title: The Yersinia enterocolitica Ysa type III secretion system is expressed during infections both in vitro and in vivo.
abstract: Yersinia enterocolitica biovar 1B maintains two type III secretion systems (T3SS) that are involved in pathogenesis, the plasmid encoded Ysc T3SS and the chromosomally encoded Ysa T3SS. In vitro, the Ysa T3SS has been shown to be expressed only at 26°C in a high-nutrient medium containing an exceptionally high concentration of salt - an artificial condition that provides no clear insight on the nature of signal that Y. enterocolitica responds to in a host. However, previous research has indicated that the Ysa system plays a role in the colonization of gastrointestinal tissues of mice. In this study, a series of Ysa promoter fusions to green fluorescent protein gene (gfp) were created to analyze the expression of this T3SS during infection. Using reporter strains, infections were carried out in vitro using HeLa cells and in vivo using the mouse model of yersiniosis. Expression of green fluorescent protein (GFP) was measured from the promoters of yspP (encoding a secreted effector protein) and orf6 (encoding a structural component of the T3SS apparatus) in vitro and in vivo. During the infection of HeLa cells GFP intensity was measured by fluorescence microscopy, while during murine infections GFP expression in tissues was measured by flow cytometry. These approaches, combined with quantification of yspP mRNA transcripts by quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR), demonstrate that the Ysa system is expressed in vitro in a contact-dependent manner, and is expressed in vivo during infection of mice.
authors.affiliation: Microbiology Graduate Group, University of California, Davis, CA; Systems Biology, Sandia National Laboratories, Livermore, CA.
authors.forename: Zachary W
authors.initials: ZW
authors.lastname: Bent
authors.forename: Steven S
authors.initials: SS
authors.lastname: Branda
authors.forename: Glenn M
authors.initials: GM
authors.lastname: Young
mesh_terms: D000818:Animals; D019677:Artificial Gene Fusion; D001426:Bacterial Proteins / Q000096:biosynthesis*; D058947:Bacterial Secretion Systems; D004195:Disease Models, Animal; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008856:Microscopy, Fluorescence; D060888:Real-Time Polymerase Chain Reaction; D015009:Yersinia Infections / Q000382:microbiology; D015008:Yersinia enterocolitica / Q000502:physiology*
chemical_list: D001426:Bacterial Proteins; D058947:Bacterial Secretion Systems; D049452:Green Fluorescent Proteins
keywords: In vivo expression; T3SS; Y. enterocolitica; Ysa; Ysp