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Célia Maria De Almeida Soares - One of the best experts on this subject based on the ideXlab platform.
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beyond melanin proteomics reveals virulence related proteins in Paracoccidioides brasiliensis and Paracoccidioides lutzii yeast cells grown in the presence of l dihydroxyphenylalanine
Journal of Fungi, 2020Co-Authors: Rodrigo Almeidapaes, Célia Maria De Almeida Soares, Marcos Abreu Almeida, Lilian Cristiane Baeza, Leticia Andrade Mendes Marmello, Monique R O Trugilho, Joshua D Nosanchuk, Richard H Valente, Rosely Maria ZancopeoliveiraAbstract:Species of the genus Paracoccidioides cause a systemic infection in human patients. Yeast cells of Paracoccidioides spp. produce melanin in the presence of L-dihydroxyphenylalanine and during infection, which may impact the pathogen’s survival in the host. To better understand the metabolic changes that occur in melanized Paracoccidioides spp. cells, a proteomic approach was performed to compare melanized and non-melanized Paracoccidioides brasiliensis and Paracoccidioides lutzii yeast cells. Melanization was induced using L-dihydroxyphenylalanine as a precursor, and quantitative proteomics were performed using reversed-phase nano-chromatography coupled to high-resolution mass spectrometry. When comparing melanized versus non-melanized cells, 1006 and 582 differentially abundant/detected proteins were identified for P. brasiliensis and P. lutzii, respectively. Functional enrichment and comparative analysis revealed 30 important KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways in melanized P. brasiliensis and 18 in P. lutzii, while differentially abundant proteins from non-melanized cells from these species were involved in 21 and 25 enriched pathways, respectively. Melanized cells presented an abundance of additional virulence-associated proteins, such as phospholipase, proteases, superoxide dis-mutases, heat-shock proteins, adhesins, and proteins related to vesicular transport. The results suggest that L-dihydroxyphenylalanine increases the virulence of Paracoccidioides spp. through complex mechanisms involving not only melanin but other virulence factors as well.
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Cell Wall Synthesis, Development of Hyphae and Metabolic Pathways Are Processes Potentially Regulated by MicroRNAs Produced Between the Morphological Stages of Paracoccidioides brasiliensis.
Frontiers in microbiology, 2018Co-Authors: Juliana S. De Curcio, Juliano D. Paccez, Evandro Novaes, Mathias Brock, Célia Maria De Almeida SoaresAbstract:MicroRNAs are molecules involved in post-transcriptional gene regulation. In pathogenic fungi, microRNAs have been described at different morphological stages by regulating targets involved in processes such as morphogenesis and energy production. Members of the Paracoccidioides complex are the main etiological agents of a systemic mycosis in Latin America. Fungi of the Paracoccidioides complex present a wide range of plasticity to colonize different niches. In response to environmental changes these fungi undergo a morphological switch, remodel their cellular metabolism and modulate structural cell wall components. However, the underlying mechanisms regulating the gene expression is not well understood. By using high performance sequencing and bioinformatics analyses, this work characterizes microRNAs produced by Paracoccidioides brasiliensis. Here, we demonstrated that the transcript encoding proteins involved in microRNA biogenesis were differentially expressed in each morphological stage. In addition, 49 microRNAs were identified in cDNA libraries with 44 differentially regulated among the libraries. Sixteen microRNAs were differentially regulated in comparison to the mycelium in the mycelium-to-yeast transition phase. The yeast parasitic phase revealed a complete remodeling of the expression of these small RNAs. Analyses of targets of the induced microRNAs, from the different libraries, revealed that these molecules may potentially regulate in the cell wall, by repressing genes involved in the synthesis and degradation of glucans and chitin. Furthermore, mRNAs involved in cellular metabolism and development were predicted to be regulated by microRNAs. Therefore, this work describes a putative post transcriptional regulation, mediated by microRNAs in P. brasiliensis and its influence on the adaptive processes of thermal dimorphic fungus.
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Image_10_Cell Wall Synthesis, Development of Hyphae and Metabolic Pathways Are Processes Potentially Regulated by MicroRNAs Produced Between the Morphological Stages of Paracoccidioides brasiliensis.TIF
2018Co-Authors: Juliana S. De Curcio, Juliano D. Paccez, Evandro Novaes, Mathias Brock, Célia Maria De Almeida SoaresAbstract:MicroRNAs are molecules involved in post-transcriptional gene regulation. In pathogenic fungi, microRNAs have been described at different morphological stages by regulating targets involved in processes such as morphogenesis and energy production. Members of the Paracoccidioides complex are the main etiological agents of a systemic mycosis in Latin America. Fungi of the Paracoccidioides complex present a wide range of plasticity to colonize different niches. In response to environmental changes these fungi undergo a morphological switch, remodel their cellular metabolism and modulate structural cell wall components. However, the underlying mechanisms regulating the gene expression is not well understood. By using high performance sequencing and bioinformatics analyses, this work characterizes microRNAs produced by Paracoccidioides brasiliensis. Here, we demonstrated that the transcript encoding proteins involved in microRNA biogenesis were differentially expressed in each morphological stage. In addition, 49 microRNAs were identified in cDNA libraries with 44 differentially regulated among the libraries. Sixteen microRNAs were differentially regulated in comparison to the mycelium in the mycelium-to-yeast transition phase. The yeast parasitic phase revealed a complete remodeling of the expression of these small RNAs. Analyses of targets of the induced microRNAs, from the different libraries, revealed that these molecules may potentially regulate in the cell wall, by repressing genes involved in the synthesis and degradation of glucans and chitin. Furthermore, mRNAs involved in cellular metabolism and development were predicted to be regulated by microRNAs. Therefore, this work describes a putative post transcriptional regulation, mediated by microRNAs in P. brasiliensis and its influence on the adaptive processes of thermal dimorphic fungus.
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Image_2_Cell Wall Synthesis, Development of Hyphae and Metabolic Pathways Are Processes Potentially Regulated by MicroRNAs Produced Between the Morphological Stages of Paracoccidioides brasiliensis.TIF
2018Co-Authors: Juliana S. De Curcio, Juliano D. Paccez, Evandro Novaes, Mathias Brock, Célia Maria De Almeida SoaresAbstract:MicroRNAs are molecules involved in post-transcriptional gene regulation. In pathogenic fungi, microRNAs have been described at different morphological stages by regulating targets involved in processes such as morphogenesis and energy production. Members of the Paracoccidioides complex are the main etiological agents of a systemic mycosis in Latin America. Fungi of the Paracoccidioides complex present a wide range of plasticity to colonize different niches. In response to environmental changes these fungi undergo a morphological switch, remodel their cellular metabolism and modulate structural cell wall components. However, the underlying mechanisms regulating the gene expression is not well understood. By using high performance sequencing and bioinformatics analyses, this work characterizes microRNAs produced by Paracoccidioides brasiliensis. Here, we demonstrated that the transcript encoding proteins involved in microRNA biogenesis were differentially expressed in each morphological stage. In addition, 49 microRNAs were identified in cDNA libraries with 44 differentially regulated among the libraries. Sixteen microRNAs were differentially regulated in comparison to the mycelium in the mycelium-to-yeast transition phase. The yeast parasitic phase revealed a complete remodeling of the expression of these small RNAs. Analyses of targets of the induced microRNAs, from the different libraries, revealed that these molecules may potentially regulate in the cell wall, by repressing genes involved in the synthesis and degradation of glucans and chitin. Furthermore, mRNAs involved in cellular metabolism and development were predicted to be regulated by microRNAs. Therefore, this work describes a putative post transcriptional regulation, mediated by microRNAs in P. brasiliensis and its influence on the adaptive processes of thermal dimorphic fungus.
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Table_2_Cell Wall Synthesis, Development of Hyphae and Metabolic Pathways Are Processes Potentially Regulated by MicroRNAs Produced Between the Morphological Stages of Paracoccidioides brasiliensis.docx
2018Co-Authors: Juliana S. De Curcio, Juliano D. Paccez, Evandro Novaes, Mathias Brock, Célia Maria De Almeida SoaresAbstract:MicroRNAs are molecules involved in post-transcriptional gene regulation. In pathogenic fungi, microRNAs have been described at different morphological stages by regulating targets involved in processes such as morphogenesis and energy production. Members of the Paracoccidioides complex are the main etiological agents of a systemic mycosis in Latin America. Fungi of the Paracoccidioides complex present a wide range of plasticity to colonize different niches. In response to environmental changes these fungi undergo a morphological switch, remodel their cellular metabolism and modulate structural cell wall components. However, the underlying mechanisms regulating the gene expression is not well understood. By using high performance sequencing and bioinformatics analyses, this work characterizes microRNAs produced by Paracoccidioides brasiliensis. Here, we demonstrated that the transcript encoding proteins involved in microRNA biogenesis were differentially expressed in each morphological stage. In addition, 49 microRNAs were identified in cDNA libraries with 44 differentially regulated among the libraries. Sixteen microRNAs were differentially regulated in comparison to the mycelium in the mycelium-to-yeast transition phase. The yeast parasitic phase revealed a complete remodeling of the expression of these small RNAs. Analyses of targets of the induced microRNAs, from the different libraries, revealed that these molecules may potentially regulate in the cell wall, by repressing genes involved in the synthesis and degradation of glucans and chitin. Furthermore, mRNAs involved in cellular metabolism and development were predicted to be regulated by microRNAs. Therefore, this work describes a putative post transcriptional regulation, mediated by microRNAs in P. brasiliensis and its influence on the adaptive processes of thermal dimorphic fungus.
Alexandre Melo Bailão - One of the best experts on this subject based on the ideXlab platform.
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employing proteomic analysis to compare Paracoccidioides lutzii yeast and mycelium cell wall proteins
Biochimica et Biophysica Acta, 2017Co-Authors: Danielle Silva Araujo, Clayton Luiz Borges, Alexandre Melo Bailão, Lilian Cristiane Baeza, Patrícia De Sousa Lima, Ana Flavia Alves Parente, Célia Maria De Almeida SoaresAbstract:Paracoccidioidomycosis is an important systemic mycosis caused by thermodimorphic fungi of the Paracoccidioides genus. During the infective process, the cell wall acts at the interface between the fungus and the host. In this way, the cell wall has a key role in growth, environment sensing and interaction, as well as morphogenesis of the fungus. Since the cell wall is absent in mammals, it may present molecules that are described as target sites for new antifungal drugs. Despite its importance, up to now few studies have been conducted employing proteomics in for the identification of cell wall proteins in Paracoccidioides spp. Here, a detailed proteomic approach, including cell wall-fractionation coupled to NanoUPLC-MSE, was used to study and compare the cell wall fractions from Paracoccidioides lutzii mycelia and yeast cells. The analyzed samples consisted of cell wall proteins extracted by hot SDS followed by extraction by mild alkali. In summary, 512 proteins constituting different cell wall fractions were identified, including 7 predicted GPI-dependent cell wall proteins that are potentially involved in cell wall metabolism. Adhesins previously described in Paracoccidioides spp. such as enolase, glyceraldehyde-3-phosphate dehydrogenase were identified. Comparing the proteins in mycelium and yeast cells, we detected some that are common to both fungal phases, such as Ecm33, and some specific proteins, as glucanase Crf1. All of those proteins were described in the metabolism of cell wall. Our study provides an important elucidation of cell wall composition of fractions in Paracoccidioides, opening a way to understand the fungus cell wall architecture.
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Paracoccidioides brasiliensis presents metabolic reprogramming and secretes a serine proteinase during murine infection
Virulence, 2017Co-Authors: Laurine Lacerda Pigosso, Clayton Luiz Borges, Alexandre Melo Bailão, Lilian Cristiane Baeza, Mariana Vieira Tomazett, Mariana Batista Rodrigues Faleiro, Veridiana Maria Brianezi Dignani De Moura, Juliana Alves Parente Rocha, Gabriel Fernandes, Gregory M GauthierAbstract:Paracoccidoides 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.
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transcriptome profile of the response of Paracoccidioides spp to a camphene thiosemicarbazide derivative
PLOS ONE, 2015Co-Authors: Livia Do Carmo Silva, Alexandre Melo Bailão, Cecília M. A. De Oliveira, Diana Patricia Tamayo Ossa, Symone Vitoriano Da Conceicao Castro, Ludmila Bringel Pires, Cleuza C Da Silva, Narcimario P Coelho, Juliana Alves Parenterocha, Célia Maria De Almeida SoaresAbstract:Paracoccidioidomycosis (PCM) is a systemic granulomatous human mycosis caused by fungi of the genus Paracoccidioides, which is geographically restricted to Latin America. Inhalation of spores, the infectious particles of the fungus, is a common route of infection. The PCM treatment of choice is azoles such as itraconazole, but sulfonamides and amphotericin B are used in some cases despite their toxicity to mammalian cells. The current availability of treatments highlights the need to identify and characterize novel targets for antifungal treatment of PCM as well as the need to search for new antifungal compounds obtained from natural sources or by chemical synthesis. To this end, we evaluated the antifungal activity of a camphene thiosemicarbazide derivative (TSC-C) compound on Paracoccidioides yeast. To determine the response of Paracoccidioides spp. to TSC-C, we analyzed the transcriptional profile of the fungus after 8 h of contact with the compound. The results demonstrate that Paracoccidioides lutzii induced the expression of genes related to metabolism; cell cycle and DNA processing; biogenesis of cellular components; cell transduction/signal; cell rescue, defense and virulence; cellular transport, transport facilities and transport routes; energy; protein synthesis; protein fate; transcription; and other proteins without classification. Additionally, we observed intensely inhibited genes related to protein synthesis. Analysis by fluorescence microscopy and flow cytometry revealed that the compound induced the production of reactive oxygen species. Using an isolate with down-regulated SOD1 gene expression (SOD1-aRNA), we sought to determine the function of this gene in the defense of Paracoccidioides yeast cells against the compound. Mutant cells were more susceptible to TSC-C, demonstrating the importance of this gene in response to the compound. The results presented herein suggest that TSC-C is a promising candidate for PCM treatment.
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analysis of Paracoccidioides secreted proteins reveals fructose 1 6 bisphosphate aldolase as a plasminogen binding protein
BMC Microbiology, 2015Co-Authors: Edilânia Gomes Araujo Chaves, Clayton Luiz Borges, Alexandre Melo Bailão, Luiz Augusto Pereira, Sônia Nair Báo, Simone Schneider Weber, Célia Maria De Almeida SoaresAbstract:Background Despite being important thermal dimorphic fungi causing Paracoccidioidomycosis, the pathogenic mechanisms that underlie the genus Paracoccidioides remain largely unknown. Microbial pathogens express molecules that can interact with human plasminogen, a protein from blood plasma, which presents fibrinolytic activity when activated into plasmin. Additionally, plasmin exhibits the ability of degrading extracellular matrix components, favoring the pathogen spread to deeper tissues. Previous work from our group demonstrated that Paracoccidioides presents enolase, as a protein able to bind and activate plasminogen, increasing the fibrinolytic activity of the pathogen, and the potential for adhesion and invasion of the fungus to host cells. By using proteomic analysis, we aimed to identify other proteins of Paracoccidioides with the ability of binding to plasminogen.
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Characterization of the Paracoccidioides Hypoxia Response Reveals New Insights into Pathogenesis Mechanisms of This Important Human Pathogenic Fungus
2015Co-Authors: Patrícia De Sousa Lima, Alexandre Melo Bailão, Dawoon Chung, Robert A. Cramer, Célia Maria De Almeida SoaresAbstract:BackgroundHypoxic microenvironments are generated during fungal infection. It has been described that to survive in the human host, fungi must also tolerate and overcome in vivo microenvironmental stress conditions including low oxygen tension; however nothing is known how Paracoccidioides species respond to hypoxia. The genus Paracoccidioides comprises human thermal dimorphic fungi and are causative agents of paracoccidioidomycosis (PCM), an important mycosis in Latin America.Methodology/Principal FindingsIn this work, a detailed hypoxia characterization was performed in Paracoccidioides. Using NanoUPLC-MSE proteomic approach, we obtained a total of 288 proteins differentially regulated in 12 and 24 h of hypoxia, providing a global view of metabolic changes during this stress. In addition, a functional characterization of the homologue to the most important molecule involved in hypoxia responses in other fungi, the SREBP (sterol regulatory element binding protein) was performed. We observed that Paracoccidioides species have a functional homologue of SREBP, named here as SrbA, detected by using a heterologous genetic approach in the srbA null mutant in Aspergillus fumigatus. Paracoccidioides srbA (PbsrbA), in addition to involvement in hypoxia, is probable involved in iron adaptation and azole drug resistance responses.Conclusions/SignificanceIn this study, the hypoxia was characterized in Paracoccidioides. The first results can be important for a better understanding of the fungal adaptation to the host and improve the arsenal of molecules for the development of alternative treatment options in future, since molecules related to fungal adaptation to low oxygen levels are important to virulence and pathogenesis in human pathogenic fungi.
Maristela Pereira - One of the best experts on this subject based on the ideXlab platform.
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identification of a new antifungal compound against isocitrate lyase of Paracoccidioides brasiliensis
Future Microbiology, 2019Co-Authors: Maristela Pereira, Luciane S Da Silva, Uessiley Ribeiro Barbosa, Livia Do Carmo Silva, Celia Ma Soares, Roosevelt Alves Da SilvaAbstract:Aim: To perform virtual screening of compounds based on natural products targeting isocitrate lyase of Paracoccidioides brasiliensis. Materials & methods: Homology modeling and molecular dynamics simulations were applied in order to obtain conformational models for virtual screening. The selected hits were tested in vitro against enzymatic activity of ICL of the dimorphic fungus P. brasiliensis and growth of the Paracoccidioides spp. The cytotoxicity and selectivity index of the compounds were defined. Results & conclusion: Carboxamide, lactone and β-carboline moieties were identified as interesting chemical groups for the design of new antifungal compounds. The compounds inhibited ICL of the dimorphic fungus P. brasiliensis activity. The compound 4559339 presented minimum inhibitory concentration of 7.3 μg/ml in P. brasiliensis with fungicidal effect at this concentration. Thus, a new potential antifungal against P. brasiliensis is proposed.
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Image_3_Metabolic Peculiarities of Paracoccidioides brasiliensis Dimorphism as Demonstrated by iTRAQ Labeling Proteomics.TIF
2019Co-Authors: Danielle Silva Araujo, Maristela Pereira, Leandro Do Prado Assuncao, Lilian Cristiane Baeza, Igor Godinho Portis, Agenor De Castro Moreira Dos Santos, Wagner Fontes, Marcelo Valle De Sousa, Alexandre Mello Bailão, Carlos André Ornelas RicartAbstract: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.
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Metabolic Peculiarities of Paracoccidioides brasiliensis Dimorphism as Demonstrated by iTRAQ Labeling Proteomics
Frontiers Media S.A., 2019Co-Authors: Maristela Pereira, Leandro Do Prado Assuncao, Danielle Silva Araujo, Igor Godinho Portis, Agenor De Castro Moreira Dos Santos, Wagner Fontes, Marcelo Valle De Sousa, Lilian Cristiane BaezaAbstract: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
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Table_1_Metabolic Peculiarities of Paracoccidioides brasiliensis Dimorphism as Demonstrated by iTRAQ Labeling Proteomics.XLSX
2019Co-Authors: Danielle Silva Araujo, Maristela Pereira, Leandro Do Prado Assuncao, Lilian Cristiane Baeza, Igor Godinho Portis, Agenor De Castro Moreira Dos Santos, Wagner Fontes, Marcelo Valle De Sousa, Alexandre Mello Bailão, Carlos André Ornelas RicartAbstract: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.
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inhibition of Paracoccidioides lutzii pb01 isocitrate lyase by the natural compound argentilactone and its semi synthetic derivatives
PLOS ONE, 2014Co-Authors: Renata Silva Do Prado, Célia Maria De Almeida Soares, Roosevelt Alves Da Silva, Lucilia Kato, Cecília M. A. De Oliveira, Guilherme Oliveira Quintino, Ricardo Justino Alves, Silvio Cunha, Maristela PereiraAbstract:The dimorphic fungus Paracoccidioides spp. is responsible for paracoccidioidomycosis, the most prevalent systemic mycosis in Latin America, causing serious public health problems. Adequate treatment of mycotic infections is difficult, since fungi are eukaryotic organisms with a structure and metabolism similar to those of eukaryotic hosts. In this way, specific fungus targets have become important to search of new antifungal compound. The role of the glyoxylate cycle and its enzymes in microbial virulence has been reported in many fungal pathogens, including Paracoccidioides spp. Here, we show the action of argentilactone and its semi-synthetic derivative reduced argentilactone on recombinant and native isocitrate lyase from Paracoccidioides lutzii Pb01 (PbICL) in the presence of different carbon sources, acetate and glucose. Additionally, argentilactone and its semi-synthetic derivative reduced argentilactone exhibited relevant inhibitory activity against P. lutzii Pb01 yeast cells and dose-dependently influenced the transition from the mycelium to yeast phase. The other oxygenated derivatives tested, epoxy argentilactone and diol argentilactone-, did not show inhibitory action on the fungus. The results were supported by in silico experiments.
Silva, Mirelle Garcia - One of the best experts on this subject based on the ideXlab platform.
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Captação de ferro mediada por sideróforos em Paracoccidioides spp
2014Co-Authors: Silva, Mirelle GarciaAbstract:Tese (doutorado)—Universidade de Brasília, Programa de Pós-Graduação em Patologia Molecular, 2014.O gênero Paracoccidioides inclui espécies fúngicas termodimórficas, causadoras da paracoccidioidomicose, uma micose sistêmica endêmica da América Latina. A infecção ocorre quando propágulos micelianos ou conídios são inalados pelo hospedeiro. Após conversão para levedura nos alvéolos pulmonares o fungo pode disseminar-se para outros órgãos e tecidos. O ferro é um micronutriente essencial para todos os eucariotos, pois participa de vários processos biológicos essenciais. Entretanto, a biodisponibilidade deste metal dentro do hospedeiro é baixa. Como consequência, micro-organismos patogênicos desenvolveram mecanismos de aquisição de alta afinidade como forma de obter ferro durante a infecção. O presente trabalho descreve a aquisição de ferro mediada por sideróforos neste fungo. Análises in silico demonstraram que as espécies do gênero Paracoccidioides possuem todos os genes necessários para síntese e captação de sideróforos, os quais são produzidos em condições de depleção de ferro. Análises de cromatografia líquida de fase reversa e espectrometria de massas revelaram que Paracoccidioides spp. produz sideróforos do tipo hidroxamato. O fungo sintetiza e secreta coprogeno B, o qual gera ácido dimerúmico como produto de degradação, e também produz ferricrocina e ferricromo C como sideróforos intracelulares. Adicionalmente, Paracoccidioides spp. é capaz de crescer na presença de sideróforos como única fonte de ferro, demonstrando que além de produzir, o fungo também utiliza siderofóros para o crescimento, incluindo o xenosideróforo ferrioxamina. A exposição prévia a ferrioxamina aumentou a sobrevivência de Paracoccidioides spp. após fagocitose por macrófagos ativados. Além disso, o fungo provavelmente induz a síntese de sideróforos quando no interior destas células, demonstrando que estas moléculas provavelmente desempenham papel importante durante a interação patógeno-hospedeiro. Ademais, sideróforos produzidos por Paracoccidioides spp. podem ser utilizados como fontes de ferro por Aspergillus nidulans. Em conjunto, estes dados demonstraram que a síntese e a utilização de sideróforos são mecanismos empregados por Paracoccidioides spp. para superar a limitação de ferro. Como a escassez deste micronutriente é encontrada no hospedeiro, a produção de sideróforos está provavelmente relacionada à patogenicidade e virulência do fungo e representa um possível alvo para terapia com antifúngicos levando-se em consideração a ausência de tal via em humanos. _______________________________________________________________________________________ ABSTRACTThe genus Paracoccidioides includes termodimorphic fungal species which causes paracoccidioidomycosis, a systemic mycosis endemic in Latin America. The infection occurs when mycelium propagules or conidia are inhaled by the host. After conversion to yeast in the alveoli the fungus may disseminate to other organs and tissues. Iron is an essential micronutrient for all eukaryotes, since it participates in a variety of essential biological processes. However, the bioavailability of this metal is low inside the host. As a consequence, pathogenic microorganisms evolved high affinity acquisition mechanisms to obtain iron during infection. Here we describe the siderophore mediated iron acquisition in this fungus. In silico analysis demonstrated that species from Paracoccidioides genus possess all the necessary genes for synthesis and uptake of siderophores, which are produced under iron limiting conditions. Reversed phase liquid chromatography and mass spectrometry analysis revealed that Paracoccidioides spp. produce siderophores of hydroxamate type. The fungus synthesizes and secretes coprogen B, which generates dimerumic acid as a breakdown product, and also produces ferricrocin and ferrichrome C as intracellular siderophores. Moreover, Paracoccidioides spp. is able to grow in presence of siderophores as the only iron sources, demonstrating that beyond producing, the fungus also utilizes siderophores for growth, including the xenosiderophore ferrioxamine. Previous exposure to ferrioxamine increased Paracoccidioides spp. survival following phagocytosis by activated macrophages. Moreover, the fungus probably induces siderophore synthesis inside these cells, demonstrating that these iron chelators play an important role during host-pathogen interaction. Additionally, siderophores produced by Paracoccidioides spp. can be utilized as iron sources by Aspergillus nidulans. Altogether, these data demonstrated that synthesis and utilization of siderophores are mechanisms employed by Paracoccidioides spp. to surpass iron limitation. As iron paucity is found within the host, siderophore production may be related to fungus pathogenicity and virulence and represents a possible target for antifungal therapy since these pathway is absent in humans
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Captação de ferro mediada por sideróforos em Paracoccidioides spp
2014Co-Authors: Silva, Mirelle GarciaAbstract:O gênero Paracoccidioides inclui espécies fúngicas termodimórficas, causadoras da paracoccidioidomicose, uma micose sistêmica endêmica da América Latina. A infecção ocorre quando propágulos micelianos ou conídios são inalados pelo hospedeiro. Após conversão para levedura nos alvéolos pulmonares o fungo pode disseminar-se para outros órgãos e tecidos. O ferro é um micronutriente essencial para todos os eucariotos, pois participa de vários processos biológicos essenciais. Entretanto, a biodisponibilidade deste metal dentro do hospedeiro é baixa. Como consequência, micro-organismos patogênicos desenvolveram mecanismos de aquisição de alta afinidade como forma de obter ferro durante a infecção. O presente trabalho descreve a aquisição de ferro mediada por sideróforos neste fungo. Análises in silico demonstraram que as espécies do gênero Paracoccidioides possuem todos os genes necessários para síntese e captação de sideróforos, os quais são produzidos em condições de depleção de ferro. Análises de cromatografia líquida de fase reversa e espectrometria de massas revelaram que Paracoccidioides spp. produz sideróforos do tipo hidroxamato. O fungo sintetiza e secreta coprogeno B, o qual gera ácido dimerúmico como produto de degradação, e também produz ferricrocina e ferricromo C como sideróforos intracelulares. Adicionalmente, Paracoccidioides spp. é capaz de crescer na presença de sideróforos como única fonte de ferro, demonstrando que além de produzir, o fungo também utiliza siderofóros para o crescimento, incluindo o xenosideróforo ferrioxamina. A exposição prévia a ferrioxamina aumentou a sobrevivência de Paracoccidioides spp. após fagocitose por macrófagos ativados. Além disso, o fungo provavelmente induz a síntese de sideróforos quando no interior destas células, demonstrando que estas moléculas provavelmente desempenham papel importante durante a interação patógeno-hospedeiro. Ademais, sideróforos produzidos por Paracoccidioides spp. podem ser utilizados como fontes de ferro por Aspergillus nidulans. Em conjunto, estes dados demonstraram que a síntese e a utilização de sideróforos são mecanismos empregados por Paracoccidioides spp. para superar a limitação de ferro. Como a escassez deste micronutriente é encontrada no hospedeiro, a produção de sideróforos está provavelmente relacionada à patogenicidade e virulência do fungo e representa um possível alvo para terapia com antifúngicos levando-se em consideração a ausência de tal via em humanos. _______________________________________________________________________________________ ABSTRACTThe genus Paracoccidioides includes termodimorphic fungal species which causes paracoccidioidomycosis, a systemic mycosis endemic in Latin America. The infection occurs when mycelium propagules or conidia are inhaled by the host. After conversion to yeast in the alveoli the fungus may disseminate to other organs and tissues. Iron is an essential micronutrient for all eukaryotes, since it participates in a variety of essential biological processes. However, the bioavailability of this metal is low inside the host. As a consequence, pathogenic microorganisms evolved high affinity acquisition mechanisms to obtain iron during infection. Here we describe the siderophore mediated iron acquisition in this fungus. In silico analysis demonstrated that species from Paracoccidioides genus possess all the necessary genes for synthesis and uptake of siderophores, which are produced under iron limiting conditions. Reversed phase liquid chromatography and mass spectrometry analysis revealed that Paracoccidioides spp. produce siderophores of hydroxamate type. The fungus synthesizes and secretes coprogen B, which generates dimerumic acid as a breakdown product, and also produces ferricrocin and ferrichrome C as intracellular siderophores. Moreover, Paracoccidioides spp. is able to grow in presence of siderophores as the only iron sources, demonstrating that beyond producing, the fungus also utilizes siderophores for growth, including the xenosiderophore ferrioxamine. Previous exposure to ferrioxamine increased Paracoccidioides spp. survival following phagocytosis by activated macrophages. Moreover, the fungus probably induces siderophore synthesis inside these cells, demonstrating that these iron chelators play an important role during host-pathogen interaction. Additionally, siderophores produced by Paracoccidioides spp. can be utilized as iron sources by Aspergillus nidulans. Altogether, these data demonstrated that synthesis and utilization of siderophores are mechanisms employed by Paracoccidioides spp. to surpass iron limitation. As iron paucity is found within the host, siderophore production may be related to fungus pathogenicity and virulence and represents a possible target for antifungal therapy since these pathway is absent in humans
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employing proteomic analysis to compare Paracoccidioides lutzii yeast and mycelium cell wall proteins
Biochimica et Biophysica Acta, 2017Co-Authors: Danielle Silva Araujo, Clayton Luiz Borges, Alexandre Melo Bailão, Lilian Cristiane Baeza, Patrícia De Sousa Lima, Ana Flavia Alves Parente, Célia Maria De Almeida SoaresAbstract:Paracoccidioidomycosis is an important systemic mycosis caused by thermodimorphic fungi of the Paracoccidioides genus. During the infective process, the cell wall acts at the interface between the fungus and the host. In this way, the cell wall has a key role in growth, environment sensing and interaction, as well as morphogenesis of the fungus. Since the cell wall is absent in mammals, it may present molecules that are described as target sites for new antifungal drugs. Despite its importance, up to now few studies have been conducted employing proteomics in for the identification of cell wall proteins in Paracoccidioides spp. Here, a detailed proteomic approach, including cell wall-fractionation coupled to NanoUPLC-MSE, was used to study and compare the cell wall fractions from Paracoccidioides lutzii mycelia and yeast cells. The analyzed samples consisted of cell wall proteins extracted by hot SDS followed by extraction by mild alkali. In summary, 512 proteins constituting different cell wall fractions were identified, including 7 predicted GPI-dependent cell wall proteins that are potentially involved in cell wall metabolism. Adhesins previously described in Paracoccidioides spp. such as enolase, glyceraldehyde-3-phosphate dehydrogenase were identified. Comparing the proteins in mycelium and yeast cells, we detected some that are common to both fungal phases, such as Ecm33, and some specific proteins, as glucanase Crf1. All of those proteins were described in the metabolism of cell wall. Our study provides an important elucidation of cell wall composition of fractions in Paracoccidioides, opening a way to understand the fungus cell wall architecture.
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Paracoccidioides brasiliensis presents metabolic reprogramming and secretes a serine proteinase during murine infection
Virulence, 2017Co-Authors: Laurine Lacerda Pigosso, Clayton Luiz Borges, Alexandre Melo Bailão, Lilian Cristiane Baeza, Mariana Vieira Tomazett, Mariana Batista Rodrigues Faleiro, Veridiana Maria Brianezi Dignani De Moura, Juliana Alves Parente Rocha, Gabriel Fernandes, Gregory M GauthierAbstract:Paracoccidoides 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.
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analysis of Paracoccidioides secreted proteins reveals fructose 1 6 bisphosphate aldolase as a plasminogen binding protein
BMC Microbiology, 2015Co-Authors: Edilânia Gomes Araujo Chaves, Clayton Luiz Borges, Alexandre Melo Bailão, Luiz Augusto Pereira, Sônia Nair Báo, Simone Schneider Weber, Célia Maria De Almeida SoaresAbstract:Background Despite being important thermal dimorphic fungi causing Paracoccidioidomycosis, the pathogenic mechanisms that underlie the genus Paracoccidioides remain largely unknown. Microbial pathogens express molecules that can interact with human plasminogen, a protein from blood plasma, which presents fibrinolytic activity when activated into plasmin. Additionally, plasmin exhibits the ability of degrading extracellular matrix components, favoring the pathogen spread to deeper tissues. Previous work from our group demonstrated that Paracoccidioides presents enolase, as a protein able to bind and activate plasminogen, increasing the fibrinolytic activity of the pathogen, and the potential for adhesion and invasion of the fungus to host cells. By using proteomic analysis, we aimed to identify other proteins of Paracoccidioides with the ability of binding to plasminogen.
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predicting copper iron and zinc binding proteins in pathogenic species of the Paracoccidioides genus
Frontiers in Microbiology, 2015Co-Authors: Gabriel Tristao, Célia Maria De Almeida Soares, Clayton Luiz Borges, Leandro Do Prado Assuncao, Luiz Paulo Dos A Santos, Mirelle Garcia Silvabailao, Gabriele Cavallaro, Alexandre M BailaoAbstract: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 cooper, 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.
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transcriptional profile of Paracoccidioides induced by oenothein b a potential antifungal agent from the brazilian cerrado plant eugenia uniflora
BMC Microbiology, 2013Co-Authors: Patricia Fernanda Zambuzzicarvalho, Patricia Kott Tomazett, Wellington Santos Martins, Pedro Henrique Ferri, Suzana C. Santos, Célia Maria De Almeida Soares, Clayton Luiz Borges, Maristela PereiraAbstract:Background The compound oenothein B (OenB), which is isolated from the leaves of Eugenia uniflora, a Brazilian Cerrado plant, interferes with Paracoccidioides yeast cell morphology and inhibits 1,3-β-D-glucan synthase (PbFKS1) transcript accumulation, which is involved in cell wall synthesis. In this work we examined the gene expression changes in Paracoccidioides yeast cells following OenB treatment in order to investigate the adaptive cellular responses to drug stress.
