The immediate goal of this project is to define the cellular and biochemical functions of 57 uncharacterized open reading frames (ORFs), hypothetical proteins, and small non-coding RNAs in B. abortus. Our preliminary data provide evidence that these genes have a functional role in general stress adaptation and chronic mammalian infection. General stress response (GSR)-dependent transcription in B. abortus is controlled by the sigma factor, SigEI. Our preliminary investigation of SigEI and its upstream signaling partner, PhyR have demonstrated that these regulatory proteins are required for adaptation to oxidative and acid stress in vitro, and for maintenance of chronic infection in a BALB/c murine model. We have experimentally defined a set of 108 genes that are directly or indirectiy regulated by SigEI. Within this regulated gene set are 32 uncharacterized open reading frames (ORFs) and 9 small non-coding RNAs. Additionally, we have identified 16 hypothetical genes that are adjacent to, or in apparent operons with SigEI-regulated ORFs, but for which we have no evidence of expression. We propose to: 1) Test strains in which these genes have been deleted in oxidative and acid stress survival assays, and in cell-based and animal infection assays, 2) elucidate the biochemical functions of these hypothetical proteins and SigEI-regulated ORFs in collaboration with out Biochemical Function Technology Core, and 3) characterize the biochemical function of putative non-coding RNAs under transcriptional control of SigEI. These experiments will provide a comprehensive (in vivo and in vitro) functional understanding of conserved and non-conserved B. abortus genes that are currently uncharacterized.
Brucella spp. are the causative agents of brucellosis, which is among the most common zooneses globally. Due to their high infectivity, easy aerosolization, and debilitating effects on infected individuals. Brucella spp. have been classified as a category B bioterror threat by the U.S. Centers for Disease Control (CDC). Experiments proposed herein will define the biochemical function of uncharacterized genes implicated in regulation of B abortus stress physiology and in the control of chronic brucellosis disease.
|CzyÅ¼, Daniel M; Jain-Gupta, Neeta; Shuman, Howard A et al. (2016) A dual-targeting approach to inhibit Brucella abortus replication in human cells. Sci Rep 6:35835|
|Meyer, Peter A; Socias, Stephanie; Key, Jason et al. (2016) Data publication with the structural biology data grid supports live analysis. Nat Commun 7:10882|
|Herrou, Julien; CzyÅ¼, Daniel M; Willett, Jonathan W et al. (2016) WrpA Is an Atypical Flavodoxin Family Protein under Regulatory Control of the Brucella abortus General Stress Response System. J Bacteriol 198:1281-93|
|Baric, Ralph S; Crosson, Sean; Damania, Blossom et al. (2016) Next-Generation High-Throughput Functional Annotation of Microbial Genomes. MBio 7:|
|Willett, Jonathan W; Herrou, Julien; CzyÅ¼, Daniel M et al. (2016) Brucella abortus Î”rpoE1 confers protective immunity against wild type challenge in a mouse model of brucellosis. Vaccine 34:5073-81|
|Willett, Jonathan W; Herrou, Julien; Briegel, Ariane et al. (2015) Structural asymmetry in a conserved signaling system that regulates division, replication, and virulence of an intracellular pathogen. Proc Natl Acad Sci U S A 112:E3709-18|
|Kim, Hye-Sook; Willett, Jonathan W; Jain-Gupta, Neeta et al. (2014) The Brucella abortus virulence regulator, LovhK, is a sensor kinase in the general stress response signalling pathway. Mol Microbiol 94:913-25|
|Tam, Christina; Demke, Owen; Hermanas, Timothy et al. (2014) YfbA, a Yersinia pestis regulator required for colonization and biofilm formation in the gut of cat fleas. J Bacteriol 196:1165-73|
|Herrou, Julien; Crosson, Sean (2013) Molecular structure of the Brucella abortus metalloprotein RicA, a Rab2-binding virulence effector. Biochemistry 52:9020-8|