Proteomic analysis of Paracoccidioides brasiliensis complex isolates: Correlation of the levels of differentially expressed proteins with in vivo virulence

Abstract

Background Paracoccidioidomycosis (PCM) is a systemic mycosis commonly found in Latin America that is caused by distinct species of Paracoccidioides genus: Paracoccidioides brasiliensis complex (S1, PS2, PS3 and PS4) and Paracoccidioides lutzii . Its pathobiology has been recently explored by different approaches to clarify the mechanisms of host-pathogen interactions underpinning PCM. The diversity of clinical forms of this disease has been attributed to both host- and fungus-related factors. Methodology/Principal findings For better understanding of the molecular underpinnings of host-fungus interactions, we evaluated in vivo virulence of nine Paracoccidioides brasiliensis complex isolates and correlated it to protein expression profiles obtained by two-dimensional gel electrophoresis. Based on the recovery of viable fungi from mouse organs, the isolates were classified as those having low, moderate, or high virulence. Highly virulent isolates overexpressed proteins related to adhesion process and stress response, probably indicating important roles of those fungal proteins in regulating the colonization capacity, survival, and ability to escape host immune system reaction. Moreover, highly virulent isolates exhibited enhanced expression of glycolytic pathway enzymes concomitantly with repressed expression of succinyl-CoA ligase beta chain, a protein related to the tricarboxylic acid cycle. Conclusions/Significance Our findings may point to the mechanisms used by highly virulent P . brasiliensis isolates to withstand host immune reactions and to adapt to transient iron availability as strategies to survive and overcome stress conditions inside the host.

Immunoproteomic Analysis Reveals Novel Candidate Antigens for the Diagnosis of Paracoccidioidomycosis Due to Paracoccidioides lutzii

“…The 16 specific immunogenic proteins of P. lutzii can be divided into three main groups. Group 1 comprises seven proteins that have been widely reported in Paracoccidioides under experimental physiological conditions according to STRING analysis, namely, triosephosphate isomerase, glyceraldehyde-3-phosphate dehydrogenase, citrate synthase, isocitrate lyase, enolase, HSP60-like protein, and an uncharacterized protein (thioredoxin-like superfamily) [ 54 , 70 , 87 , 88 , 89 , 90 , 91 , 92 , 93 ]. Proteins included in group 2 are antigens that already have been described in human PCM, such as triosephosphate isomerase, glyceraldehyde-3-phosphate dehydrogenase, fructose-bisphosphate aldolase [ 70 ], or Paracoccidioides antigens that already have been related to eliciting a humoral immune response in BALB/c mice, such as transketolase, HSP60-like protein, and isocitrate lyase (all shared with PS3) [ 71 ].…”

“…The 25 specific immunogenic proteins of P. brasiliensis can be divided into three main groups. Group 1 comprises 13 proteins that have already been described in Paracoccidioides under experimental physiological condition, namely, triosephosphate isomerase, spermidine synthase, glyceraldehyde-3-phosphate dehydrogenase, 4-hydroxyphenylpyruvate dioxygenase, thioredoxin reductase, ATP synthase subunit beta, hexokinase, HSP75-like protein, HSP60-like protein, HSP7-like protein, aconitate hydratase, mitochondrial, 1-Cys peroxiredoxin, and an uncharacterized protein (XP_010758132) [ 54 , 70 , 87 , 88 , 89 , 90 , 91 , 92 , 93 , 94 , 95 ], all of them enriched in our STRING network. Proteins in group 2 correspond to antigens that have already been described in human PCM [ 70 ] or immunogens that have already been described to elicit a humoral immune response in BALB/c mice, such as HSP7-like protein and HSP 60-like protein [ 71 ].…”

“…Therefore, to avoid any bias among Paracoccidioides spp. isolates at the start of in vitro culturing, isolates were passed through BALB/c and then reisolated before protein extraction, as previously described by our group [ 54 ].…”

An Overview of Genomics, Phylogenomics and Proteomics Approaches in Ascomycota

“…Pathways described as essential for the survival of pathogens inside macrophages (i.e., phosphate pentoses pathway, synthesis of cell wall components and mitochondrial activity) were in fact observed only or with higher intensity in yeast cells recovered from non-primed macrophages. For a better understanding of the molecular underpinnings of host-fungus interactions, Do Amaral et al [ 256 ], carried out a quantitative proteomic study (2D-E, LC-MS/MS) in an attempt to correlate expression profiles to in vivo virulence of a number of P. brasiliensis strains isolated from environmental and animal sources, including Pb18. Highly virulent isolates that caused disseminated disease in a murine model of PCM were found to overexpress proteins related to pathogenicity, metabolism and redox homeostasis (e.g., glutathione reductase, peroxisomal catalase, RNA binding protein), indicated as critical in regulating the fugus ability to escape the host immune system.…”

“… [ 255 ] 2D-E, LC-MS/MS 11 Identification of proteins commonly overexpressed in highly virulent P. brasiliensis spp. complex isolates causing disseminated disease in a murine model of PCM [ 256 ] Nano-ESI-UPLC-MS E 308 Characterization of the proteomic response to macrophage internalization. [ 410 ] Secretome LC-MS/MS 205; 260 Vesicle and vesicle-free extracellular proteome.…”