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It is widely accepted that science is universal by nature. However, to make science universal, access to research findings is imperative. The open access model of publication of academic articles was established and consolidated during the last two decades. However, most of the open access journals apply article-processing charges (APCs), which can cost more than USD 10,000.00. In regions where support for research is scarce, these funds are usually not available. Similar problems occur in countries with weak economies and, consequently, unfavorable currency conversion rates. This situation reveals a barrier to the alleged universality of science and the access to research findings. In this manuscript, the barriers faced by authors and institutions from low-to-middle income regions to cover APCs and make their science freely available are discussed and illustrated with recent numbers.
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Brazilian science is under attack. In this manuscript, we will discuss the most recent events that, if not reverted, will make Brazilian science inviable. We urge the scientific community in Brazil and abroad to stand up and resist in defense of more than a century of essential scientific contributions.
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Paracoccidioides brasiliensis and P. lutzii cause human paracoccidioidomycosis. We have previously characterized the <200-nt RNA sub-populations contained in fungal extracellular vesicles (EVs) from P. brasiliensis Pb18 and other pathogenic fungi. We have presently used the RNA-seq strategy to compare the <200- and >200-nt RNA fractions contained in EVs isolated from culture supernatants of P. brasiliensis Pb18, Pb3, and P. lutzii Pb01. Shared mRNA sequences were related to protein modification, translation, and DNA metabolism/biogenesis, while those related to transport and oxidation-reduction were exclusive to Pb01. The presence of functional full-length mRNAs was validated by in vitro translation. Among small non-coding (nc)RNA, 15 were common to all samples; small nucleolar (sno)RNAs were enriched in P. brasiliensis EVs, whereas for P. lutzii there were similar proportions of snoRNA, rRNA, and tRNA. Putative exonic sRNAs were highly abundant in Pb18 EVs. We also found sRNA sequences bearing incomplete microRNA structures mapping to exons. RNA-seq data suggest that extracellular fractions containing Pb18 EVs can modulate the transcriptome of murine monocyte-derived dendritic cells in a transwell system. Considering that sRNA classes are involved in transcription/translation modulation, our general results may indicate that differences in virulence among fungal isolates can be related to their distinct EV-RNA content.
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Vesículas Extracelulares/genética , Paracoccidioides , Paracoccidioidomicose/microbiologia , RNA/genética , Animais , Células Cultivadas , Paracoccidioides/genética , Paracoccidioides/patogenicidade , VirulênciaRESUMO
BACKGROUND: Chagas' disease, caused by Trypanosoma cruzi, was described for the first time over a hundred years ago. Nonetheless, clinically available drugs still lack effective and selective properties. Nitric oxide (NO) produced by activated macrophages controls the progression of disease by killing the parasite. METHODS AND RESULTS: Here, chitosan nanoparticles (CS NPs) were synthesized and mercaptosuccinic acid (MSA), the NO donor precursor, was encapsulated into CS NPs, forming MSA-CS NPs, which had hydrodynamic size of 101.0±2.535 nm. Encapsulated MSA was nitrosated forming NO donor S-nitrosomercaptosuccinic acid-containing nanoparticles (S-nitroso-MSA-CS NPs). Kinetic data revealed a sustained release of NO from the nanoparticles. S-nitroso-MSA-CS NPs inhibited epimastigote proliferation and trypomastigote viability of T. cruzi, with IC50=75.0±6.5 µg·mL-1 and EC50=25.0±5.0 µg·mL-1, respectively. Treatment of peritoneal macrophages with nanoparticles decreased the number of T. cruzi-infected cells and the average number of intracellular replicative amastigotes per infected cells. Besides, the results have showed a selective behaviour of S-nitroso-MSA-CS NPs to parasites. Morphological and biochemical changes induced by these NO-releasing nanoparticles, such as cell shrinkage, cell cycle arrest, mitochondrial membrane depolarization and phosphatidylserine exposure on cell surface indicate that epimastigotes death is associated to the apoptotic pathway. CONCLUSION: S-nitroso-MSA-CS NPs are promising nanocarriers for the treatment of Chagas's disease.
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Antiprotozoários/farmacologia , Quitosana/farmacologia , Nanopartículas/química , Óxido Nítrico/química , Trypanosoma cruzi/efeitos dos fármacos , Animais , Quitosana/química , Quitosana/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/parasitologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Testes de Sensibilidade Microbiana , Óxido Nítrico/metabolismo , Testes de Sensibilidade ParasitáriaRESUMO
BACKGROUND Eukaryotic ribonucleoprotein (RNP) granules are important for the regulation of RNA fate. RNP granules exist in trypanosomatids; however, their roles in controlling gene expression are still not understood. XRNA is a component of granules in Trypanosoma brucei but has not been investigated in Trypanosoma cruzi. OBJECTIVES This study aimed to investigate the TcXRNA dynamic assembly and its interaction with RNP components under conditions that affect the mRNA availability. METHODS We used in vitro metacyclogenesis of T. cruzi to observe changes in RNP granules during the differentiation process. TcXRNA expression was analysed by Western blot and immunofluorescence. Colocalisation assays were performed to investigate the interaction of TcXRNA with other RNP components. FINDINGS TcXRNA is constantly present during metacyclogenesis and is localised in cytoplasmic granules. TcXRNA does not colocalise with TcDHH1 and TcCAF1 granules in the cytoplasm. However, TcXRNA granules colocalise with mRNP granules at the nuclear periphery when mRNA processing is inhibited. MAIN CONCLUSIONS TcXRNA plays a role in mRNA metabolism as a component of mRNP granules whose assembly is dependent on mRNA availability. TcXRNA granules colocalise with distinct RNP granules at the nuclear periphery, suggesting that the perinuclear region is a regulatory compartment in T. cruzi mRNA metabolism.
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Humanos , RNA/sangue , RNA Mensageiro/análise , Metaciclina/uso terapêutico , RNA Nuclear PequenoRESUMO
Abstract RNA-binding proteins (RBPs) have important functions in the regulation of gene expression. RBPs play key roles in post-transcriptional processes in all eukaryotes, such as splicing regulation, mRNA transport and modulation of mRNA translation and decay. RBPs assemble into different mRNA-protein complexes, which form messenger ribonucleoprotein complexes (mRNPs). Gene expression regulation in trypanosomatids occurs mainly at the post-transcriptional level and RBPs play a key role in all processes. However, the functional characterization of RBPs in Trypanosoma cruzi has been impaired due to the lack of reliable reverse genetic manipulation tools. The comparison of RBPs from Saccharomyces cerevisiae and T. cruzi might allow inferring on the function of these proteins based on the information available for the orthologous RNA-binding proteins from the S. cerevisiae model organism. In this review, we discuss the role of some RBPs from T. cruzi and their homologues in regulating gene expression in yeast.
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Asymmetric mRNA localization is a sophisticated tool for regulating and optimizing protein synthesis and maintaining cell polarity. Molecular mechanisms involved in the regulated localization of transcripts are widespread in higher eukaryotes and fungi, but not in protozoa. Trypanosomes are ancient eukaryotes that branched off early in eukaryote evolution. We hypothesized that these organisms would have basic mechanisms of mRNA localization. FISH assays with probes against transcripts coding for proteins with restricted distributions showed a discrete localization of the mRNAs in the cytoplasm. Moreover, cruzipain mRNA was found inside reservosomes suggesting new unexpected functions for this vacuolar organelle. Individual mRNAs were also mobilized to RNA granules in response to nutritional stress. The cytoplasmic distribution of these transcripts changed with cell differentiation, suggesting that localization mechanisms might be involved in the regulation of stage-specific protein expression. Transfection assays with reporter genes showed that, as in higher eukaryotes, 3'UTRs were responsible for guiding mRNAs to their final location. Our results strongly suggest that Trypanosoma cruzi have a core, basic mechanism of mRNA localization. This kind of controlled mRNA transport is ancient, dating back to early eukaryote evolution.
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Transporte de RNA , RNA Mensageiro/metabolismo , Trypanosoma cruzi/metabolismo , Regiões 3' não Traduzidas/genética , Animais , Compartimento Celular , Cisteína Endopeptidases/genética , Cisteína Endopeptidases/metabolismo , Citoplasma/metabolismo , Hibridização in Situ Fluorescente , Estágios do Ciclo de Vida , Luciferases/metabolismo , Parasitos/crescimento & desenvolvimento , Parasitos/metabolismo , Proteínas de Protozoários/metabolismo , Estresse Fisiológico , Frações Subcelulares/metabolismo , Trypanosoma cruzi/crescimento & desenvolvimentoRESUMO
Adipocyte stem cells (hASCs) can proliferate and self-renew and, due to their multipotent nature, they can differentiate into several tissue-specific lineages, making them ideal candidates for use in cell therapy. Most attempts to determine the mRNA profile of self-renewing or differentiating stem cells have made use of total RNA for gene expression analysis. Several lines of evidence suggest that self-renewal and differentiation are also dependent on the control of protein synthesis by posttranscriptional mechanisms. We used adipogenic differentiation as a model, to investigate the extent to which posttranscriptional regulation controlled gene expression in hASCs. We focused on the initial steps of differentiation and isolated both the total mRNA fraction and the subpopulation of mRNAs associated with translating ribosomes. We observed that adipogenesis is committed in the first days of induction and three days appears as the minimum time of induction necessary for efficient differentiation. RNA-seq analysis showed that a significant percentage of regulated mRNAs were posttranscriptionally controlled. Part of this regulation involves massive changes in transcript untranslated regions (UTR) length, with differential extension/reduction of the 3'UTR after induction. A slight correlation can be observed between the expression levels of differentially expressed genes and the 3'UTR length. When we considered association to polysomes, this correlation values increased. Changes in the half lives were related to the extension of the 3'UTR, with longer UTRs mainly stabilizing the transcripts. Thus, changes in the length of these extensions may be associated with changes in the ability to associate with polysomes or in half-life.
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Adipócitos/fisiologia , Polirribossomos/fisiologia , Células-Tronco/fisiologia , Regiões 3' não Traduzidas , Adipócitos/citologia , Adipócitos/metabolismo , Adipogenia , Adulto , Diferenciação Celular/fisiologia , Feminino , Regulação da Expressão Gênica , Glutationa/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Polirribossomos/genética , Polirribossomos/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Células-Tronco/citologia , Células-Tronco/metabolismo , Transcrição Gênica , Adulto JovemRESUMO
To characterise the trypanosomatid-exclusive RNA-binding protein TcRBP19, we analysed the phenotypic changes caused by its overexpression. Although no evident changes were observed when TcRBP19 was ectopically expressed in epimastigotes, the metacyclogenesis process was affected. Notably, TcRBP19 overexpression also led to a decrease in the number of infected mammalian cells. These findings suggest that TcRBP19 may be involved in the life cycle progression of the Trypanosoma cruzi parasite.
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Animais , Proteínas de Protozoários/fisiologia , Proteínas de Ligação a RNA/genética , Trypanosoma cruzi/genética , Regulação da Expressão Gênica , Estágios do Ciclo de Vida , Processamento Pós-Transcricional do RNA/genética , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/fisiologia , Trypanosoma cruzi/crescimento & desenvolvimento , Trypanosoma cruzi/metabolismoRESUMO
Trypanosomes are parasitic protozoa in which gene expression is primarily controlled through the regulation of mRNA stability and translation. This post-transcriptional control is mediated by various families of RNA-binding proteins, including those with zinc finger CCCH motifs. CCCH zinc finger proteins have been shown to be essential to differentiation events in trypanosomatid parasites. Here, we functionally characterise TcZFP2 as a predicted post-transcriptional regulator of differentiation in Trypanosoma cruzi. This protein was detected in cell culture-derived amastigotes and trypomastigotes, but it was present in smaller amounts in metacyclic trypomastigote forms of T. cruzi. We use an optimised recombinant RNA immunopreciptation followed by microarray analysis assay to identify TcZFP2 target mRNAs. We further demonstrate that TcZFP2 binds an A-rich sequence in which the adenosine residue repeats are essential for high-affinity recognition. An analysis of the expression profiles of the genes encoding the TcZFP2-associated mRNAs throughout the parasite life cycle by microarray hybridisation showed that most of the associated mRNAs were upregulated in the metacyclic trypomastigote forms, also suggesting a role for TcZFP2 in metacyclic trypomastigote differentiation. Knockdown of the orthologous Trypanosoma brucei protein levels showed ZFP2 to be a positive regulator of specific target mRNA abundance.
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Proteínas de Ligação a DNA/metabolismo , Proteínas de Protozoários/metabolismo , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Trypanosoma cruzi/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Estabilidade de RNA , Trypanosoma cruzi/crescimento & desenvolvimentoRESUMO
Trypanosomes are parasitic protozoa in which gene expression is primarily controlled through the regulation of mRNA stability and translation. This post-transcriptional control is mediated by various families of RNA-binding proteins, including those with zinc finger CCCH motifs. CCCH zinc finger proteins have been shown to be essential to differentiation events in trypanosomatid parasites. Here, we functionally characterise TcZFP2 as a predicted post-transcriptional regulator of differentiation in Trypanosoma cruzi. This protein was detected in cell culture-derived amastigotes and trypomastigotes, but it was present in smaller amounts in metacyclic trypomastigote forms of T. cruzi. We use an optimised recombinant RNA immunopreciptation followed by microarray analysis assay to identify TcZFP2 target mRNAs. We further demonstrate that TcZFP2 binds an A-rich sequence in which the adenosine residue repeats are essential for high-affinity recognition. An analysis of the expression profiles of the genes encoding the TcZFP2-associated mRNAs throughout the parasite life cycle by microarray hybridisation showed that most of the associated mRNAs were upregulated in the metacyclic trypomastigote forms, also suggesting a role for TcZFP2 in metacyclic trypomastigote differentiation. Knockdown of the orthologous Trypanosoma brucei protein levels showed ZFP2 to be a positive regulator of specific target mRNA abundance.
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Proteínas de Ligação a DNA/metabolismo , Proteínas de Protozoários/metabolismo , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Trypanosoma cruzi/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Estabilidade de RNA , Trypanosoma cruzi/crescimento & desenvolvimentoRESUMO
Small non-coding RNAs derived from transfer RNAs have been identified as a broadly conserved prokaryotic and eukaryotic response to stress. Their presence coincides with changes in developmental state associated with gene expression regulation. In the epimastigote form of Trypanosoma cruzi, tRNA fragments localize to posterior cytoplasmic granules. In the infective metacyclic form of the parasite, we found tRNA-derived fragments to be abundant and evenly distributed within the cytoplasm. The fragments were not associated with polysomes, suggesting that the tRNA-derived fragments may not be directly involved in translation control in metacyclics.
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Grânulos Citoplasmáticos/genética , RNA de Protozoário/análise , RNA de Transferência/análise , Trypanosoma cruzi/genética , Grânulos Citoplasmáticos/química , RNA de Protozoário/genética , RNA de Transferência/genéticaRESUMO
Stem cells can either differentiate into more specialized cells or undergo self-renewal. Several lines of evidence from different organisms suggest that these processes depend on the post-transcriptional regulation of gene expression. The presence of the PUF [Pumilio/FBF (fem-3 binding factor)] domain defines a conserved family of RNA binding proteins involved in repressing gene expression. It has been suggested that a conserved function of PUF proteins is to repress differentiation and sustain the mitotic proliferation of stem cells. In humans, Pumilio-2 (PUM2) is expressed in embryonic stem cells and adult germ cells. Here we show that PUM2 is expressed in a subpopulation of adipose-derived stem cell (ASC) cultures, with a granular pattern of staining in the cytoplasm. Protein levels of PUM2 showed no changes during the differentiation of ASCs into adipocytes. Moreover, RNAi knockdown of pum2 did not alter the rate of adipogenic differentiation compared with wild-type control cells. A ribonomic approach was used to identify PUM2-associated mRNAs. Microarray analysis showed that PUM2-bound mRNAs are part of gene networks involved in cell proliferation and gene expression control. We studied pum2 expression in cell cultures with low or very high levels of proliferation and found that changes in pum2 production were dependent on the proliferation status of the cell. Transient knockdown of pum2 expression by RNAi impaired proliferation of ASCs in vitro. Our results suggest that PUM2 does not repress differentiation of ASCs but rather is involved in the positive control of ASCs division and proliferation.
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Adipócitos/metabolismo , Regulação da Expressão Gênica , Células-Tronco Mesenquimais/metabolismo , Mitose/genética , RNA Mensageiro/genética , Proteínas de Ligação a RNA/genética , Adipócitos/citologia , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Técnicas de Silenciamento de Genes , Inativação Gênica , Humanos , Células-Tronco Mesenquimais/citologia , Análise de Sequência com Séries de Oligonucleotídeos , RNA Interferente Pequeno/genéticaRESUMO
Trypanosoma cruzi alternates between different morphological and functional types during its life cycle. Since the discovery of this parasite at the beginning of the twentieth century, efforts have been made to determine the basis of its pathogenesis in the course of Chagas disease and its biochemical constituents. There has also been work to develop tools and strategies for prophylaxis of the important disease caused by these parasites which affects millions of people in Latin America. The identification of axenic conditions allowing T. cruzi growth and differentiation has led to the identification and characterization of stage-specific antigens as well as a better characterization of the biological properties and biochemical particularities of each individual developmental stage. The recent availability of genomic data should pave the way to new progress in our knowledge of the biology and pathogenesis of T. cruzi. This review addresses the differentiation and major stage-specific antigens of T. cruzi and attempts to describe the complexity of the parasite and of the disease it causes.
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Antígenos de Protozoários/imunologia , Estágios do Ciclo de Vida , Trypanosoma cruzi/crescimento & desenvolvimento , Animais , Adesão Celular , Doença de Chagas/imunologia , Doença de Chagas/parasitologia , Cisteína Proteases/metabolismo , Regulação da Expressão Gênica , Glicoproteínas/imunologia , Mamíferos , Neuraminidase/imunologia , Fosfoproteínas/imunologia , Proteínas de Protozoários/imunologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Trypanosoma cruzi/genética , Trypanosoma cruzi/imunologia , Trypanosoma cruzi/metabolismo , Glicoproteínas Variantes de Superfície de Trypanosoma/imunologiaRESUMO
In trypanosomatids, Ca²+-binding proteins can affect parasite growth, differentiation and invasion. Due to their importance for parasite maintenance, they become an attractive target for drug discovery and design. Phytomonas serpens 15T is a non-human pathogenic trypanosomatid that expresses important protein homologs of human pathogenic trypanosomatids. In this study, the coding sequence of calmodulin, a Ca²+-binding protein, of P. serpens 15T was cloned and characterized. The encoded polypeptide (CaMP) displayed high amino acid identity to homolog protein of Trypanosoma cruzi and four helix-loop-helix motifs were found. CaMP sequence analysis showed 20 amino acid substitutions compared to its mammalian counterparts. This gene is located on a chromosomal band with estimated size of 1,300 kb and two transcripts were detected by Northern blot analysis. A polyclonal antiserum raised against the recombinant protein recognized a polypeptide with an estimated size of 17 kDa in log-phase promastigote extracts. The recombinant CaMP retains its Ca²+-binding capacity.
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Calmodulina/química , Calmodulina/metabolismo , Clonagem Molecular , Proteínas de Protozoários/química , Proteínas de Protozoários/metabolismo , Trypanosomatina/genética , Motivos de Aminoácidos , Sequência de Aminoácidos , Cálcio/metabolismo , Calmodulina/genética , Humanos , Dados de Sequência Molecular , Ligação Proteica , Estrutura Secundária de Proteína , Proteínas de Protozoários/genética , Alinhamento de Sequência , Trypanosoma cruzi/química , Trypanosoma cruzi/genética , Trypanosoma cruzi/metabolismo , Trypanosomatina/química , Trypanosomatina/metabolismoRESUMO
The life cycle of the protozoan Trypanosoma cruzi exposes it to several environmental stresses in its invertebrate and vertebrate hosts. Stress conditions are involved in parasite differentiation, but little is known about the stress response proteins involved. We report here the first characterization of stress-induced protein-1 (STI-1) in T. cruzi (TcSTI-1). This co-chaperone is produced in response to stress and mediates the formation of a complex between the stress proteins HSP70 and HSP90 in other organisms. Despite the similarity of TcSTI-1 to STI-1 proteins in other organisms, its expression profile in response to various stress conditions, such as heat shock, acidic pH or nutrient starvation, is quite different. Neither polysomal mRNA nor protein levels changed in exponentially growing epimastigotes cultured under any of the stress conditions studied. Increased levels of TcSTI-1 were observed in epimastigotes subjected to nutritional stress in the late growth phase. Co-immunoprecipitation assays revealed an association between TcSTI-1 and TcHSP70 in T. cruzi epimastigotes. Immunolocalization demonstrated that TcSTI-1 was distributed throughout the cytoplasm and there was some colocalization of TcSTI-1 and TcHSP70 around the nucleus. Thus, TcSTI-1 associates with TcHSP70 and TcSTI-1 expression is induced when the parasites are subjected to stress conditions during specific growth phase.
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Proteínas de Choque Térmico , Trypanosoma cruzi , Núcleo Celular , Citoplasma , Imunofluorescência , Proteínas de Choque Térmico HSP90 , Proteínas de Choque Térmico HSP90 , Proteínas de Choque Térmico , ImunoprecipitaçãoRESUMO
The genus Phytomonas comprises trypanosomatids that can parasitize a broad range of plant species. These flagellates can cause diseases in some plant families with a wide geographic distribution, which can result in great economic losses. We have demonstrated previously that Phytomonas serpens 15T, a tomato trypanosomatid, shares antigens with Trypanosoma cruzi, the agent of human Chagas disease. Herein, two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS) were used to identify proteins of P. serpens 15T that are recognized by sera from patients with Chagas disease. After 2D-electrophoresis of whole-cell lysates, 31 peptides were selected and analyzed by tandem mass spectrometry. Twenty-eight polypeptides were identified, resulting in 22 different putative proteins. The identified proteins were classified into 8 groups according to biological process, most of which were clustered into a cellular metabolic process category. These results generated a collection of proteins that can provide a starting point to obtain insights into antigenic cross reactivity among trypanosomatids and to explore P. serpens antigens as candidates for vaccine and immunologic diagnosis studies.
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Antígenos de Protozoários/imunologia , Doença de Chagas/imunologia , Leishmania/imunologia , Solanum lycopersicum/parasitologia , Trypanosoma cruzi/imunologia , Animais , Antígenos de Protozoários/isolamento & purificação , Reações Cruzadas , Eletroforese em Gel Bidimensional , Ensaio de Imunoadsorção Enzimática , Humanos , Espectrometria de MassasRESUMO
The use of conditioned medium (CM) from human cardiac explants (HCEs) as a potential source of paracrine factors for adult stem cell signaling has never been evaluated. We hypothesized that HCEs might provide a source of soluble factors triggering the differentiation of mesenchymal stem cells (MSCs) into cardiomyocyte-like cells. By using two-dimensional electrophoresis (2-DE) gels/mass spectrometry and antibody macroarray assays, we found that HCEs release macromolecules, including cytokines, growth factors and myocardial and metabolism-related proteins into the culture medium. We identified a total of 20 proteins in the HCE-CM. However, as shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and 2-DE, these 20 proteins account for only a fraction of the total number of proteins present in the HCE-CM. We also found that CM increased the proliferation of bone marrow-derived-MSCs (BM-MSCs) in vitro. Unlike the other effects, this effect was most evident after 48 h of culture. Moreover, we examined the effect of HCE-CM on levels of mRNA and protein for specific cardiac markers. We showed that a surprisingly big fraction of BM-MSCs (3.4-5.0%) treated in vitro with HCE-CM became elongated and began to express cardiac markers, consistent with their possible differentiation into cardiomyocyte-like cells. Our in vitro model may be useful not only per se, but also for studies of the mechanisms of action of soluble factors involved in cell differentiation, paving the way for possible new protein-based treatments in the future.
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Células da Medula Óssea/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Meios de Cultivo Condicionados/química , Meios de Cultivo Condicionados/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Miócitos Cardíacos/efeitos dos fármacos , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Células Cultivadas , Técnicas de Cultura , Citocinas/química , Citocinas/farmacologia , Eletroforese em Gel Bidimensional , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/química , Peptídeos e Proteínas de Sinalização Intercelular/farmacologia , Espectrometria de Massas , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Análise em Microsséries , Miócitos Cardíacos/citologia , Miócitos Cardíacos/metabolismo , Proteínas/química , Proteínas/farmacologia , RNA Mensageiro/metabolismo , Solubilidade , Fatores de Tempo , Técnicas de Cultura de TecidosRESUMO
The genus Phytomonas comprises trypanosomatids that can parasitize a broad range of plant species. These fagellates can cause diseases in some plant families with a wide geographic distribution, which can result in great economic losses. We have demonstrated previously that Phytomonas serpens 15T, a tomato trypanosomatid, shares antigens with Trypanosoma cruzi, the agent of human Chagas disease. Herein, two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS) were used to identify proteins of P. serpens 15T that are recognized by sera from patients with Chagas disease. After 2D-electrophoresis of whole-cell lysates, 31 peptides were selected and analyzed by tandem mass spectrometry. Twenty-eight polypeptides were identifed, resulting in 22 different putative proteins. The identifed proteins were classifed into 8 groups according to biological process, most of which were clustered into a cellular metabolic process category. These results generated a collection of proteins that can provide a starting point to obtain insights into antigenic cross reactivity among trypanosomatids and to explore P. serpens antigens as candidates for vaccine and immunologic diagnosis studies.