Paracoccidioidomycosis (PCM), a systemic mycosis with a high incidence in Latin America, is caused by thermodimorphic fungi of the Paracoccidioides genus. The contact with host occurs by the inhalation of conidia or mycelial propagules which once reaching the pulmonary alveoli differentiate into yeast cells. This transition process is vital in the pathogenesis of PCM allowing the fungus survival in the host. Thus, the present work performed a comparative proteome analysis of mycelia, mycelia-to-yeast transition, and yeast cells of Paracoccidioides brasiliensis . For that, tryptic peptides were labeled with iTRAQ and identified by LC–MS/MS and computational data analysis, which allowed the identification of 312 proteins differentially expressed in different morphological stages. Data showed that P. brasiliensis yeast cells preferentially employ aerobic beta-oxidation and the tricarboxylic acid cycle accompanied by oxidative phosphorylation for ATP production, in comparison to mycelia and the transition from mycelia-to-yeast cells. Furthermore, yeast cells show a metabolic reprogramming in amino acid metabolism and in the induction of virulence determinants and heat shock proteins allowing adaptation to environmental conditions during the increase of the temperature. In opposite of that, the alcoholic fermentation found to P. lutzii , at least under laboratory conditions, is strongly favored in mycelium compared to yeast cells. Thereby, the data strongly support substantial metabolic differences among members of the Paracoccidioides complex, when comparing the saprobiotic mycelia and the yeast parasitic phases.

Fungi of the complex Paracoccidioides spp. are thermodimorphic organisms that cause Paracoccidioidomycosis, one of the most prevalent mycoses in Latin America. These fungi present metabolic mechanisms that contribute to the fungal survival in host tissues. Paracoccidioides lutzii activates glycolysis and fermentation while inactivates aerobic metabolism in iron deprivation, a condition found during infection. In lungs Paracoccidioides brasiliensis face a glucose poor environment and relies on the beta-oxidation to support energy requirement. During mycelium to yeast transition P. lutzii cells up-regulate transcripts related to lipid metabolism and cell wall remodeling in order to cope with the host body temperature. Paracoccidioides spp. cells also induce transcripts/enzymes of the methylcitrate cycle (MCC), a pathway responsible for propionyl-CoA metabolism. Propionyl-CoA is a toxic compound formed during the degradation of odd-chain fatty acids, branched chain amino acids and cholesterol. Therefore, fungi require a functional MCC for full virulence and the ability to metabolize propionyl-CoA is related to the virulence traits in Paracoccidioides spp. On this way we sought to characterize the propionate metabolism in Paracoccidioides spp. The data collected showed that P. lutzii grows in propionate and activates the MCC by accumulating transcripts and proteins of methylcitrate synthase (MCS), methylcitrate dehydratase (MCD) and methylisocitrate lyase (MCL). Biochemical characterization of MCS showed that the enzyme is regulated by phosphorylation, an event not yet described. Proteomic analyses further indicate that P. lutzii yeast cells degrades lipids and amino acids to support the carbon requirement for propionate metabolism. The induction of a putative propionate kinase suggests that fungal cells use propionyl-phosphate as an intermediate in the production of toxic propionyl-CoA. Concluding, the metabolism of propionate in P. lutzii is under regulation at transcriptional and phosphorylation levels and that survival on this carbon source requires additional mechanisms other than activation of MCC.

Approximately one-third of all proteins have been estimated to contain at least one metal cofactor, and these proteins are referred to as metalloproteins. These represent one of the most diverse classes of proteins, containing metal ions that bind to specific sites to perform catalytic, regulatory and structural functions. Bioinformatic tools have been developed to predict metalloproteins encoded by an organism based only on its genome sequence. Its function and the type of metal binder can also be predicted via a bioinformatics approach. Paracoccidioides complex includes termodimorphic pathogenic fungi that are found as saprobic mycelia in the environment and as yeast, the parasitic form, in host tissues. They are the etiologic agents of Paracoccidioidomycosis, a prevalent systemic mycosis in Latin America. Many metalloproteins are important for the virulence of several pathogenic microorganisms. Accordingly, the present work aimed to predict the copper, iron and zinc proteins encoded by the genomes of three phylogenetic species of Paracoccidioides (Pb01, Pb03, and Pb18). The metalloproteins were identified using bioinformatics approaches based on structure, annotation and domains. Cu-, Fe-, and Zn-binding proteins represent 7% of the total proteins encoded by Paracoccidioides spp. genomes. Zinc proteins were the most abundant metalloproteins, representing 5.7% of the fungus proteome, whereas copper and iron proteins represent 0.3 and 1.2%, respectively. Functional classification revealed that metalloproteins are related to many cellular processes. Furthermore, it was observed that many of these metalloproteins serve as virulence factors in the biology of the fungus. Thus, it is concluded that the Cu, Fe, and Zn metalloproteomes of the Paracoccidioides spp. are of the utmost importance for the biology and virulence of these particular human pathogens.

Histoplasma capsulatum is a thermodimorphic fungus that causes histoplasmosis, a mycosis of global incidence. The disease is prevalent in temperate and tropical regions such as North America, South America, Europe, and Asia. It is known that during infection macrophages restrict Zn availability to H. capsulatum as a microbicidal mechanism. In this way the present work aimed to study the response of H. capsulatum to zinc deprivation. In silico analyses showed that H. capsulatum has eight genes related to zinc homeostasis ranging from transcription factors to CDF and ZIP family transporters. The transcriptional levels of ZAP1, ZRT1, and ZRT2 were induced under zinc-limiting conditions. The decrease in Zn availability increases fungicidal macrophage activity. Proteomics analysis during zinc deprivation at 24 and 48 h showed 265 proteins differentially expressed at 24 h and 68 at 48 h. Proteins related to energy production pathways, oxidative stress, and cell wall remodeling were regulated. The data also suggested that low metal availability increases the chitin and glycan content in fungal cell wall that results in smoother cell surface. Metal restriction also induces oxidative stress triggered, at least in part, by reduction in pyridoxin synthesis.

Esse projeto de extensão foi apresentado durante a XII Edição do Movimento Científico e Cultural de Aparecida de Goiânia, Goiás, Brasil (MOCCA). O objetivo foi promover o conhecimento científico básico sobre o fluxo da informação genética a partir da experimentação por métodos ilustrativos. Cada participante respondeu um questionário para avaliar seu conhecimento prévio sobre o tema. Em seguida, foi direcionado a um monitor que o acompanhava, no ambiente interativo, apresentando ilustrações com maquetes, painéis, modelos tridimensionais, bem como, na realização da extração simples de DNA de uma fruta. Ao final, o mesmo questionário foi reaplicado. A análise comparativa do desempenho dos participantes no questionário antes e depois da realização das atividades mostrou que as abordagens interativas e as práticas ilustrativas contribuíram, positivamente, para a melhor compreensão do conteúdo sugerindo que essa metodologia possa ser incorporada ao processo de ensino-aprendizagem.