Depicting the metabolism of Paracoccidioides brasiliensis during infection by transcriptional and proteomic approaches

Species in the Paracoccidioides spp. complex are the causative agents of paracoccidioidomycosis (PCM), considered one of the most important systemic endemic mycosis in Latin America. The genus Paracoccidioides comprises 2 species, P. brasiliensis and P. lutzii with the former being classified into 4 different phylogenetic groups named S1, PS2, PS3 and PS4. More recently, the S1 group was sub-divided into 2 lineages (S1a and S1b). The development of the disease depends on several factors dependent on both the host and the fungal virulence. Once the fungal pathogen enters in contact with the host, it should adapt itself to and override the various microenvironmental factors to survive inside the host. Fungal adaptation depends on its metabolism and capability to produce and secrete several molecules that play an important role in virulence. Among these molecules, Paracoccidioides species produce enzymes that participate in the most important pathways of central metabolism, involving carbohydrates, lipids, amino acids and nucleotides. In a previous study and using proteomic analysis, it was found that after internalization of P. brasiliensis into macrophages, a total of 308 differentially expressed proteins were detected in this fungal pathogen. The upregulated proteins included those related to b-oxidation of fatty acids and amino acids’ catabolism, as well as proteins associated with the alternative carbon metabolism and enzymes involved in the gluconeogenesis pathway. As it concerns the downregulated proteins included those related to glycolysis and protein synthesis. Interestingly, it has been described that most of the Paracoccidioides spp enzymes that participate in both the glycolytic pathway and the tricarboxylic acid (TCA) cycle including glyceraldehyde-3-phosphate dehydrogenase (GADPH), enolase (ENO), fructose-1–6-bisphosphate aldolase (FBA), triose phosphate isomerase (TPI), malate synthase (MLS) and isocitrate lyase (ICL), function as moonlight proteins, this is, they are multifunctional proteins capable of accomplishing different functions often unrelated. Thus, these glycolytic enzymes allow Paracoccidioides spp binding to extracellular matrix (ECM) proteins such as laminin, fibronectin, fibrinogen, type I and IV collagens and plasminogen, among others. These interactions are involved in the adhesion and invasion process leading to Paracoccidioides infection. Paracoccidioides spp also has the capacity to defend itself against the oxidative and nitrosative stress molecules produced by the host s immune cells especially those produced by phagocytic cells (mainly macrophages). The oxidative and nitrosative stress are characterized by production of reactive oxygen species (ROS) and reactive nitrogen species (RNS), respectively. ROS comprises superoxide anion radical (O2), hydroxyl radical ( OH) and hydrogen peroxide (H2O2) while RNS comprises nitric oxide (NO ) and peroxynitrite (ONOO-), among others. These oxidative and nitrosative molecules act on fungal cells through the inactivation of proteins, lipids and cellular enzymes inhibiting respiration, and damaging the DNA and membranes thus leading to the fungal death. Therefore, several reports have been shown that Paracoccidioides spp has a powerful antioxidant defense system that allows this fungal pathogen to survive ROS and RNS production; this system includes expression/production of several enzymes such as catalases (CATs), superoxide dismutases (SODs), alternative oxidase (AOX), cytochrome c peroxidase (CCP) and thioredoxins (THX). The important role in virulence of most of these detoxifying molecules has been demonstrated in elegant studies using knock-down or silenced P. brasiliensis strains obtained by antisense RNA (aRNA) approaches both in