Abstract
ABSTRACTParacoccidoides brasiliensis and Paracoccidioides lutzii, the etiologic agents of paracoccidioidomycosis, cause disease in healthy and immunocompromised persons in Latin America. We developed a method for harvesting P. brasiliensis yeast cells from infected murine lung to facilitate in vivo transcriptional and proteomic profiling. P. brasiliensis harvested at 6 h post-infection were analyzed using RNAseq and LC-MSE. In vivo yeast cells had 594 differentially expressed transcripts and 350 differentially expressed proteins. Integration of transcriptional and proteomic data indicated that early in infection (6 h), P. brasiliensis yeast cells underwent a shift in metabolism from glycolysis to β-oxidation, upregulated detoxifying enzymes to defend against oxidative stress, and repressed cell wall biosynthesis. Bioinformatics and functional analyses also demonstrated that a serine proteinase was upregulated and secreted in vivo. To our knowledge this is the first study depicting transcriptional and proteomic data of P. brasiliensis yeast cells upon 6 h post-infection of mouse lung.
Highlights
Paracoccidioides brasiliensis and Paracoccidioides lutzii, the etiological agents of paracoccidioidomycosis (PCM), are fungal pathogens that are acquired following the inhalation of mycelia fragments or conidia into the lungs of a human host.[1]
Prior studies have profiled gene expression in P. lutzii yeast cells incubated in human blood at 36C, using cDNA representational difference analysis, as well as P. brasiliensis yeast cells internalized by macrophages, using microarray technology.[4,5]
For transcriptome and proteome analyses, we focused our attention at 6 hours post-inhalation because our data demonstrated that at this time point, P. brasiliensis invaded mouse lung tissue
Summary
Paracoccidioides brasiliensis and Paracoccidioides lutzii, the etiological agents of paracoccidioidomycosis (PCM), are fungal pathogens that are acquired following the inhalation of mycelia fragments or conidia into the lungs of a human host.[1] A defining feature of Paracoccidioides pathogenesis is the morphologic transition from mycelia or conidia to yeast cells, which is governed by temperature and occurs when mycelia and conidia enter the lungs. Our goal is to characterize genes involved with the adaptive response of P. brasiliensis during the early stages of pulmonary infection. Identification of these genes is crucial for the understanding of fungal pathogenesis, the limited abundance of P. brasiliensis yeast cells at the sites of infection, makes gene and protein assays challenging. Approaches used to identify genes relevant for infection at the proteomic level, include the use of various in vitro experimental conditions such as iron deprivation, hypoxia, oxidative, nitrosative stress, and fungal-macrophage interactions.[6,7,8,9,10] The later approach demonstrated that during macrophage infection, yeast cells remodel their
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