Developing a transgenic mouse model to better understand the role of CD8 and granulysin against Mycobacterium tuberculosis infection.

Abstract

Abstract Human cytotoxic lymphocytes kill infected cells through release of lytic proteins contained in cytotoxic granules. The cytotoxic granules of human NK and CD8 T cells contain perforin, granzyme and granulysin (GNLY), and GNLY has direct antimicrobial activity against Mycobacterium tuberculosis (Mtb). Murine studies suggest that CD8 T cells have only a modest role in immunity to tuberculosis (TB). We hypothesize that mouse CD8 T cells are ineffective at containing Mycobacterium tuberculosis (Mtb) as they don’t express GNLY. To test this hypothesis, GNLY-transgenic mice that express human granulysin protein under the control of human regulatory elements were infected with Mtb. GNLY-Tg and non-tg control mice control Mtb similarly. There was no difference in the ability of GNY-Tg and non-tg CD8 T cells to transfer protection to T cell deficient mice. Anti-CD3 stimulation led to expression of granulysin in human CD8 T cells, but not in murine CD8 T cells. However, after Mtb infection, GNLY was expressed in NK cells but not CD8 T cells. We conclude that GNLY-expressing NK cells do not alter resistance to Mtb. Importantly, GNLY-Tg mice are inadequate model to study CD8 function during TB. We are developing a new strategy to express granulysin in CD8 T cells. We expressed GNLY (isoform 2) in primary murine CD8 T cells by retroviral gene transfer and detected GNLY expression in 60% of CD8 T cells. Future directions We are assessing the cytolytic activity of GNLY-expressing CD8 T cells and their ability to kill intracellular pathogens in vitro and in vivo. We expect to develop a new GNLY-Tg model that can be used to study the function of CD8 T cells and granulysin in many different diseases. This work is supported by NIH AI159374. This work is supported by NIH grant R21 AI159374.