UPR determinants of Brucella virulence

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PROJECT SUMMARY Brucellosis remains the most prevalent zoonosis worldwide, incurring significant human morbidity and economic impact. No safe human vaccine exists. The species most often implicated in human disease, B. melitensis has proven refractory to eradication. Following ingestion or inhalation, Brucella invade macrophages and other phagocytes, and traffic within vacuoles to the endoplasmic reticulum (ER) where they establish a replicative niche. During this process, Brucella induce a host stress response known as the “Unfolded Protein Response” (UPR). The UPR enables cells to cope with ER stress by transiently decreasing protein production and expanding ER capacity. However, if ER stress is prolonged or profound, the UPR initiates apoptosis. We have noted a striking correlation between extent of UPR induction and human virulence. In contrast to B. abortus and the vaccine strain S19, B. melitensis (the most virulent species) induces marked upregulation of the UPR target gene C/EBP homologous protein (CHOP) in murine and human macrophages. In certain bacterial infections, CHOP enhances virulence. CHOP critically contributed to apoptosis and abortion in a Brucella murine model. CHOP also stimulates the production of pro-inflammatory cytokines (e.g. IL-23) that could lead to human disease manifestations such as spondyloarthritis. Currently, the role of CHOP in Brucella infection of macrophages and spondyloarthritis is unknown, and there are no appropriate disease models to address this specific question. We hypothesize that the induction of a more extensive UPR by B. melitensis contributes to the virulence and pathogenicity of this species. To gain a greater understanding of the critical interactions between Brucella, host UPR and the development of arthritis, we propose the following aims: Aim 1: Test the hypothesis that CHOP contributes to the virulence of B. melitensis via greater bacterial replication and induction of specific host inflammatory responses. We will confirm our preliminary findings of species-specific UPR induction and expand upon these results by comparing macrophage gene expression induced by B. melitensis vs. B. abortus. Further, ddit3 (CHOP)-/- macrophages will delineate the portion of differential gene expression due to CHOP. We will determine if CHOP regulates apoptosis, replication or cell-cell spread of B. melitensis. Finally, the requirement for CHOP in B. melitensis replication and host inflammatory responses will be determined in vivo. Aim 2: Establish a mouse model of Brucella spondyloarthritis using spondyloarthritis-susceptible mice. Within recent years, several murine spondyloarthritis models have been developed that require either cytokine or infectious triggers, namely the SKG-ZAP-70 mutant model and B10.RIII mice. We will systematically test these mice strains to establish a robust, tractable model of Brucella-induced spondyloarthritis that will enable testing of our overarching hypothesis. An increasing number of pathogens have been recognized to manipulate the UPR. This work will contribute to the understanding of how the UPR mediates the relationships between bacterial invaders, host responses and disease manifestations.