Granulysin and the antimicrobial activity of CD8T cells - development of a better model

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Abstract. CD8 T cells are an important arm of adaptive immunity, and make critical contributions to protection against viruses, other intracellular pathogens, and tumors. Our lab has a long-standing interest in the role of CD8 T cells in immunity to tuberculosis (TB), which is the leading cause of death in the world from an infectious agent. Recent data from the non-human primate (NHP) model of TB shows that CD8 T cells make a crucial contribution to both primary and vaccine-mediated immunity against Mycobacterium tuberculosis (Mtb) [1-5]. Increases in CD8 T cell number and function are associated with vaccine-induced protection in the NHP TB model. Furthermore, using single cell RNASeq, cytotoxic NK and CD8 T cells are significantly associated with host-beneficial outcome (i.e., Mtb control). Importantly, CTL frequently associated with restrictive granulomas more frequently express the combination of perforin, granzymes, and granulysin. Granulysin (GNLY) is an important cytotoxic granule component produced by cytotoxic CD8 T cells and NK cells, and can kill many different types of intracellular bacterial pathogens. The granulysin protein has antimicrobial activity against Mtb when directly applied to bacteria and can kill drug resistant Mtb strains [6]. However, it has been difficult to study the role of granulysin in the mouse model, as rodents lack an ortholog of the gene. A human granulysin transgenic mouse (hGNLY-tg) was developed by Allan Krensky [7]. In that mouse, granulysin is expressed primarily in NK cells, and in CD8 T cells only after vigorous in vitro stimulation. Here, we propose to develop new mouse models to study the role of granulysin in antimicrobial immunity. Our approach will address whether granulysin expression is crucial for CD8 T cell control of Mtb. Our expression system will enable better modelling of human CD8 T cell responses in the mouse. This model will provide an experimental system to support investigation of the role of granulysin in immunity to TB. We hypothesize that the suboptimal antibacterial function of CD8 T cells in the murine model is their lack of granulysin expression. A corollary is that granulysin expression by murine CD8 T cells would improve their ability to control Mtb infection after low dose aerosol Mtb infection. We will answer this question as part of this project.