Genetic Control of Immune Response to Recombinant Antigens Carried by an Attenuated Salmonella typhimurium Vaccine Strain: Nramp1 Influences T-Helper Subset Responses and Protection against Leishmanial Challenge

Attenuated strains of Salmonella typhimurium have been widely used as vehicles for delivery and expression of vaccine antigens in murine models of infectious disease. In mice, early bacterial replication following infection with S. typhimurium is controlled by the gene (Nramp1, formerly Ity/Lsh/Bcg) encoding the natural-resistance-associated macrophage protein (Nramp1). Nramp1 regulates macrophage activation and has multiple pleiotropic effects, including regulation of tumor necrosis factor alpha, interleukin 1β (IL-1β), and major histocompatibility complex class II molecules, all of which influence antigen processing and presentation. Nramp1 also has a direct effect on antigen processing, possibly by regulating the activity of proteases in the late endosomal compartment. Hence, there are multiple ways (regulation of bacterial load or recombinant antigen dose, class II molecule expression, costimulatory or adjuvant activity, and antigen processing) that Nramp1 might influence responses to recombinant salmonella vaccines. To test the hypothesis that Nramp1 influences responses to vaccination, congenic mouse strains have been used to analyze immune responses to recombinant antigens (tetanus toxoid antigen and leishmanial gp63) carried by live attenuated S. typhimurium aroA aroD mutants. Results show that congenic mice carrying the wild-type (S. typhimurium resistance) Nramp1 allele mount a predominantly T-helper-1 (IL-2 and gamma interferon) response to vaccination and show enhanced resolution of lesions following challenge infection with Leishmania major. In contrast, mice carrying mutant (S. typhimurium susceptibility) Nramp1 mount a T-helper-2 (immunoglobulin E and IL-4) response and show exacerbated lesion growth upon challenge.Attenuated strains of Salmonella have been widely used as vehicles for delivery and expression of a heterogeneous range of antigens (Ag) from other pathogens (for a review, see reference 18). In murine models of infectious disease, genetically engineered Salmonella typhimurium has been used to induce immunity to viral (influenza virus A [40]), bacterial (tetanus toxin [10], Streptococcus pyogenes M1 protein [31], and Francisella tularensis [36]), protozoan (Leishmania major [44, 45] and Plasmodium yoelii circumsporozoite protein [33]), and helminth (schistosomiasis [22, 23]) Ag. Since a similar vaccine strategy (e.g., attenuated Salmonella typhi aro mutants in human trials [16, 39]) might ultimately be adopted for use in genetically diverse human populations, it is important to investigate the influence that host genetics might have on the ability to induce protective immune responses to recombinant salmonella vaccines. While major histocompatibility complex (MHC) class I and class II molecule genes with polymorphisms will be obvious candidate genes because of their ability to restrict vaccine responses to certain antigenic epitopes, a role for non-MHC genes acting independently of Ag specificity should also be considered.In mice, early bacterial replication following infection with S. typhimurium is regulated by the gene (Nramp1, formerly Ity/Lsh/Bcg) encoding the natural-resistance-associated macrophage protein (Nramp1) (for reviews, see references 4, 6, and 37). Nramp1 was identified by positional cloning (42). Gene knockout was then used (41) to formally demonstrate that the gene (renamed Nramp1) mediated resistance to all three intramacrophage pathogens S. typhimurium (Ity), Leishmania donovani (Lsh), and Mycobacterium bovis (Bcg). The protein (Nramp1) encoded by Nramp1 regulates the cascade of gene-inductive events which follow interaction of macrophages with bacterial lipopolysaccharide (LPS) and/or natural killer- or T-cell-derived gamma interferon (IFN-γ). The gene has multiple pleiotropic effects, including regulation of tumor necrosis factor alpha (TNF-α), interleukin 1β (IL-1β), and MHC class II molecules. All of these influence Ag processing and presentation, either directly (class II) or indirectly through their costimulatory or adjuvant (IL-1β and TNF-α) activity. Recent studies also demonstrate that Nramp1 has a more direct effect on Ag processing, possibly by regulating the activity of proteases in the late endosomal compartment (26). Hence, there are multiple ways (regulation of bacterial load or recombinant Ag dose, class II molecule expression, costimulatory or adjuvant activity, and Ag processing) that Nramp1 might influence responses to recombinant salmonella vaccines.To test the hypothesis that Nramp1 influences responses to vaccination, congenic mouse strains have been used to analyze immune responses to recombinant Ag (tetanus toxoid Ag and leishmanial gp63) carried by live attenuated S. typhimurium aroA aroD mutants. Results show that congenic mice carrying the wild-type (S. typhimurium resistance) Nramp1 allele mount a predominantly T-helper-1 (IL-2 and IFN-γ) response to vaccination and show enhanced resolution of lesions following challenge infection with L. major. In contrast, mice carrying mutant (S. typhimurium susceptibility) Nramp1 mount a T-helper-2 (immunoglobulin E [IgE] and IL-4) response and show exacerbated lesion growth upon challenge.