Updated project metadata. Fungal pathogens are emerging threats to global health and the rise of incidence is associated with climate change and increased geographical distribution; factors also influencing host susceptibility to infection. Accurate detection and diagnosis of fungal infections is paramount to align options for rapid and effective therapeutic treatments. For improved diagnostics, the discovery and development of biomarkers presents a promising avenue; however, this approach requires a priori knowledge of markers of infection. To uncover putative novel biomarkers of disease, profiling of the host immune response and pathogen virulence factor production is indispensable. In this study, we use mass spectrometry-based proteomics to resolve the temporal and spatial proteomes of Cryptococcus neoformans infection of the spleen following a murine model of infection. Dual perspective proteome profiling defines global remodeling of the host over a time course of infection, confirming activation of immune associated proteins connected to fungal infections. Conversely, pathogen proteomes detect well-characterized C. neoformans virulence determinants, along with novel mapped patterns of pathogenesis during progression of disease. Together, our systematic approach confirms immune protection against fungal pathogens and explores the discovery of putative biomarkers from complementary biological systems for monitoring the presence and progression of cryptococcal disease.