Fungal pathogens cause life-threatening diseases in humans, and the increasing prevalence of these diseases emphasizes the need for new targets for therapeutic intervention. Nutrient acquisition during
infection is a promising target, and recent studies highlight the contributions of endomembrane trafficking, mitochondria, and vacuoles in the sensing and acquisition of
heme by fungi. These studies have been facilitated by genetically encoded biosensors and other tools to quantitate
heme in subcellular compartments and to investigate the dynamics of trafficking in living cells. In particular, the applications of biosensors in fungi have been extended beyond the detection of metabolites, cofactors, pH, and redox status to include the detection of
heme. Here, we focus on studies that make use of biosensors to examine mechanisms of
heme uptake and degradation, with guidance from the model fungus Saccharomyces cerevisiae and an emphasis on the pathogenic fungi Candida albicans and Cryptococcus neoformans that threaten human health. These studies emphasize a role for endocytosis in
heme uptake, and highlight membrane contact sites involving mitochondria, the endoplasmic reticulum and vacuoles as mediators of intracellular
iron and
heme trafficking.