title: Toxicity of an α-pore-forming toxin depends on the assembly mechanism on the target membrane as revealed by single molecule imaging.
abstract: α-Pore-forming toxins (α-PFTs) are ubiquitous defense tools that kill cells by opening pores in the target cell membrane. Despite their relevance in host/pathogen interactions, very little is known about the pore stoichiometry and assembly pathway leading to membrane permeabilization. Equinatoxin II (EqtII) is a model α-PFT from sea anemone that oligomerizes and forms pores in sphingomyelin-containing membranes. Here, we determined the spatiotemporal organization of EqtII in living cells by single molecule imaging. Surprisingly, we found that on the cell surface EqtII did not organize into a unique oligomeric form. Instead, it existed as a mixture of oligomeric species mostly including monomers, dimers, tetramers, and hexamers. Mathematical modeling based on our data supported a new model in which toxin clustering happened in seconds and proceeded via condensation of EqtII dimer units formed upon monomer association. Furthermore, altering the pathway of EqtII assembly strongly affected its toxic activity, which highlights the relevance of the assembly mechanism on toxicity.
authors.affiliation: Max Planck Institute for Intelligent Systems, Heisenbergstr. 3, 70569 Stuttgart, Germany; German Cancer Research Center, Bioquant, Im Neuenheimer Feld 267, 69120 Heidelberg, Germany.
authors.forename: Yamunadevi
authors.initials: Y
authors.lastname: Subburaj
authors.affiliation: Center for Protein Studies, Faculty of Biology, Calle 25 #455, Plaza de la Revolución, La Habana, Cuba.
authors.forename: Uris
authors.initials: U
authors.lastname: Ros
authors.affiliation: Max Planck Institute for Intelligent Systems, Heisenbergstr. 3, 70569 Stuttgart, Germany; German Cancer Research Center, Bioquant, Im Neuenheimer Feld 267, 69120 Heidelberg, Germany,; Interfaculty Institute of Biochemistry, University of Tübingen, Hoppe-Seyler-Str. 4, 72076 Tübingen, Germany.
authors.forename: Eduard
authors.initials: E
authors.lastname: Hermann
authors.affiliation: Max Planck Institute for Intelligent Systems, Heisenbergstr. 3, 70569 Stuttgart, Germany.
authors.forename: Rudi
authors.initials: R
authors.lastname: Tong
authors.affiliation: Max Planck Institute for Intelligent Systems, Heisenbergstr. 3, 70569 Stuttgart, Germany; German Cancer Research Center, Bioquant, Im Neuenheimer Feld 267, 69120 Heidelberg, Germany,; Interfaculty Institute of Biochemistry, University of Tübingen, Hoppe-Seyler-Str. 4, 72076 Tübingen, Germany. Electronic address: ana.garcia@uni-tuebingen.de.
authors.forename: Ana J
authors.initials: AJ
authors.lastname: García-Sáez
mesh_terms: D002463:Cell Membrane Permeability; D003064:Cnidarian Venoms / Q000737:chemistry*; D004910:Erythrocyte Membrane / Q000737:chemistry*; D006461:Hemolysis; D006801:Humans; D052899:Pore Forming Cytotoxic Proteins / Q000737:chemistry*; D055503:Protein Multimerization
chemical_list: D003064:Cnidarian Venoms; D052899:Pore Forming Cytotoxic Proteins; C012307:equinatoxin
keywords: Actinoporins; Cell Death; Equinatoxin II; Membrane Biophysics; Membrane Protein; Permeability; Toxin