title: Identification of a conserved membrane localization domain within numerous large bacterial protein toxins.
abstract: Vibrio cholerae is the causative agent of the diarrheal disease cholera. Many virulence factors contribute to intestinal colonization and disease including the Multifunctional Autoprocessing RTX toxin (MARTX(Vc)). The Rho-inactivation domain (RID) of MARTX(Vc) is responsible for inactivating the Rho-family of small GTPases, which leads to depolymerization of the actin cytoskeleton. Based on a deletion analysis of RID to determine the minimal functional domain, we have identified a subdomain at the N terminus of RID that is homologous to the membrane targeting C1 domain of Pasteurella multocida toxin. A GFP fusion to this subdomain from RID colocalized with a plasma membrane marker when transiently expressed within HeLa cells and can be found in the membrane fraction following subcellular fractionation. This C1-like subdomain is present in multiple families of bacterial toxins, including all of the clostridial glucosyltransferase toxins and various MARTX toxins. GFP-fusions to these homologous domains are also membrane associated, indicating that this is a conserved membrane localization domain (MLD). We have identified three residues (Y23, S68, R70) as necessary for proper localization of one but not all MLDs. In addition, we found that substitution of the RID MLD with the MLDs from two different effector domains from the Vibrio vulnificus MARTX toxin restored RID activity, indicating that there is functional overlap between these MLDs. This study describes the initial recognition of a family of conserved plasma membrane-targeting domains found in multiple large bacterial toxins.
authors.affiliation: Department of Microbiology-Immunology, Northwestern University, Chicago, IL 60611, USA.
authors.forename: Brett
authors.initials: B
authors.lastname: Geissler
authors.forename: Rehman
authors.initials: R
authors.lastname: Tungekar
authors.forename: Karla J F
authors.initials: KJ
authors.lastname: Satchell
mesh_terms: D000595:Amino Acid Sequence; D001427:Bacterial Toxins / Q000737:chemistry* / Q000235:genetics / Q000633:toxicity; D002462:Cell Membrane / Q000737:chemistry; D017124:Conserved Sequence; D005816:Genetic Complementation Test; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000633:toxicity; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D055672:Static Electricity; D014734:Vibrio cholerae / Q000737:chemistry* / Q000235:genetics / Q000472:pathogenicity*
chemical_list: D001427:Bacterial Toxins; D011993:Recombinant Fusion Proteins; C434464:RtxA protein, Vibrio cholerae; D049452:Green Fluorescent Proteins