Bacterial Virulence Regulation and Targets of CD8+ T Cell Immunity During Listeria monocytogenes Infection

Abstract

Listeria monocytogenes (Lm) is an intracellular bacterial pathogen that causes a severe invasive disease in humans. Upon infection, Lm must sense its presence within a host and upregulate bacterial genes necessary for productive infection. One of the Lm factors involved in this response is VirR, the response regulator component of a two-component regulatory system. VirR is necessary for in vivo virulence and Lm innate resistance to antimicrobial peptides. We found VirR function depends on the presence of an ABC-transporter that we have named VirAB. Deletion of virAB led to a decrease in transcription of VirR-regulated genes. The ΔvirAB mutant also exhibited similar defects in intracellular spread and in vivo virulence to a ΔvirR strain. To investigate the role of VirAB in antimicrobial resistance, we determined the sensitivity of ΔvirAB bacteria to nisin and bacitracin. We found that expression of VirAB was necessary for nisin resistance, but not resistance to bacitracin. We propose that VirAB is a sensor required for VirR signaling during host infection and exposure to nisin. Since exposure to serum and blood has been shown to upregulate virulence genes in Lm, we performed a transposon mutagenesis screen to identify genes involved in Lm serum sensing. The genes identified from this screen included virR, virA, and the VirR-regulated operon dltABCD. We show that VirR and DltABCD are required for the upregulation of the principle Lm virulence determinants LLO and ActA in response to growth in serum, and that this induction primarily takes place post-transcriptionally. Clearance of Lm infection is primarily mediated by CD8+ T cells. Of the thousands of potential T cell antigens produced by Lm, protective CD8+ T cells appear to target only a few secreted proteins. Despite decades of study as a model pathogen for cell-mediated immunity, only a handful of Lm-specific CD8+ T cell antigens have been identified from a single mouse haplotype. Here we describe the generation and validation of a system to identify which surface and secreted Lm proteins are T cell antigens for any mammalian host. This information could be used in the future to inform immunotherapies based on modified Lm strains.