Internalization of Clostridium botulinum C2 Toxin Is Regulated by Cathepsin B Released from Lysosomes
Abstract
1. Introduction
2. Results
2.1. C2 Toxin-Induced Lysosomal Exocytosis
2.2. Lysosomal Proteases Are Needed for Internalization of C2 Toxin
2.3. Effects of Cathepsin siRNAs on C2-Toxin-Induced Cytotoxicity
3. Discussion
4. Conclusions
5. Materials and Methods
5.1. Materials
5.2. Cell Cultures and Cytotoxicity Assays
5.3. β-hexosaminidase Assays
5.4. Determination of Acidic Sphingomyelinase Activities
5.5. Small Interfering RNA Silencing and Western Blots
5.6. Immunofluorescence Analysis
5.7. Statistical Analyses
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
- Aktories, K.; Schwan, C.; Papatheodorou, P.; Lang, A.E. Bidirectional attack on the actin cytoskeleton. Bacterial protein toxins causing polymerization or depolymerization of actin. Toxicon 2012, 60, 572–581. [Google Scholar] [CrossRef] [PubMed]
- Stiles, B.G.; Pradhan, K.; Fleming, J.M.; Samy, R.P.; Barth, H.; Popoff, M.R. Clostridium and bacillus binary enterotoxins: Bad for the bowels, and eukaryotic being. Toxins 2014, 6, 2626–2656. [Google Scholar] [CrossRef] [PubMed]
- Takehara, M.; Takagishi, T.; Seike, S.; Oda, M.; Sakaguchi, Y.; Hisatsune, J.; Ochi, S.; Kobayashi, K.; Nagahama, M. Cellular entry of Clostridium perfringens iota-toxin and Clostridium botulinum C2 toxin. Toxins 2017, 9, 247. [Google Scholar] [CrossRef] [PubMed]
- Knapp, O.; Benz, R.; Popoff, M.R. Pore-forming activity of clostridial binary toxins. Biochim. Biophys. Acta 2015, 1858, 512–525. [Google Scholar] [CrossRef]
- Eckhardt, M.; Barth, H.; Blöcker, D.; Aktories, K. Binding of Clostridium botulinum C2 toxin to asparagine-linked complex and hybrid carbohydrates. J. Biol. Chem. 2000, 275, 2328–2334. [Google Scholar] [CrossRef]
- Nagahama, M.; Hagiyama, T.; Kojima, T.; Aoyanagi, K.; Takahashi, C.; Oda, M.; Sakaguchi, Y.; Oguma, K.; Sakurai, J. Binding and internalization of Clostridium botulinum C2 toxin. Infect. Immun. 2009, 77, 5139–5148. [Google Scholar] [CrossRef]
- Nagahama, M.; Takahashi, C.; Aoyanagi, K.; Tashiro, R.; Kobayashi, K.; Sakaguchi, Y.; Ishidoh, K.; Sakurai, J. Intracellular trafficking of Clostridium botulinum C2 toxin. Toxicon 2014, 82, 76–82. [Google Scholar] [CrossRef]
- Nagahama, M.; Takehara, M.; Takagishi, T.; Seike, S.; Miyamoto, K.; Kobayashi, K. Cellular uptake of Clostridium botulinum C2 toxin requires acid sphingomyelinase activity. Infect. Immun. 2017, 85, e00966-16. [Google Scholar] [CrossRef]
- Idone, V.; Tam, C.; Goss, J.W.; Toomre, D.; Pypaert, M.; Andrews, N.W. Repair of injured plasma membrane by rapid Ca2+-dependent endocytosis. J. Cell Biol. 2008, 180, 905–914. [Google Scholar] [CrossRef]
- Tam, C.; Idone, V.; Devlin, C.; Fernandes, M.C.; Flannery, A.; He, X.; Schuchman, E.; Tabas, I.; Andrews, N.W. Exocytosis of acid sphingomyelinase by wounded cells promotes endocytosis and plasma membrane repair. J. Cell Biol. 2010, 189, 1027–1038. [Google Scholar] [CrossRef]
- Corrotte, M.; Fernandes, M.C.; Tam, C.; Andrews, N.W. Toxin pores endocytosed during plasma membrane repair traffic into the lumen of MVBs for degradation. Traffic 2012, 13, 483–494. [Google Scholar] [CrossRef]
- Andrews, N.W.; Corrotte, M.; Castro-Gomes, T. Above the fray: Surface remodeling by secreted lysosomal enzymes leads to endocytosis-mediated plasma membrane repair. Semin. Cell Dev. Biol. 2015, 45, 10–17. [Google Scholar] [CrossRef]
- Castro-Gomes, T.; Corrotte, M.; Tam, C.; Andrews, N.W. Plasma membrane repair is regulated extracellularly by proteases released from lysosomes. PLoS ONE 2016, 11, e0152583. [Google Scholar] [CrossRef]
- Brito, C.; Cabanes, D.; Sarmento Mesquita, F.; Sousa, S. Mechanisms protecting host cells against bacterial pore-forming toxins. Cell. Mol. Life Sci. 2019, 76, 1319–1339. [Google Scholar] [CrossRef]
- Benes, P.; Vetvicka, V.; Fusek, M. Cathepsin D--many functions of one aspartic protease. Crit. Rev. Oncol. Hematol. 2008, 68, 12–28. [Google Scholar] [CrossRef]
- Braulke, T.; Bonifacino, J.S. Sorting of lysosomal proteins. Biochim. Biophys. Acta 2009, 1793, 605–614. [Google Scholar] [CrossRef]
- Heinrich, M.; Wickel, M.; Schneider-Brachert, W.; Sandberg, C.; Gahr, J.; Schwandner, R.; Weber, T.; Saftig, P.; Peters, C.; Brunner, J.; et al. Cathepsin D targeted by acid sphingomyelinase-derived ceramide [published correction appears in EMBO J 2000 Jan 17;19(2):315]. EMBO J. 1999, 18, 5252–5263. [Google Scholar] [CrossRef]
- Heinrich, M.; Wickel, M.; Winoto-Morbach, S.; Schneider-Brachert, W.; Weber, T.; Brunner, J.; Saftig, P.; Peters, C.; Krönke, M.; Schütze, S. Ceramide as an activator lipid of cathepsin D. Adv. Exp. Med. Biol. 2000, 477, 305–315. [Google Scholar]
- Fonović, M.; Turk, B. Cysteine cathepsins and extracellular matrix degradation. Biochim. Biophys. Acta 2014, 1840, 2560–2570. [Google Scholar] [CrossRef]
- Diment, S.; Leech, M.S.; Stahl, P.D. Cathepsin D is membrane-associated in macrophage endosomes. J. Biol. Chem. 1988, 263, 6901–6907. [Google Scholar] [CrossRef]
- Kanda, Y. Investigation of the freely available easy-to-use software ‘EZR’ for medical statistics. Bone Marrow Transplant 2013, 48, 452–458. [Google Scholar] [CrossRef]
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. |
© 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
Share and Cite
Nagahama, M.; Kobayashi, K.; Ochi, S.; Takehara, M. Internalization of Clostridium botulinum C2 Toxin Is Regulated by Cathepsin B Released from Lysosomes. Toxins 2021, 13, 272. https://doi.org/10.3390/toxins13040272
Nagahama M, Kobayashi K, Ochi S, Takehara M. Internalization of Clostridium botulinum C2 Toxin Is Regulated by Cathepsin B Released from Lysosomes. Toxins. 2021; 13(4):272. https://doi.org/10.3390/toxins13040272
Chicago/Turabian StyleNagahama, Masahiro, Keiko Kobayashi, Sadayuki Ochi, and Masaya Takehara. 2021. "Internalization of Clostridium botulinum C2 Toxin Is Regulated by Cathepsin B Released from Lysosomes" Toxins 13, no. 4: 272. https://doi.org/10.3390/toxins13040272
APA StyleNagahama, M., Kobayashi, K., Ochi, S., & Takehara, M. (2021). Internalization of Clostridium botulinum C2 Toxin Is Regulated by Cathepsin B Released from Lysosomes. Toxins, 13(4), 272. https://doi.org/10.3390/toxins13040272