The environmental fungus Cryptococcus neoformans is capable of adapting to the human host, and in susceptible individuals, can cause deadly meningitis that is fatal without antifungal therapy. C. neoformans is responsible for ~15% of AIDS-related deaths worldwide, and is a significant cause of mortality and allograft loss in the transplant population. Interactions with macrophages are important to the outcome of infection. In response to the macrophage intracellular environment, C. neoformans expresses factors that promote survival, including the enzyme urease that prevents acidification of the phago-lysosome, and reactive oxygen stress response factors that detoxify reactive oxygen species and repair oxidative damage. We present compelling preliminary data to suggest that both urease and ROS response effectors are regulated at the level of mRNA translation through the kinase Gcn2. Gcn2 is the sole kinase in C. neoformans that phosphorylates eukaryotic translation initiation factor 2 (eIF2), thereby regulating translation initiation. In this collaboration, the expertise of the Panepinto lab in translational regulation in C. neoformans synergizes with the innovative computational and genomics approaches developed by the McManus lab to investigate the translational response to oxidative stress and perform the first study of translational regulation in C. neoformans during macrophage infection. The proposal consists of two aims. The first aim will use ribosome profiling and RNA-seq to investigate the genome wide translational regulation in response to oxidative stress and determine the consequences of GCN2 deletion on ROS response gene expression and susceptibility to oxidative killing by macrophages. The second Aim will examine the translational regulation employed by C. neoformans during intracellular growth in bone marrow derived macrophages, and its dependence on Gcn2. These studies will provide a comprehensive view of how C. neoformans reprograms its translatome in response to oxidative stress, and during macrophage infection. Due to the fundamental importance of translational control, the processes selected for translation during macrophage adaptation are expected to include potential targets for future therapeutic development.