title: A cellular metalloproteinase activates Vibrio cholerae pro-cytolysin.
abstract: Many strains of Vibrio cholerae produce a cytolysin (VCC) that forms oligomeric transmembrane pores in animal cells. The molecule is secreted as a procytolysin (pro-VCC) of 79 kDa that must be cleaved at the N terminus to generate the active 65-kDa toxin. Processing can occur in solution, and previous studies have described the action of mature VCC thus generated. However, little is known about the properties of pro-VCC itself. In this study, it is shown that pro-VCC exist as a monomer in solution and binds as a monomer to eukaryotic cells. Bound pro-VCC can then be activated either by exogenous, extracellular, or by endogenous, cell-bound proteases. In both cases, cleavage generates the 65-kDa VCC that oligomerizes to form transmembrane pores. A wide variety of exogenous proteinases can mediate activation. In contrast, the activating cellular protease is selectively inhibited by the hydroxamate inhibitor TAPI, and thus probable candidates are members of the ADAM-metalloproteinase family. Furin, MMP-2, MMP-9, and serine proteinases were excluded. Cells over-expressing ADAM-17, also known as tumor necrosis factor alpha converting enzyme, displayed increased activation of VCC, and knockout cells lacking ADAM-17 had a markedly decreased capacity to cleave the protoxin. The possibility is raised that pro-VCC is targeted to membrane sites that selectively contain or are accessible to cellular ADAM-metalloproteinases. Although many microbial toxins are activated by furin, this is the first evidence for processing by a cellular metalloproteinase. We identified ADAM-17 as a potent activator of pro-VCC.
authors.affiliation: Institute of Medical Microbiology and Hygiene, University of Mainz, D55101 Mainz, Germany. avaleva@uni-mainz.de
authors.forename: Angela
authors.initials: A
authors.lastname: Valeva
authors.forename: Ivan
authors.initials: I
authors.lastname: Walev
authors.forename: Silvia
authors.initials: S
authors.lastname: Weis
authors.forename: Fatima
authors.initials: F
authors.lastname: Boukhallouk
authors.forename: Trudy M
authors.initials: TM
authors.lastname: Wassenaar
authors.forename: Kristina
authors.initials: K
authors.lastname: Endres
authors.forename: Falk
authors.initials: F
authors.lastname: Fahrenholz
authors.forename: Sucharit
authors.initials: S
authors.lastname: Bhakdi
authors.forename: Alexander
authors.initials: A
authors.lastname: Zitzer
mesh_terms: D051722:ADAM Proteins; D000072198:ADAM17 Protein; D000818:Animals; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism; D002772:Cholera Toxin / Q000378:metabolism*; D006224:Cricetinae; D003603:Cytotoxins / Q000378:metabolism*; D006801:Humans; D008666:Metalloendopeptidases / Q000502:physiology*; D051379:Mice; D011498:Protein Precursors / Q000378:metabolism*; D011817:Rabbits
chemical_list: D003603:Cytotoxins; D011498:Protein Precursors; D002772:Cholera Toxin; D051722:ADAM Proteins; D008666:Metalloendopeptidases; D000072198:ADAM17 Protein; C000606462:ADAM17 protein, human; C000606463:Adam17 protein, mouse