 791–797 (2012).
Masignani, V. et al. In silico identification of novel bacterial ADP-ribosyltransferases. Int. J. Med. Microbiol. 293, 471–478 (2004).
Fieldhouse, R. J., Turgeon, Z., White, D. & Merrill, A. R. Cholera- and anthrax-like toxins are among several new ADP-ribosyltransferases. PLoS Comput. Biol. 6, e1001029 (2010). This paper shows that bioinformatics could be used to discover new toxins using protein-fold homology and amino acid sequence conservation searches; it also characterizes several toxins in silico , several of which were later experimentally confirmed to have ADP-ribosyltransferase activity.
Choe, S. et al. The crystal structure of diphtheria toxin. Nature 357, 216–222 (1992).
Gill, D. M. & Dinius, L. L. Observations on the structure of diphtheria toxin. J. Biol. Chem. 246, 1485–1491 (1971).
Collier, R. J. & Kandel, J. Structure and activity of diphtheria toxin. I. Thiol-dependent dissociation of a fraction of toxin into enzymically active and inactive fragments. J. Biol. Chem. 246, 1496–1503 (1971).
Gill, D. M. & Pappenheimer, A. M. Jr. Structure–activity relationships in diphtheria toxin. J. Biol. Chem. 246, 1492–1495 (1971).
Zhang, R.-G. et al. The three-dimensional crystal structure of cholera toxin. J. Mol. Biol. 251, 563–573 (1995).
Sixma, T. K. et al. Refined structure of Escherichia coli heat-labile enterotoxin, a close relative of cholera toxin. J. Mol. Biol. 230, 890–918 (1993).
Blöcker, D. et al. The C terminus of component C2II of Clostridium botulinum C2 toxin is essential for receptor binding. Infect. Immun. 68, 4566–4573 (2000).
Schleberger, C., Hochmann, H., Barth, H., Aktories, K. & Schulz, G. E. Structure and action of the binary C2 toxin from Clostridium botulinum. J. Mol. Biol. 364, 705–715 (2006).
Aktories, K., Weller, U. & Chhatwal, G. S. Clostridium botulinum type C produces a novel ADP-ribosyltransferase distinct from botulinum C2 toxin. FEBS Lett. 212, 109–113 (1987).
Han, S., Arvai, A. S., Clancy, S. B. & Tainer, J. A. Crystal structure and novel recognition motif of Rho ADP-ribosylating C3 exoenzyme from Clostridium botulinum: structural insights for recognition specificity and catalysis. J. Mol. Biol. 305, 95–107 (2001).
Song, J., Gao, X. & Galan, J. E. Structure and function of the Salmonella Typhi chimaeric A2B5 typhoid toxin. Nature 499, 350–354 (2013). This paper solves the crystal structure of TT, showing its novel A2B5 arrangement and its similarity to PT.
Lang, A. E. et al. Photorhabdus luminescens toxins ADP-ribosylate actin and RhoA to force actin clustering. Science 327, 1139–1142 (2010). This study identifies the substrates of the TcC components of P. luminescens toxin complex proteins and shows that these toxins promote actin polymerization.
Blackburn, M., Golubeva, E., Bowen, D. & Ffrench-Constant, R. H. A. Novel insecticidal toxin from Photorhabdus luminescens, toxin complex a (Tca), and its histopathological effects on the midgut of Manduca sexta. Appl. Environ. Microbiol. 64, 3036–3041 (1998).
Han, S. & Tainer, J. The ARTT motif and a unified structural understanding of substrate recognition in ADP-ribosylating bacterial toxins and eukaryotic ADP-ribosyltransferases. Int. J. Med. Microbiol. 291, 523–529 (2002). This paper proposes that toxins could be identified by a core ARTT motif and suggests that substrate specificity is dependent on external protein structure.
Wilson, B. A., Reich, K. A., Weinstein, B. R. & Collier, R. J. Active-site mutations of diphtheria toxin: effects of replacing glutamic acid-148 with aspartic acid, glutamine, or serine. Biochemistry 29, 8643–8651 (1990).
Carroll, S. F. & Collier, R. J. NAD binding site of diphtheria toxin: identification of a residue within the nicotinamide subsite by photochemical modification with NAD. Proc. Natl Acad. Sci. USA 81, 3307–3311 (1984).
Carroll, S. F. & Collier, R. J. Active site of Pseudomonas aeruginosa exotoxin A. Glutamic acid 553 is photolabeled by NAD and shows functional homology with glutamic acid 148 of diphtheria toxin. J. Biol. Chem. 262, 8707–8711 (1987).
Douglas, D. & Collier, R. Exotoxin A of Pseudomonas aeruginosa: substitution of glutamic acid 553 with aspartic acid drastically reduces toxicity and enzymatic activity. J. Bacteriol. 169, 4967–4971 (1987).
Bell, C. E. & Eisenberg, D. Crystal structure of diphtheria toxin bound to nicotinamide adenine dinucleotide. Biochemistry 35, 1137–1149 (1996).
Oliver, A. W. et al. Crystal structure of the catalytic fragment of murine poly(ADP–ribose) polymerase-2. Nucleic Acids Res. 32, 456–464 (2004).
Ruf, A., Mennissier de Murcia, J., de Murcia, G. & Schulz, G. E. Structure of the catalytic fragment of poly(AD-ribose) polymerase from chicken. Proc. Natl Acad. Sci. USA 93, 7481–7485 (1996).
Tsuge, H. et al. Crystal structure and site-directed mutagenesis of enzymatic components from Clostridium perfringens iota-toxin. J. Mol. Biol. 325, 471–483 (2003).
Domenighini, M. & Rappuoli, R. Three conserved consensus sequences identify the NAD-binding site of ADP-ribosylating enzymes, expressed by eukaryotes, bacteria and T-even bacteriophages. Mol. Microbiol. 21, 667–674 (1996).
Roux Jr., E. & Yersin, A. Contribution a l'etude de la diphtherie. Ann. Inst. Pasteur 2, 620–629 (1888).
Eaton, M. D. The purification and concentration of diphtheria toxin. J. Bacteriol. 31, 347 (1936).
Pappenheimer, A. M. Jr. Diphtheria toxin: I. isolation and characterization of a toxic protein from C. diphtheriae filtrates. J. Biol. Chem. 125, 543–553 (1937).
Freeman, V. J. Studies on the virulence of bacteriophage infected strains of C. diphtheriae. J. Bacteriol. 61, 675–688 (1951).
Naglich, J. G., Metherall, J. E., Russell, D. W. & Eidels, L. Expression cloning of a diphtheria toxin receptor: identity with a heparin-binding EGF-like growth factor precursor. Cell 69, 1051–1061 (1992).
Louie, G. V., Yang, W., Bowman, M. E. & Choe, S. Crystal structure of the complex of diphtheria toxin with an extracellular fragment of its receptor. Mol. Cell 1, 67–78 (1997).
Lemichez, E. et al. Membrane translocation of diphtheria toxin fragment A exploits early to late endosome trafficking machinery. Mol. Microbiol. 23, 445–457 (1997).
Gordon, V. M., Klimpel, K. R., Arora, N., Henderson, M. A. & Leppla, S. H. Proteolytic activation of bacterial toxins by eukaryotic cells is performed by furin and by additional cellular proteases. Infect. Immun. 63, 82–87 (1995).
Sandvig, K. & Olsnes, S. Diphtheria toxin entry into cells is facilitated by low pH. J. Cell Biol. 87, 828–832 (1980).
Draper, R. K. & Simon, M. I. The entry of diphtheria toxin into the mammalian cell cytoplasm: evidence for lysosomal involvement. J. Cell Biol. 87, 849–854 (1980).
Collier, R. J. Effect of diphtheria toxin on protein synthesis: inactivation of one of the transfer factors. J. Mol. Biol. 25, 83–98 (1967).
Honjo, T., Nishizuka, Y., Hayaishi, O. & Kato, I. Diphtheria toxin-dependent adenosine diphosphate ribosylation of aminoacyl transferase II and inhibition of protein synthesis. J. Biol. Chem. 243, 3553–3555 (1968).
Van Ness, B. G., Howard, J. B. & Bodley, J. W. ADP-ribosylation of elongation factor 2 by diphtheria toxin. Isolation and properties of the novel ribosyl-amino acid and its hydrolysis products. J. Biol. Chem. 255, 10717–10720 (1980).
Su, X., Lin, Z. & Lin, H. The biosynthesis and biological function of diphthamide. Crit. Rev. Biochem. Mol. Biol. 48, 515–521 (2013).
Strauss, N. & Hendee, E. D. The effect of diphtheria toxin on the metabolism of HeLa cells. J. Exp. Med. 109, 145–163 (1959).
Taylor, D. et al. Critical movements of a single diphthamide residue of eukaryotic elongation factor 2 monitored by cryo-EM. Microsc. Microanalysis 13, 388–389 (2007).
Liu, S. et al. Diphthamide modification on eukaryotic elongation factor 2 is needed to assure fidelity of mRNA translation and mouse development. Proc. Natl Acad. Sci. USA 109, 13817–13822 (2012).
Gupta, P. K., Liu, S., Batavia, M. P. & Leppla, S. H. The diphthamide modification on elongation factor-2 renders mammalian cells resistant to ricin. Cell. Microbiol. 10, 1687–1694 (2008).
Gill, D. M. Bacterial toxins: a table of lethal amounts. Microbiol. Rev. 46, 86–94 (1982).
Carroll, S. F. & Collier, R. J. Amino acid sequence homology between the enzymic domains of diphtheria toxin and Pseudomonas aeruginosa exotoxin A. Mol. Microbiol. 2, 293–296 (1988).
Collier, R. J. & McKay, D. B. Crystallization of exotoxin A from Pseudomonas aeruginosa. J. Mol. Biol. 157, 413–415 (1982).
Allured, V. S., Collier, R. J., Carroll, S. F. & McKay, D. B. Structure of exotoxin A of Pseudomonas aeruginosa at 3.0-Angstrom resolution. Proc. Natl Acad. Sci. USA 83, 1320–1324 (1986).
Kounnas, M. Z. et al. The α2-macroglobulin receptor/low density lipoprotein receptor-related protein binds and internalizes Pseudomonas exotoxin A. J. Biol. Chem. 267, 12420–12423 (1992).
Ogata, M., Fryling, C. M., Pastan, I. & FitzGerald, D. J. Cell-mediated cleavage of Pseudomonas exotoxin between Arg279 and Gly280 generates the enzymatically active fragment which translocates to the cytosol. J. Biol. Chem. 267, 25396–25401 (1992).
Fitzgerald, D., Morris, R. E. & Saelinger, C. B. Receptor-mediated internalization of Pseudomonas toxin by mouse fibroblasts. Cell 21, 867–873 (1980).
Weldon, J. E. & Pastan, I. A guide to taming a toxin — recombinant immunotoxins constructed from Pseudomonas exotoxin A for the treatment of cancer. FEBS J. 278, 4683–4700 (2011).
McKee, M. L. & FitzGerald, D. J. Reduction of furin-nicked Pseudomonas exotoxin A: an unfolding story. Biochemistry 38, 16507–16513 (1999).
Iglewski, B. H., Liu, P. V. & Kabat, D. Mechanism of action of Pseudomonas aeruginosa exotoxin A: adenosine diphosphate ribosylation of mammalian elongation factor 2 in vitro and in vivo. Infection Immun. 15, 138–144 (1977).
Faruque, S. M. et al. Genetic diversity and virulence potential of environmental Vibrio cholerae population in a cholera-endemic area. Proc. Natl Acad. Sci. USA 101, 2123–2128 (2004).
Colwell, R. R. Global climate and infectious disease: the cholera paradigm. Science 274, 2025–2031 (1996).
Purdy, A. E. et al. Diversity and distribution of cholix toxin, a novel ADP-ribosylating factor from Vibrio cholerae. Environ. Microbiol. Rep. 2, 198–207 (2010).
Chen, Y., Johnson, J. A., Pusch, G. D., Morris, J. G. & Stine, O. C. The genome of non-O1 Vibrio cholerae NRT36S demonstrates the presence of pathogenic mechanisms that are distinct from those of O1 Vibrio cholerae. Infection Immun. 75, 2645–2647 (2007). This is the first study to identify chxA in strains of Vibrio cholerae that do not express CT using sequence analysis.
Awasthi, S. P. et al. Novel cholix toxin variants, ADP-ribosylating toxins in Vibrio cholerae non-O1/non-O139 strains, and their pathogenicity. Infect. Immun. 81, 531–541 (2013).
Jørgensen, R. et al. Cholix toxin, a novel ADP-ribosylating factor from Vibrio cholerae. J. Biol. Chem. 283, 10671–10678 (2008). This paper characterizes the structure of ChxA and shows its similarity to other DT-like toxins. It also proposes a new mechanism for DT-like toxin ADP-ribosylation activity.
Fieldhouse, R. J., Jørgensen, R., Lugo, M. R. & Merrill, A. R. The 1.8 Å cholix toxin crystal structure in complex with NAD+ and evidence for a new kinetic model. J. Biol. Chem. 287, 21176–21188 (2012).
Stover, C. K. et al. Complete genome sequence of Pseudomonas aeruginosa PAO1, an opportunistic pathogen. Nature 406, 959–964 (2000).
Koch, R. Sechster Bericht der deutschen Wissenschaftlichen Commission zur Enforschung der Cholera. Dtsch Med Wochenschr 10, 191–192 (in German) (1884).
Howard-Jones, N. Robert Koch and the cholera vibrio: a centenary. BMJ 288, 379–381 (1984).
De, S. N. Enterotoxicity of bacteria-free culture-filtrate of Vibrio cholerae. Nature 183, 1533–1534 (1959).
Lencer, W. I., Hirst, T. R. & Holmes, R. K. Membrane traffic and the cellular uptake of cholera toxin. Biochim. Biophys. Acta 1450, 177–190 (1999).
Eidels, L., Proia, R. & Hart, D. Membrane receptors for bacterial toxins. Microbiol. Rev. 47, 596–620 (1983).
Ewers, H. & Helenius, A. Lipid-mediated endocytosis. Cold Spring Harb. Perspect. Biol. 3, a004721 (2011).
Bobak, D. et al. Mechanism of activation of cholera toxin by ADP-ribosylation factor (ARF): both low- and high-affinity interactions of ARF with guanine nucleotides promote toxin activation. Biochemistry 29, 855–861 (1990).
O'Neal, C. J., Jobling, M. G., Holmes, R. K. & Hol, W. G. J. Structural basis for the activation of cholera toxin by human ARF6-GTP. Science 309, 1093–1096 (2005).
Kahn, R. A. & Gilman, A. G. The protein cofactor necessary for ADP-ribosylation of Gs by cholera toxin is itself a GTP binding protein. J. Biol. Chem. 261, 7906–7911 (1986).
Gill, D. M. & Meren, R. ADP-ribosylation of membrane proteins catalyzed by cholera toxin: basis of the activation of adenylate cyclase. Proc. Natl Acad. Sci. USA 75, 3050–3054 (1978).
Cassel, D. & Pfeuffer, T. Mechanism of cholera toxin action: covalent modification of the guanyl nucleotide-binding protein of the adenylate cyclase system. Proc. Natl Acad. Sci. USA 75, 2669–2673 (1978).
Moss, J. Activation of adenylate cyclase by heat-labile Escherichia coli enterotoxin: evidence for ADP-ribosyl transferase activity similar to that of choleragen. J. Clin. Invest. 62, 281–285 (1978).
Chang, P. P., Moss, J., Twiddy, E. M. & Holmes, R. K. Type II heat-labile enterotoxin of Escherichia coli activates adenylate cyclase in human fibroblasts by ADP ribosylation. Infect. Immun. 55, 1854–1858 (1987).
Kantor, H. S., Tao, P. & Wisdom, C. Action of Escherichia coli enterotoxin: adenylate cyclase behavior of intestinal epithelial cells in culture. Infecti. Immun. 9, 1003–1010 (1974).
World Health Organization. Immunization, Vaccines and Biologicals: Pertussis [online], (WHO, 2011).
Tamura, M. et al. Subunit structure of islet-activating protein, pertussis toxin, in conformity with the A–B model. Biochemistry 21, 5516–5522 (1982).
Stein, P. E. et al. The crystal structure of pertussis toxin. Structure 2, 45–57 (1994).
Armstrong, G. D., Howard, L. A. & Peppler, M. S. Use of glycosyltransferases to restore pertussis toxin receptor activity to asialoagalactofetuin. J. Biol. Chem. 263, 8677–8684 (1988).
Stein, P. et al. Structure of a pertussis toxin–sugar complex as a model for receptor binding. Nature Struct. Biol. 1, 591–596 (1994).
Brennan, M. J., David, J. L., Kenimer, J. G. & Manclark, C. R. Lectin-like binding of pertussis toxin to a 165-kilodalton Chinese hamster ovary cell glycoprotein. J. Biol. Chem. 263, 4895–4899 (1988).
el Bayâ, A., Linnemann, R., von Olleschik-Elbheim, L., Robenek, H. & Schmidt, M. Endocytosis and retrograde transport of pertussis toxin to the Golgi complex as a prerequisite for cellular intoxication. Eur. J. Cell Biol. 73, 40–48 (1997).
Burns, D. L. & Manclark, C. R. Role of cysteine 41 of the A subunit of pertussis toxin. J. Biol. Chem. 264, 564–568 (1989).
Antoine, R. & Locht, C. Roles of the disulfide bond and the carboxy-terminal region of the S1 subunit in the assembly and biosynthesis of pertussis toxin. Infection Immun. 58, 1518–1526 (1990).
Bokoch, G. M., Katada, T., Northup, J. K., Hewlett, E. L. & Gilman, A. G. Identification of the predominant substrate for ADP-ribosylation by islet activating protein. J. Biol. Chem. 258, 2072–2075 (1983).
Katada, T. & Ui, M. Direct modification of the membrane adenylate cyclase system by islet-activating protein due to ADP-ribosylation of a membrane protein. Proc. Natl Acad. Sci. USA 79, 3129–3133 (1982).
Kurose, H., Katada, T., Amano, T. & Ui, M. Specific uncoupling by islet-activating protein, pertussis toxin, of negative signal transduction via α-adrenergic, cholinergic, and opiate receptors in neuroblastoma x glioma hybrid cells. J. Biol. Chem. 258, 4870–4875 (1983).
Simpson, L. L., Stiles, B. G., Zepeda, H. H. & Wilkins, T. D. Molecular basis for the pathological actions of Clostridium perfringens iota toxin. Infect. Immun. 55, 118–122 (1987).
Perelle, S., Gibert, M., Bourlioux, P., Corthier, G. & Popoff, M. R. Production of a complete binary toxin (actin-specific ADP-ribosyltransferase) by Clostridium difficile CD196. Infect. Immun. 65, 1402–1407 (1997).
Simpson, L. L., Stiles, B. G., Zepeda, H. & Wilkins, T. D. Production by Clostridium spiroforme of an iotalike toxin that possesses mono(ADP-ribosyl)transferase activity: identification of a novel class of ADP-ribosyltransferases. Infect. Immun. 57, 255–261 (1989).
Aktories, K., Lang, A. E., Schwan, C. & Mannherz, H. G. Actin as target for modification by bacterial protein toxins. FEBS J. 278, 4526–4543 (2011).
Young, J. A. T. & Collier, R. J. Anthrax toxin: receptor binding, internalization, pore formation, and translocation. Annu. Rev. Biochem. 76, 243–265 (2007).
Papatheodorou, P. et al. Lipolysis-stimulated lipoprotein receptor (LSR) is the host receptor for the binary toxin Clostridium difficile transferase (CDT). Proc. Natl Acad. Sci. USA 108, 16422–16427 (2011).
Han, S., Craig, J. A., Putnam, C. D., Carozzi, N. B. & Tainer, J. A. Evolution and mechanism from structures of an ADP-ribosylating toxin and NAD complex. Nature Struct. Biol. 6, 932–936 (1999).
Vandekerckhove, J., Schering, B., Bärmann, M. & Aktories, K. Clostridium perfringens iota toxin ADP-ribosylates skeletal muscle actin in Arg-177. FEBS Lett. 225, 48–52 (1987).
Vandekerckhove, J., Schering, B., Bärmann, M. & Aktories, K. Botulinum C2 toxin ADP-ribosylates cytoplasmic β/γ-actin in arginine 177. J. Biol. Chem. 263, 696–700 (1988).
Gülke, I. et al. Characterization of the enzymatic component of the ADP-ribosyltransferase toxin CDTa from Clostridium difficile. Infect. Immun. 69, 6004–6011 (2001).
Gerding, D. N., Johnson, S., Rupnik, M. & Aktories, K. Clostridium difficile binary toxin CDT: mechanism, epidemiology, and potential clinical importance. Gut Microbes 5, 6–18 (2014).
Schwan, C. et al. Clostridium difficile toxin CDT induces formation of microtubule-based protrusions and increases adherence of bacteria. PLoS Pathog. 5, e1000626 (2009).
Schwan, C. et al. Clostridium difficile toxin CDT hijacks microtubule organization and reroutes vesicle traffic to increase pathogen adherence. Proc. Natl Acad. Sci. USA 111, 2313–2318 (2014).
Lesnick, M. L., Reiner, N. E., Fierer, J. & Guiney, D. G. The Salmonella spvB virulence gene encodes an enzyme that ADP-ribosylates actin and destabilizes the cytoskeleton of eukaryotic cells. Mol. Microbiol. 39, 1464–1470 (2001).
Otto, H. et al. The spvB gene-product of the Salmonella enterica virulence plasmid is a mono(ADP-ribosyl)transferase. Mol. Microbiol. 37, 1106–1115 (2000). This paper shows that the spvB gene is similar to other mono-ADP-ribosyltransferases using PSI-BLAST protein homology searches.
Braun, M. et al. Characterization of an ADP-ribosyltransferase toxin (AexT) from Aeromonas salmonicida subsp. salmonicida. J. Bacteriol. 184, 1851–1858 (2002). This study uses bioinformatics to show that AexT is similar to ExoS, using sequence similarity, and characterizes the cytotoxic activity of AexT.
Fehr, D. et al. Aeromonas exoenzyme T of Aeromonas salmonicida is a bifunctional protein that targets the host cytoskeleton. J. Biol. Chem. 282, 28843–28852 (2007). This paper identifies AexT as a bifunctional toxin and characterizes its ADP-ribosyltransferase substrates.
Visschedyk, D. D. et al. Photox, a novel actin-targeting mono-ADP-ribosyltransferase from Photorhabdus luminescens. J. Biol. Chem. 285, 13525–13534 (2010). This paper uses bioinformatics to identify photox using amino acid homology with the previously identified toxin SpvB; it shows structural and functional similarity between photox and other actin-ADP-ribosylating toxins.
Hochmann, H., Pust, S., von Figura, G., Aktories, K. & Barth, H. Salmonella enterica SpvB ADP-ribosylates actin at position arginine-177. Characterization of the catalytic domain within the SpvB protein and a comparison to binary clostridial actin-ADP-ribosylating toxins. Biochemistry 45, 1271–1277 (2006). This paper shows that SpvB has amino acid residues that are conserved in bacterial actin-ADP-ribosylating toxins, despite not being a binary toxin. It also functionally confirms this conservation by demonstrating actin ADP-ribosylation at Arg177.
Vilches, S. et al. Aeromonas hydrophila AH-3 AexT is an ADP-ribosylating toxin secreted through the type III secretion system. Microb. Pathog. 44, 1–12 (2008).
Barbieri, J. T. & Sun, J. in Reviews of Physiology, Biochemistry and Pharmacology Vol. 152 (eds Aktories, K. & Just, I.) 79–92 (Springer Berlin Heidelberg, 2005).
Coburn, J., Dillon, S. T., Iglewski, B. H. & Gill, D. M. Exoenzyme S of Pseudomonas aeruginosa ADP-ribosylates the intermediate filament protein vimentin. Infection Immun. 57, 996–998 (1989).
Coburn, J., Wyatt, R. T., Iglewski, B. H. & Gill, D. M. Several GTP-binding proteins, including p21c-H-ras, are preferred substrates of Pseudomonas aeruginosa exoenzyme S. J. Biol. Chem. 264, 9004–9008 (1989).
Litvak, Y. & Selinger, Z. Aeromonas salmonicida toxin AexT has a Rho family GTPase-activating protein domain. J. Bacteriol. 189, 2558–2560 (2007).
Rubin, E. J., Gill, D. M., Boquet, P. & Popoff, M. R. Functional modification of a 21-kilodalton G protein when ADP-ribosylated by exoenzyme C3 of Clostridium botulinum. Mol. Cell. Biol. 8, 418–426 (1988).
Wilde, C., Vogelsgesang, M. & Aktories, K. Rho-specific Bacillus cereus ADP-ribosyltransferase C3cer cloning and characterization. Biochemistry 42, 9694–9702 (2003).
Hanna, S. & El-Sibai, M. Signaling networks of Rho GTPases in cell motility. Cell Signall. 25, 1955–1961 (2013).
Heasman, S. J. & Ridley, A. J. Mammalian Rho GTPases: new insights into their functions from in vivo studies. Nature Rev. Mol. Cell Biol. 9, 690–701 (2008).
Sekine, A., Fujiwara, M. & Narumiya, S. Asparagine residue in the rho gene product is the modification site for botulinum ADP-ribosyltransferase. J. Biol. Chem. 264, 8602–8605 (1989).
Genth, H., Schmidt, M., Gerhard, R., Aktories, K. & Just, I. Activation of phospholipase D1 by ADP-ribosylated RhoA. Biochem. Biophys. Res. Commun. 302, 127–132 (2003).
Genth, H. et al. Entrapment of Rho ADP-ribosylated by Clostridium botulinum C3 exoenzyme in the Rho-guanine nucleotide dissociation inhibitor-1 complex. J. Biol. Chem. 278, 28523–28527 (2003).
Barth, H. et al. Neosynthesis and activation of Rho by Escherichia coli cytotoxic necrotizing factor (CNF1) reverse cytopathic effects of ADP-ribosylated Rho. J. Biol. Chem. 274, 27407–27414 (1999).
Pautsch, A., Vogelsgesang, M., Tränkle, J., Herrmann, C. & Aktories, K. Crystal structure of the C3bot–RalA complex reveals a novel type of action of a bacterial exoenzyme. EMBO J. 24, 3670–3680 (2005).
Wilde, C., Chhatwal, G. S., Schmalzing, G., Aktories, K. & Just, I. A. Novel C3-like ADP-ribosyltransferase from Staphylococcus aureus. Modifying RhoE and Rnd3. J. Biol. Chem. 276, 9537–9542 (2001).
Molinari, G. et al. Localization of the C3-Like ADP-ribosyltransferase from Staphylococcus aureus during bacterial invasion of mammalian cells. Infect. Immun. 74, 3673–3677 (2006).
Fahrer, J. et al. Selective and specific internalization of clostridial C3 ADP-ribosyltransferases into macrophages and monocytes. Cell. Microbiol. 12, 233–247 (2010).
Wollein Waldetoft, K. & Råberg, L. To harm or not to harm? On the evolution and expression of virulence in group A streptococci. Trends Microbiol. 22, 7–13 (2014).
Ferretti, J. J. et al. Complete genome sequence of an M1 strain of Streptococcus pyogenes. Proc. Natl Acad. Sci. USA 98, 4658–4663 (2001). This paper proposes, on the basis of sequence data, that the spyA gene encodes a C3-like ADP-ribosyltransferase.
Coye, L. H. & Collins, C. M. Identification of SpyA, a novel ADP-ribosyltransferase of Streptococcus pyogenes. Mol. Microbiol. 54, 89–98 (2004). This study characterizes the ADP-ribosyltransferase activity of SpyA and shows that, despite its predicted C3-like activity, SpyA has activity towards different eukaryotic substrates.
Korotkova, N. et al. SpyA is a membrane-bound ADP-ribosyltransferase of Streptococcus pyogenes which modifies a streptococcal peptide, SpyB. Mol. Microbiol. 83, 936–952 (2012).
Ivaska, J., Pallari, H.-M., Nevo, J. & Eriksson, J. E. Novel functions of vimentin in cell adhesion, migration, and signaling. Exp. Cell Res. 313, 2050–2062 (2007).
Eckes, B. et al. Impaired wound healing in embryonic and adult mice lacking vimentin. J. Cell Sci. 113, 2455–2462 (2000).
Icenogle, L. M. et al. Molecular and biological characterization of streptococcal SpyA-mediated ADP-ribosylation of intermediate filament protein vimentin. J. Biol. Chem. 287, 21481–21491 (2012).
Gohara, R. et al. Phosphorylation of vimentin head domain inhibits interaction with the carboxyl-terminal end of α-helical rod domain studied by surface plasmon resonance measurements. FEBS Lett. 489, 182–186 (2001).
Hoff, J. S., DeWald, M., Moseley, S. L., Collins, C. M. & Voyich, J. M. SpyA, a C3-Like ADP-ribosyltransferase, contributes to virulence in a mouse subcutaneous model of Streptococcus pyogenes infection. Infection Immun. 79, 2404–2411 (2011).
Guttman, D. S. et al. A functional screen for the type III (Hrp) secretome of the plant pathogen Pseudomonas syringae. Science 295, 1722–1726 (2002).
Petnicki-Ocwieja, T. et al. Genomewide identification of proteins secreted by the Hrp type III protein secretion system of Pseudomonas syringae pv. tomato DC3000. Proc. Natl Acad. Sci. USA 99, 7652–7657 (2002).
Lindeberg, M. et al. Closing the circle on the discovery of genes encoding Hrp regulon members and type III secretion system effectors in the genomes of three model Pseudomonas syringae strains. Mol. Plant Microbe Interact. 19, 1151–1158 (2006).
Fouts, D. E. et al. Genomewide identification of Pseudomonas syringae pv. tomato DC3000 promoters controlled by the HrpL alternative sigma factor. Proc. Natl Acad. Sci. USA 99, 2275–2280 (2002).
Buell, C. R. et al. The complete genome sequence of the Arabidopsis and tomato pathogen Pseudomonas syringae pv. tomato DC3000. Proc. Natl Acad. Sci. USA 100, 10181–10186 (2003).
Fu, Z. Q. et al. A type III effector ADP-ribosylates RNA-binding proteins and quells plant immunity. Nature 447, 284–288 (2007). This paper identifies the substrate of effector HopU1 ADP-ribosyltransferase activity and shows the unique immunosuppressive phenotype of HopU1 activity.
Jeong, B.R. et al. Structure function analysis of an ADP-ribosyltransferase type III effector and its RNA-binding target in plant immunity. J. Biol. Chem. 286, 43272–43281 (2011). This study solves the crystal structure of HopU1 and shows that unique loops that frame the NAD-binding pocket are necessary for substrate recognition.
Sun, J. & Barbieri, J. T. Pseudomonas aeruginosa ExoT ADP-ribosylates CT10 regulator of kinase (Crk) proteins. J. Biol. Chem. 278, 32794–32800 (2003).
Sun, J., Maresso, A. W., Kim, J. J. & Barbieri, J. T. How bacterial ADP-ribosylating toxins recognize substrates. Nature Struct. Mol. Biol. 11, 868–876 (2004).
Heintzen, C., Nater, M., Apel, K. & Staiger, D. AtGRP7, a nuclear RNA-binding protein as a component of a circadian-regulated negative feedback loop in Arabidopsis thaliana. Proc. Natl Acad. Sci. USA 94, 8515–8520 (1997).
Nicaise, V. et al. Pseudomonas HopU1 modulates plant immune receptor levels by blocking the interaction of their mRNAs with GRP7. EMBO J. 32, 701–712 (2013).
Jessen, T. H., Oubridge, C., Teo, C. H., Pritchard, C. & Nagai, K. Identification of molecular contacts between the U1, a small nuclear ribonucleoprotein and U1 RNA. EMBO J. 10, 3447–3456 (1991).
Burd, C. & Dreyfuss, G. Conserved structures and diversity of functions of RNA-binding proteins. Science 265, 615–621 (1994).
Nagai, K., Oubridge, C., Jessen, T. H., Li, J. & Evans, P. R. Crystal structure of the RNA-binding domain of the U1 small nuclear ribonucleoprotein A. Nature 348, 515–520 (1990).
Lee, A. L. et al. Chemical shift mapping of the RNA-binding interface of the multiple-RBD protein sex-lethal. Biochemistry 36, 14306–14317 (1997).
Schöning, J. C. et al. Auto-regulation of the circadian slave oscillator component AtGRP7 and regulation of its targets is impaired by a single RNA recognition motif point mutation. Plant J. 52, 1119–1130 (2007).
Boller, T. & He, S. Y. Innate immunity in plants: an arms race between pattern recognition receptors in plants and effectors in microbial pathogens. Science 324, 742–744 (2009).
Dodds, P. N. & Rathjen, J. P. Plant immunity: towards an integrated view of plant–pathogen interactions. Nature Rev. Genet. 11, 539–548 (2010).
Segonzac, C. & Zipfel, C. Activation of plant pattern-recognition receptors by bacteria. Curr. Opin. Microbiol. 14, 54–61 (2011).
Maurice, J. A first step in bringing typhoid fever out of the closet. Lancet 379, 699–700 (2012).
Haghjoo, E. & Galán, J. E. Salmonella Typhi encodes a functional cytolethal distending toxin that is delivered into host cells by a bacterial-internalization pathway. Proc. Natl Acad. Sci. USA 101, 4614–4619 (2004).
Spanò, S., Ugalde, J. E. & Galán, J. E. Delivery of a Salmonella Typhi exotoxin from a host intracellular compartment. Cell Host Microbe 3, 30–38 (2008).
Guidi, R. et al. Salmonella enterica delivers its genotoxin through outer membrane vesicles secreted from infected cells. Cell. Microbiol. 15, 2034–2050 (2013).
Forst, S., Dowds, B., Boemare, N. & Stackebrandt, E. Xenorhabdus and Photorhabdus spp.:bugs that kill bugs. Annu. Rev. Microbiol. 51, 47–72 (1997).
Waterfield, N., Hares, M., Ffrench-Constant, R., Wren, B. & Hinchliffe, S. in The Genus Yersinia (eds Perry, R. & Fetherston, J.) 247–257 (Springer, 2007).
Waterfield, N. R., Ciche, T. & Clarke, D. Photorhabdus and a host of hosts. Annu. Rev. Microbiol. 63, 557–574 (2009).
Waterfield, N. R., Bowen, D. J., Fetherston, J. D., Perry, R. D. & Ffrench-Constant, R. H. The tc genes of Photorhabdus: a growing family. Trends Microbiol. 9, 185–191 (2001).
Ffrench-Constant, R. & Waterfield, N . in Advances in Applied Microbiology (eds Laskin, A. I., Bennett, J. W., Gadd, G. M. & Sariaslani, S.) 169–183 (Academic Press, 2005).
Gatsogiannis, C. et al. A syringe-like injection mechanism in Photorhabdus luminescens toxins. Nature 495, 520–523 (2013).
Ffrench-Constant, R. et al. Photorhabdus: towards a functional genomic analysis of a symbiont and pathogen. FEMS Microbiol. Rev. 26, 433–456 (2003).
Cassimeris, L., Safer, D., Nachmias, V. T. & Zigmond, S. H. Thymosin β4 sequesters the majority of G-actin in resting human polymorphonuclear leukocytes. J. Cell Biol. 119, 1261–1270 (1992).
Mannherz, H. G. & Hannappel, E. The β-thymosins: intracellular and extracellular activities of a versatile actin binding protein family. Cell. Motil. Cytoskeleton 66, 839–851 (2009).
Vetter, I. R. & Wittinghofer, A. The guanine nucleotide-binding switch in three dimensions. Science 294, 1299–1304 (2001).
Meusch, D. et al. Mechanism of Tc toxin action revealed in molecular detail. Nature 508, 61–65 (2014).
Falkow, S. Molecular Koch's postulates applied to microbial pathogenicity. Rev. Infect. Dis. 10 (Suppl. 2) 274–276 (1988). This paper explains the molecular application of Koch's postulates by Falkow for confirmation of the role of putative virulence factors in bacterial pathogenesis.
Falkow, S. Molecular Koch's postulates applied to bacterial pathogenicity — a personal recollection 15 years later. Nature Rev. Microbiol. 2, 67–72 (2004).
Zheng, L.-L. et al. A comparison of computational methods for identifying virulence factors. PLoS ONE 7, e42517 (2012). This paper shows the functionality and importance of protein amino acid sequence analyses and explains caveats and issues with common search methods.
Jones, D. T. & Swindells, M. B. Getting the most from PSI-BLAST. Trends Biochem. Sci. 27, 161–164 (2002).
Qi, Y., Sadreyev, R. I., Wang, Y., Kim, B. H. & Grishin, N. V. A comprehensive system for evaluation of remote sequence similarity detection. BMC Bioinformatics 8, 314 (2007).
Palmiter, R. D. et al. Cell lineage ablation in transgenic mice by cell-specific expression of a toxin gene. Cell 50, 435–443 (1987).
Saito, M. et al. Diphtheria toxin receptor-mediated conditional and targeted cell ablation in transgenic mice. Nature Biotech. 19, 746–750 (2001).
Buch, T. et al. A Cre-inducible diphtheria toxin receptor mediates cell lineage ablation after toxin administration. Nature Meth. 2, 419–426 (2005).
Chaudhary, V. K. et al. A recombinant immunotoxin consisting of two antibody variable domains fused to Pseudomonas exotoxin. Nature 339, 394–397 (1989).
Pastan, I. Immunotoxins containing Pseudomonas exotoxin A: a short history. Cancer Immunol. Immunother. 52, 338–341 (2003).
Kreitman, R. J. & Pastan, I. Antibody fusion proteins: anti-CD22 recombinant immunotoxin moxetumomab pasudotox. Clin. Cancer Res. 17, 6398–6405 (2011).
Kreitman, R. J. et al. Efficacy of the anti-CD22 recombinant immunotoxin BL22 in chemotherapy-resistant hairy-cell leukemia. New Engl. J. Med. 345, 241–247 (2001).
Chaudhary, V. K. et al. Selective killing of HIV-infected cells by recombinant human CD4-Pseudomonas exotoxin hybrid protein. Nature 335, 369–372 (1988).
Berger, E. A. & Pastan, I. Immunotoxin complementation of HAART to deplete persisting HIV-infected cell reservoirs. PLoS Pathog. 6, e1000803 (2010).
Sarnovsky, R. et al. Initial characterization of an immunotoxin constructed from domains II and III of cholera exotoxin. Cancer Immunol. Immunother. 59, 737–746 (2010).
Francis, J. W. et al. Enhancement of diphtheria toxin potency by replacement of the receptor binding domain with tetanus toxin C-fragment. J. Neurochem. 74, 2528–2536 (2000).
Ahnert-Hilger, G. et al. Differential effects of Rho GTPases on axonal and dendritic development in hippocampal neurones. J. Neurochem. 90, 9–18 (2004).
Bertrand, J., Di Polo, A. & McKerracher, L. Enhanced survival and regeneration of axotomized retinal neurons by repeated delivery of cell-permeable C3-like Rho antagonists. Neurobiol. Dis. 25, 65–72 (2007).
Höltje, M. et al. A 29-amino acid fragment of Clostridium botulinum C3 protein enhances neuronal outgrowth, connectivity, and reinnervation. FASEB J. 23, 1115–1126 (2009).
Bertrand, J., Winton, M. J., Rodriguez-Hernandez, N., Campenot, R. B. & McKerracher, L. Application of Rho antagonist to neuronal cell bodies promotes neurite growth in compartmented cultures and regeneration of retinal ganglion cell axons in the optic nerve of adult rats. J. Neurosci. 25, 1113–1121 (2005).
Boato, F. et al. C3 peptide enhances recovery from spinal cord injury by improved regenerative growth of descending fiber tracts. J. Cell Sci. 123, 1652–1662 (2010).
Acknowledgements
N.C.S. and J.T.B. are supported by a US National Institutes of Health (NIH) grant (grant number NIH AI30162). K.A. is supported by the Deutsche Forschungsgemeinschaft (AK6/23-1 and AK6//22-2)
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
J.T.B. is a consultant for Syntaxin Ltd. N.C.S. and K.A. declare no competing interests.
Glossary
- SN1 strain-alleviation mechanism
-
An electrophilic reaction mechanism that starts with the hydrolytic release of nicotinamide from the NAD donor, followed by formation of an oxocarbenium cation intermediate in a 'strained' conformation. Rotation around the phosphodiester bond forms a second oxocarbenium cation, which results in strain relief and moving of the ribose moiety near to the acceptor residue to complete the ADP-ribose transfer.
- P-site
-
(Peptidyl site). The site on the small ribosomal subunit that holds the tRNA molecule that is linked to the growing end of the polypeptide chain.
- Retrograde trafficking
-
Trafficking of vesicles in a direction from the host cell surface to the ER; for example, trafficking from the Golgi complex to the ER.
- ER-associated degradation
-
(ERAD). A cellular pathway that targets misfolded proteins in the ER for ubiquitylation and subsequent degradation by the proteasome.
- Intermediate filament
-
A filament that is formed by coiled-coil-rich cytoskeletal proteins, such as keratin.
- Pattern recognition receptors
-
(PRRs). Soluble or membrane-associated receptors that are displayed by the metazoan host and can recognize complex molecular patterns on the surface of microorganisms.
- Autocrine
-
A term used to describe the activation of cellular receptors on the same cell that produces the ligand.
- Paracrine
-
A term used to describe the activation of cellular receptors on cells adjacent to the cell that produces the ligand.
Rights and permissions
About this article
Cite this article
Simon, N., Aktories, K. & Barbieri, J. Novel bacterial ADP-ribosylating toxins: structure and function. Nat Rev Microbiol 12, 599–611 (2014). https://doi.org/10.1038/nrmicro3310
Published:
Issue Date:
DOI: https://doi.org/10.1038/nrmicro3310
This article is cited by
-
Molecular basis of threonine ADP-ribosylation of ubiquitin by bacterial ARTs
Nature Chemical Biology (2024)
-
Pathogenic Factors of Plant Pathogenic Streptomyces
Potato Research (2024)
-
Enzymology of extracellular NAD metabolism
Cellular and Molecular Life Sciences (2021)
-
The C. difficile toxin B membrane translocation machinery is an evolutionarily conserved protein delivery apparatus
Nature Communications (2020)
-
Structural and biochemical evidence supporting poly ADP-ribosylation in the bacterium Deinococcus radiodurans
Nature Communications (2019)