title: Legionella pneumophila OxyR Is a Redundant Transcriptional Regulator That Contributes to Expression Control of the Two-Component CpxRA System.
abstract: Nominally an environmental organism,Legionella pneumophilais an intracellular parasite of protozoa but is also the causative agent of the pneumonia termed Legionnaires' disease, which results from inhalation of aerosolized bacteria by susceptible humans. Coordination of gene expression by a number of identified regulatory factors, including OxyR, assistsL. pneumophilain adapting to the stresses of changing environments.L. pneumophilaOxyR (OxyRLp) is an ortholog ofEscherichia coliOxyR; however, OxyRLpwas shown elsewhere to be functionally divergent, such that it acts as a transcription regulator independently of the oxidative stress response. In this study, the use of improved gene deletion methods has enabled us to generate an unmarked in-frame deletion ofoxyRinL. pneumophilaLack of OxyRLpdid not affectin vitrogrowth or intracellular growth inAcanthamoeba castellaniiprotozoa and U937-derived macrophages. The expression of OxyRLpdoes not appear to be regulated by CpxR, even though purified recombinant CpxR bound a DNA sequence similar to that reported for CpxR elsewhere. Surprisingly, a lack of OxyRLpresulted in elevated activity of the promoters located upstream oficmRand thelpg1441-cpxAoperon, and OxyRLpdirectly bound to these promoter regions, suggesting that OxyRLpis a direct repressor. Interestingly, a strain overexpressing OxyRLpdemonstrated reduced intracellular growth inA. castellaniibut not in U937-derived macrophages, suggesting that balanced expression control of the two-component CpxRA system is necessary for survival in protozoa. Taken together, this study suggests that OxyRLpis a functionally redundant transcriptional regulator inL. pneumophilaunder the conditions evaluated herein.IMPORTANCELegionella pneumophilais an environmental pathogen, with its transmission to the human host dependent upon its ability to replicate in protozoa and survive within its aquatic niche. Understanding the genetic factors that contribute toL. pneumophilasurvival within each of these unique environments will be key to limiting future point-source outbreaks of Legionnaires' disease. The transcriptional regulatorL. pneumophilaOxyR (OxyRLp) has been previously identified as a potential regulator of virulence traits warranting further investigation. This study demonstrated thatoxyRis nonessential forL. pneumophilasurvivalin vitroandin vivovia mutational analysis. While the mechanisms of how OxyRLpexpression is regulated remain elusive, this study shows that OxyRLpnegatively regulates the expression of thecpxRAtwo-component system necessary for intracellular survival in protozoa.
authors.affiliation: Department of Microbiology, Faculty of Science, University of Manitoba, Winnipeg, Manitoba, Canada.
authors.forename: Jennifer R
authors.initials: JR
authors.lastname: Tanner
authors.affiliation: Department of Microbiology, Faculty of Science, University of Manitoba, Winnipeg, Manitoba, Canada.
authors.forename: Palak G
authors.initials: PG
authors.lastname: Patel
authors.affiliation: Department of Microbiology, Faculty of Science, University of Manitoba, Winnipeg, Manitoba, Canada.
authors.forename: Jacqueline R
authors.initials: JR
authors.lastname: Hellinga
authors.affiliation: Department of Microbiology, Faculty of Science, University of Manitoba, Winnipeg, Manitoba, Canada.
authors.forename: Lynda J
authors.initials: LJ
authors.lastname: Donald
authors.affiliation: Department of Microbiology, Faculty of Science, University of Manitoba, Winnipeg, Manitoba, Canada.
authors.forename: Celine
authors.initials: C
authors.lastname: Jimenez
authors.affiliation: Department of Pathology, Medicine, and Microbiology & Immunology, Faculty of Medicine, Dalhousie University, Halifax, Nova Scotia, Canada.|Division of Microbiology, Department of Pathology and Laboratory Medicine, Nova Scotia Health Authority (NSHA), Halifax, Nova Scotia, Canada.
authors.forename: Jason J
authors.initials: JJ
authors.lastname: LeBlanc
authors.affiliation: Department of Microbiology, Faculty of Science, University of Manitoba, Winnipeg, Manitoba, Canada Ann.Brassinga@umanitoba.ca.
authors.forename: Ann Karen C
authors.initials: AKC
authors.lastname: Brassinga
mesh_terms: D048248:Acanthamoeba castellanii / Q000382:microbiology; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D006801:Humans; D016952:Legionella pneumophila / Q000235:genetics / Q000378:metabolism*; D008264:Macrophages / Q000382:microbiology; D009154:Mutation; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D020298:U937 Cells
chemical_list: D001426:Bacterial Proteins; D012097:Repressor Proteins; D014157:Transcription Factors
keywords: CpxRA; Legionella pneumophila; OxyR; gene expression; intracellular pathogen; microbial genetics; protozoa; transcriptional regulation