Navegando por Palavras-chave "trypanosoma cruzi"
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- ItemSomente MetadadadosCaracterização de uma proteína quinase do fator de início de tradução, eif2, regulada por heme e envolvida no controle do crescimento e diferenciação de trypanosoma cruzi(Universidade Federal de São Paulo (UNIFESP), 2014-11-30) Augusto, Leonardo da Silva [UNIFESP]; Schenkman, Sergio Schenkman [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)Environmental variations cause a decrease in protein synthesis at the expense of the translation of certain proteins by the phosphorylation of the ? subunit of translation initiation factor (eIF2?) of eukaryotes. This mechanism also seems to be essential to the alternation of developmental stages of protozoan parasites on their hosts. Therefore, in this thesis, we investigated the role of eIF2? phosphorylation in the biology of the Trypanosoma cruzi, the protozoan that causes Chagas? disease. We showed that this phosphorylation is essential for the generation of infective forms. Unlike other eukaryotes, the eIF2? of Trypanosoma have an N-terminal domain that is phosphorylated at the serine 43, besides the canonical site of phosphorylation of threonine 169, equivalent to the serine 51 of other organisms. Phosphorylation of both residues is involved in triggering the differentiation of T. cruzi. In addition, we study one of the three eIF2? kinases foretold in the genome of trypanosomes. We showed that the protein kinase 2, called TcK2, is inserted into the endosomal compartment membranes of proliferative forms of T. cruzi, where are stored proteins, lipids and heme. The heme binds to TcK2 causing its inhibition and enabling proliferation. In the absence of the heme TcK2 is activated leading to a halt in protein synthesis and formation of infective forms. TcK2 gene deletion prevented differentiation, and heme was not retained in the endosomes. The parasite showed increased levels of H2O2 due to the inhibition of peroxidase and activation of superoxide dismutase activities, which results in an increase of reactive oxygen species causing damage to parasites. We concluded that TcK2 plays a key role in the development of T. cruzi through the control of protein synthesis and of the redox balance based on the quantities of heme available for the parasite.
- ItemSomente MetadadadosCaracterização molecular de uma nova família de proteínas de membrana de trypanosoma cruzi (tc-smp) similar à proteína de membrana (pssa-2) de trypanosoma brucei(Universidade Federal de São Paulo (UNIFESP), 2014-09-30) Martins, Nadini Oliveira [UNIFESP]; Silveira Filho, Jose Franco da Silveira Filho [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)We identified a new family of Trypanosoma cruzi membrane proteins (Tc-SMP)that shares 40% identity with the procyclic-form surface glycoproteins (PSSA-2) from Trypanosoma brucei, a stage-specific surface antigen with properties of type I integral membrane proteins. We identified 12 Tc-SMP genes in the T. cruzi genome (clone CL Brener) that encode 380-470 amino acid proteins with 2-3 conserved transmembrane domains. Tc-SMP genes are arranged in tandem and share high levels of identity between them, suggesting that they were evolved by gene duplication. The chromosome region harboring Tc-SMP loci is very well conserved across the Trypanosoma species of mammals, and also in T. grayi, isolated from African crocodiles, indicating that genes TcSMP existed in early trypanosome ancestry. Tc-SMP proteins can be classified according to their size into two groups: TcSMP_L (large) and Tc-SMP_S (small). Tc-SMP_L proteins are predicted to have three transmembrane (TM) domains, the first TM domain corresponding to a putative signal peptide. While Tc-SMP_S have two TM domains and the first could act as a signal peptide. While Tc-SMP_S have two TM domains and the first could act as a signalanchor. The presence of signal peptide suggests that some members of Tc-SMP family may be addressed to cell surface and/or secreted, which was confirmed by immunofluorescence and proteomic assays. Tc-SMP transcripts were identified in all stages of parasite life cycle. Although transcripts of various sizes have been detected in different parasite stages, Tc-SMP proteins express a single molecular mass protein of 40 kDa. This prediction is consistent with the reaction of an anti-Tc-SMP serum with a single band of the same molecular mass in extracts of different parasite forms. The presence of Tc-SMP protein at the parasite?s surface and in the secreted vesicles suggests that they may interact with molecules on the surface of the host cell by egulating the parasite-host cell interaction. By using invasion assays we demonstrated that a recombinant Tc-SMP inhibits 38% the rate of invasion of HeLa cells by metacyclic trypomastigotes. Our data suggests a role of Tc-SMP in the internalization of metacyclic trypomastigostes process in mammalian cells, to confirm the hypothesis that T. cruzi exploits multiple ligand-receptor interactions for invading mammalian cells. Although Tc-SMP proteins are minor components on the surface of T. cruzi, they can play important role in host-parasite interaction.
- ItemSomente MetadadadosDistribution of Trypanosoma cruzi stage-specific epitopes in cardiac muscle of Calomys callosus, BALB/c mice, and cultured cells infected with different infective forms(Elsevier B.V., 2007-07-01) Taniwaki, Noerni N.; Silva, Claudio Vieira da; Silva, Solange da; Mortara, Renato A.; Universidade Federal de São Paulo (UNIFESP); Secao Microscopia Eletron Inst Adolfo LutzTo examine whether distinct parasite infective forms or the mammalian host could affect the distribution of Trypanosoma cruzi stage-specific epitopes defined by monoclonal antibodies (Mabs) raised against mammalian-stage parasite forms, immunofluorescence studies followed the intracellular life cycle of the parasite in the cardiac muscle of Calomys callosus and BALB/c mice in the acute phase of the disease and in LLC-MK(2) Cultured cells. Animals and cells were infected either with tissue-culture derived trypomastigotes (TCT) or bloodstream trypornastigotes (BT) from the Y strain of T cruzi. Samples were examined under confocal fluorescence microscopy after labeling with Mabs 2C2, 1D9, 2B7, 3G8, 3B9, and 4B9 that react with carbohydrate epitopes on Ssp-4, a major amastigote surface glycoprotein; Mab 4B5 that identifies a noncarbohydrate epitope on all intracellular parasites stages, and Mab 3B2 that also recognizes a noncarbohydrate epitope expressed only in flagellated forms. Samples were double labeled with DAPI to visualize parasites' kinetoplasts and nuclei. Most of the Mabs used in this work displayed a surface labeling pattern on amastigotes present in Calomys and mice hearts, and in LLC-MK2 cultured cells infected with BT or TCT. Mab 2B7, however, displayed a marked polymorphic distribution in antigen expression between both mammalian hosts, independent on the infective form. Beyond the polymorphic distribution of amastigote surface epitopes, Calomys, and mice heart sections presented several inflammatory cells around amastigotes and trypornastigotes nests. (c) 2007 Elsevier B.V. All rights reserved.
- ItemSomente MetadadadosEstudo das acetilações da histona h4 em trypanosoma cruzi(Universidade Federal de São Paulo (UNIFESP), 2015-05-27) Ramos, Thiago Cesar Prata [UNIFESP]; Schenkman, Sergio Schenkman [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)Histones have a series of post-translational modifications such as acetylation, methylation, fosforizações, ADP-ribosilações and ubiquitinações, that modulate access of these factors regulating replication processes, repair and recombination of DNA and transcription of mRNA. Histones are conserved in most eukaryotes. However, trypanosoma, which include protozoan parasites that early diverged during evolution, have very different histones, differing in sequence, charge and / or size when compared to their counterparts in eukaryotes. In addition, trypanosomatides have specific mechanisms of control of gene expression. In our laboratory we found that the acetylated histone 4 can be in three different lysines (K4, K10 and K14) and obtained antibodies that specifically recognize each of these modifications. Unpublished results from our group show that acetylation at K10 and K14 vary during the cell cycle and in states where there is DNA repair. In this project, we aim to understand the role of these changes, especially in K10 and K14 in Trypanosoma cruzi. For this we plan to: 1) analyze the effect of deacetylase inhibitors, 2) change the susceptibility of histone H4 expressing this modified protein in the parasite itself, 3) to evaluate the effect on the over-expression of acetylation and knockout of proteins involved in repair DNA, 4) identify acetylases and deacilases directly involved in these changes, and 5) generate lines that over-express or have decreased expression for these acetylases. Together, we hope to understand the relationship between acetylation of histone H4 and replication processes, DNA repair and transcription Trypanosoma.
- ItemSomente MetadadadosInvasion of MDCK epithelial cells with altered expression of Rho GTPases by Trypanosoma cruzi amastigotes and metacyclic trypomastigotes of strains from the two major phylogenetic lineages(Elsevier B.V., 2004-04-01) Fernandes, A. B.; Mortara, R. A.; Universidade Federal de São Paulo (UNIFESP)In order to invade mammalian cells, Trypanosoma cruzi infective forms cause distinct rearrangements of membrane and host cell cytoskeletal components. Rho GTPases have been shown to regulate three separate signal transduction pathways, linking plasma membrane receptors to the assembly of distinct actin filament structures. Here, we examined the role of Rho GTPases on the interaction between different T cruzi infective forms of strains from the two major phylogenetic lineages with nonpolarized MDCK cells transfected with different Rho GTPase constructs. We compared the infectivity of amastigotes isolated from infected cells (intracellular amastigotes) with forms generated from the axenic differentiation of trypomastigotes (extracellular amastigotes), and also with metacyclic trypomastigotes.No detectable effect of GTPase expression was observed on metacyclic trypomastigote invasion and parasites of Y and CL (T cruzi 11) strains invaded to similar degrees all MDCK transfectants, and were more infective than either G or Tulahuen (T cruzi I) strains. Intracellular amastigotes were complement sensitive and showed very low infectivity towards the different transfectants regardless of the parasite strain. Complement-resistant T cruzi I extracellular amastigotes, especially of the G strain, were more infective than T cruzi 11 parasites, particularly for the Rac1V12 constitutively active GTPase transfectant. the fact that in Rac1N17 dominant-negative cells, the invasion of G strain extracellular amastigotes was specifically inhibited suggested an important role for Rac1 in this process. (C) 2004 Elsevier SAS. All rights reserved.
- ItemSomente MetadadadosTrypanosoma cruzi infection by oral route How the interplay between parasite and host components modulates infectivity(Elsevier B.V., 2008-06-01) Yoshida, Nobuko [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)Trypanosoma cruzi infection by oral route constitutes the most important mode of transmission in some geographical regions, as illustrated by reports on microepidemics and outbreaks of acute Chagas' disease acquired by ingestion of food contaminated with parasites from triatomine insects. in the mouse model, T cruzi metacyclic trypomastigotes invade the gastric mucosal epithelium, a unique portal of entry for systemic infection. High efficiency of metacyclic forms in establishing infection by oral route is associated with expression of gp82, a stage-specific surface molecule that binds to gastric mucin and to epithelial cells. Gp82 promotes parasite entry by triggering the signaling cascades leading to intracellular Ca2+ mobilization. T cruzi strains deficient in gp82 can effectively invade cells in vitro, by engaging the Ca2+ signal-inducing surface glycoprotein gp30. However, they are poorly infective in mice by oral route because gp30 has low affinity for gastric mucin. Metacyclic forms also express gp90, a stage-specific surface glycoprotein that binds to host cells and acts as a negative regulator of invasion. T cruzi strains expressing gp90 at high levels, in addition to gp82 and gp30, are all poor cell invaders in vitro. Notwithstanding, their infectivity by oral route may vary because, unlike gp82 and gp30, which resist degradation by pepsin in the gastric milieu, the gp90 isoforms of different strains have varying susceptibility to peptic digestion. for instance, in a T cruzi isolate, derived from an acute case of Chagas' disease acquired by oral route, gp90 is extensively degraded by gastric juice in the mouse stomach and this renders the parasite highly invasive towards target cells. If such an exacerbation of infectivity occurs in humans, it may be responsible for the severity of the disease reported in outbreaks of oral infection. (c) 2007 Elsevier Ireland Ltd. All rights reserved.