title: A conserved alpha-helix essential for a type VI secretion-like system of Francisella tularensis.
abstract: Francisella tularensis harbors genes with similarity to genes encoding components of a type VI secretion system (T6SS) recently identified in several gram-negative bacteria. These genes include iglA and iglB encoding IglA and IglB, homologues of which are conserved in most T6SSs. We used a yeast two-hybrid system to study the interaction of the Igl proteins of F. tularensis LVS. We identified a region of IglA, encompassing residues 33 to 132, necessary for efficient binding to IglB, as well as for IglAB protein stability and intramacrophage growth. In particular, residues 103 to 122, overlapping a highly conserved alpha-helix, played an absolutely essential role. Point mutations within this domain caused modest defects in IglA-IglB binding in the yeast Saccharomyces cerevisiae but markedly impaired intramacrophage replication and phagosomal escape, resulting in severe attenuation of LVS in mice. Thus, IglA-IglB complex formation is clearly crucial for Francisella pathogenicity. This interaction may be universal to type VI secretion, since IglAB homologues of Yersinia pseudotuberculosis, Pseudomonas aeruginosa, Vibrio cholerae, Salmonella enterica serovar Typhimurium, and Escherichia coli were also shown to interact in yeast, and the interaction was dependent on preservation of the same alpha-helix. Heterologous interactions between nonnative IglAB proteins further supported the notion of a conserved binding site. Thus, IglA-IglB complex formation is clearly crucial for Francisella pathogenicity, and the same interaction is conserved in other human pathogens.
authors.affiliation: Department of Clinical Microbiology, Clinical Bacteriology, Umeå University, SE-901 85 Umeå, Sweden. jeanette.broms@climi.umu.se
authors.forename: Jeanette E
authors.initials: JE
authors.lastname: Bröms
authors.forename: Moa
authors.initials: M
authors.lastname: Lavander
authors.forename: Anders
authors.initials: A
authors.lastname: Sjöstedt
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D002460:Cell Line; D017124:Conserved Sequence; D005604:Francisella tularensis / Q000235:genetics / Q000472:pathogenicity*; D005816:Genetic Complementation Test; D008264:Macrophages / Q000382:microbiology; D026901:Membrane Transport Proteins / Q000235:genetics* / Q000378:metabolism*; D051379:Mice; D011485:Protein Binding; D025941:Protein Interaction Mapping; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011550:Pseudomonas aeruginosa / Q000235:genetics; D012441:Saccharomyces cerevisiae; D012486:Salmonella typhimurium / Q000235:genetics; D020798:Two-Hybrid System Techniques; D014734:Vibrio cholerae / Q000235:genetics; D014774:Virulence; D015011:Yersinia pseudotuberculosis / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D026901:Membrane Transport Proteins