Paracoccidioides brasiliensis is a thermodimorphic human pathogenic fungus that causes paracoccidioidomycosis (PCM), which is the most prevalent systemic mycosis in Latin America. Differentiation from the mycelial to the yeast form (M-to-Y) is an essential step for the establishment of PCM. We evaluated the involvement of mitochondria and intracellular oxidative stress in M-to-Y differentiation. M-to-Y transition was delayed by the inhibition of mitochondrial complexes III and IV or alternative oxidase (AOX) and was blocked by the association of AOX with complex III or IV inhibitors. The expression of P. brasiliensis aox (Pbaox) was developmentally regulated through M-to-Y differentiation, wherein the highest levels were achieved in the first 24 h and during the yeast exponential growth phase; Pbaox was upregulated by oxidative stress. Pbaox was cloned, and its heterologous expression conferred cyanide-resistant respiration in Saccharomyces cerevisiae and Escherichia coli and reduced oxidative stress in S. cerevisiae cells. These results reinforce the role of PbAOX in intracellular redox balancing and demonstrate its involvement, as well as that of other components of the mitochondrial respiratory chain complexes, in the early stages of the M-to-Y differentiation of P. brasiliensis.Paracoccidioides brasiliensis, which is a thermally dimorphic fungus, is the etiological agent of paracoccidioidomycosis (PCM), which is the most prevalent human systemic mycosis in Latin America (5, 52) and affects almost 10 million individuals in Latin America (54). It is acquired by the inhalation of airborne microconidia, which reach the pulmonary alveolar epithelium and transform into the pathogenic yeast form (52,54,59). The human form of PCM that is caused by this fungus is characterized by a range of clinical manifestations, ranging from asymptomatic forms to severe, disseminated, and often fatal disease. Usually, fibrosis sequelae in the affected organs may interfere with the well-being of the patient (50).During infection, thermodimorphic fungi, such as Histoplasma capsulatum, Blastomyces dermatitidis, and P. brasiliensis, differentiate from a mycelial into a pathogenic yeast form, a transition that can also be induced (in vitro) by temperature changes from between 23 and 26°C to between 35 and 37°C. In addition, these pathogens are often subjected to significant environmental stresses, including exposure to the reactive oxygen species (ROS) and reactive nitrogen species (RNS) that are produced by the host cells (6,27,45).Elevated sublethal temperatures (thermal stress or heat shock) increase ROS generation and oxidative damage in a variety of eukaryotic cells (16,49,60,73). Mitochondria are the primary intracellular sources of ROS, which are generated in respiratory chain complexes I and III and can damage biomolecules, such as nucleic acids, lipids, and proteins (20)(21)(22). In order to control ROS levels, cells employ diverse detoxification mechanisms, including superoxide dismutase, catalase, the glutathione/thioredoxin...
