Caracterização do fator de transcrição AreA de P. lutzii

Abstract

Paracoccidioidomycosis is a mycosis endemic to subtropical areas of Latin America, caused by fungi of the genus Paracoccidioides. Infection occurs via inhalation of conidia or fragments of mycelium. In the lungs of the host, phagocytic cells engage in the process of phagocytosis to combat the microorganism. Within the phagosome, alterations in pH, oxidative stress, and nutritional limitation present challenges to the pathogen's survival. In this context, nutritional immunity can be defined as a process that deprives the pathogen of preferred sources of essential nutrients required for the synthesis of biomolecules, such as nitrogen sources. Pathogenic fungi such as Paracoccidioides spp. have a coordinated mechanism for capturing non-preferred sources of nitrogen, which is known as Nitrogen Catabolic Repression (NCR). The transcription factor GATA AreA is responsible for regulating this process, ensuring the catabolism of nitrogen available in the environment by controlling the genes responsible for the uptake and assimilation of these sources. The objective of this study was to examine the influence of areA gene silencing on the assimilation of nitrogen sources, growth, cell wall composition, and cell size of P. lutzii, with comparisons between mutant and wild-type strains grown in varying concentrations of glutamine. The gap and areA genes were evaluated by RT-qPCR. The growth curve was measured by absorbance at 0, 12, 24, and 48 hours. The viability of the cells was estimated by propidium iodide, followed by a subsequent analysis under a fluorescence microscope. To evaluate the composition of the cell wall and cell size, the cells were stained with calcofluor white and examined under a fluorescence microscope. The gap and areA genes exhibited elevated expression levels in the glutamine mutant strain. The CFW analysis indicates a discrepancy in cell wall composition between the strains in the presence of 10 mM glutamine. Furthermore, the mutant exhibited increased cell size under these conditions. Consequently, the results indicate that the strain in which the areA gene has been silenced displays a distinct biological response when grown in 10 mM glutamine, which differs from that described in the literature. An analysis of the expression of new gene targets and an evaluation of the proteomic profile in the therefore mentioned conditions may assist in identifying crucial factors for understanding the role of AreA in P. lutzii.