scholarly journals Cell Adhesion and Ca2+ Signaling Activity in Stably Transfected Trypanosoma cruzi Epimastigotes Expressing the Metacyclic Stage-Specific Surface Molecule gp82

2003 ◽  
Vol 71 (3) ◽  
pp. 1561-1565 ◽  
Author(s):  
Patricio M. Manque ◽  
Ivan Neira ◽  
Vanessa D. Atayde ◽  
Esteban Cordero ◽  
Alice T. Ferreira ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Cell Invasion ◽  
Hela Cells ◽  
Plasmid Vector ◽  
Surface Glycoprotein ◽  
Target Cells ◽  
Developmentally Regulated ◽  
Intracellular Ca ◽  
Tyrosine Phosphorylated Protein ◽  
Metacyclic Trypomastigote

ABSTRACT Metacyclic trypomastigotes of Trypanosoma cruzi express a developmentally regulated 82-kDa surface glycoprotein (gp82) that has been implicated in host cell invasion. gp82-mediated interaction of metacyclic forms with target cells induces in both cells activation of the signal transduction pathways, leading to intracellular Ca2+ mobilization, which is required for parasite internalization. Noninfective epimastigotes do not express detectable levels of gp82 and are unable to induce a Ca2+ response. We stably transfected epimastigotes with a T. cruzi expression vector carrying the metacyclic stage gp82 cDNA. These transfectants produced a functional gp82, which bound to and triggered a Ca2+ response in HeLa cells, in the same manner as the metacyclic trypomastigote gp82. Such properties were not found in epimastigotes transfected with the plasmid vector alone. Epimastigotes expressing gp82 on the surface adhered to HeLa cells but were not internalized. Treatment of gp82-expressing epimastigotes with forskolin, an activator of adenylyl cyclase that increases the metacyclic trypomastigote entry into target cells, did not promote parasite internalization. P175, an intracellular tyrosine phosphorylated protein, which appears to play a role in gp82-dependent signaling cascade in metacyclic forms, was undetectable in epimastigotes, either transfected or not with pTEX-gp82. Overall, our results indicate that gp82 is required but not sufficient for target cell invasion.

scholarly journals Actin Cytoskeleton-Dependent and -Independent Host Cell Invasion by Trypanosoma cruzi Is Mediated by Distinct Parasite Surface Molecules

2006 ◽  
Vol 74 (10) ◽  
pp. 5522-5528 ◽  
Author(s):  
Daniele Ferreira ◽  
Mauro Cortez ◽  
Vanessa D. Atayde ◽  
Nobuko Yoshida
Keyword(s):  
Trypanosoma Cruzi ◽  
Actin Cytoskeleton ◽  
Host Cell ◽  
Cell Invasion ◽  
Hela Cells ◽  
Cytochalasin D ◽  
Target Cells ◽  
Host Cell Invasion ◽  
Latrunculin B ◽  
Surface Molecules

ABSTRACT The disassembly of host cell actin cytoskeleton as a facilitator of Trypanosoma cruzi invasion has been reported by some authors, while other workers claim that it instead inhibits internalization of the parasite. In this study we aimed at elucidating the basis of this discrepancy. We performed experiments with metacyclic trypomastigotes of T. cruzi strains G and CL, which differ markedly in infectivity and enter target cells by engaging the surface molecules gp35/50 and gp82, respectively, which have signaling activity. Treatment of HeLa cells with the F-actin-disrupting drug cytochalasin D or latrunculin B inhibited the invasion by strain G but not the invasion by strain CL. In contrast to cells penetrated by strain CL, which were previously shown to have a disrupted actin cytoskeleton architecture, no such alteration was observed in HeLa cells invaded by strain G, and parasites were found to be closely associated with target cell actin. Coinfection with enteroinvasive Escherichia coli (EIEC), which recruits host cell actin for internalization, drastically reduced entry of strain CL into HeLa cells but not entry of strain G. In contrast to gp82 in its recombinant form, which induces disruption of F-actin and inhibits EIEC invasion, purified mucin-like gp35/50 molecules promoted an increase in EIEC uptake by HeLa cells. These data, plus the finding that drugs that interfere with mammalian cell signaling differentially affect the internalization of metacyclic forms of strains G and CL, indicate that the host cell invasion mediated by gp35/50 is associated with signaling events that favor actin recruitment, in contrast to gp82-dependent invasion, which triggers the signaling pathways leading to disassembly of F-actin.

The Escherichia coli AIDA autotransporter adhesin recognizes an integral membrane glycoprotein as receptor

Microbiology ◽  
2003 ◽  
Vol 149 (7) ◽  
pp. 1871-1882 ◽  
Author(s):  
Sven Laarmann ◽  
M. Alexander Schmidt
Keyword(s):  
Hela Cells ◽  
Cell Types ◽  
Surface Glycoprotein ◽  
Receptor Protein ◽  
Target Cells ◽  
Cellular Receptor ◽  
Membrane Glycoprotein ◽  
Two Dimensional Gel Electrophoresis ◽  
Specific Attachment ◽  
Complementary Techniques

The AIDA-I autotransporter adhesin, as a prototype of the AIDA adhesin family, represents a tripartite antigen consisting of the functional adhesin AIDA-I (α-domain), which mediates the specific attachment of bacteria to target cells, and a two-domain translocator (AIDAc) organized in the β 1- and β 2-domains. Cellular receptor moieties for the adhesin AIDA-I have not been identified. Here, it is demonstrated that the purified adhesin binds specifically to a high-affinity class of receptors on HeLa cells. Additionally, the adhesin was found to bind to a variety of mammalian cell types, indicating a broad tissue distribution of the receptor moiety. By using complementary techniques, including co-immunoprecipitation and one- and two-dimensional gel electrophoresis, the AIDA-I binding protein on HeLa cells was identified as a surface glycoprotein of about 119 kDa (gp119). The gp119 AIDA-I cellular receptor protein was characterized biochemically and found to be an integral N-glycosylated membrane protein with a pI of 5·2.

scholarly journals Infection by Trypanosoma cruzi Metacyclic Forms Deficient in gp82 but Expressing a Related Surface Molecule, gp30

2003 ◽  
Vol 71 (11) ◽  
pp. 6184-6191 ◽  
Author(s):  
Mauro Cortez ◽  
Ivan Neira ◽  
Daniele Ferreira ◽  
Alejandro O. Luquetti ◽  
Anis Rassi ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Cell Invasion ◽  
Target Cell ◽  
Surface Profile ◽  
Cell Entry ◽  
Gastric Mucin ◽  
Surface Glycoprotein ◽  
Metacyclic Trypomastigotes ◽  
Mucosal Infection

ABSTRACT Trypanosoma cruzi metacyclic trypomastigotes invade and replicate in the gastric mucosal epithelium after oral infection. In this study we analyzed the process of infection by T. cruzi isolates deficient in the expression of gp82, the metacyclic stage-specific surface glycoprotein implicated in target cell entry in vitro and in promoting mucosal infection in mice after oral challenge. Mice infected by the oral route with metacyclic forms of gp82-deficient isolate 569 or 588 developed patent parasitemia but at greatly reduced levels compared to those infected with the gp82-expressing isolate CL. Metacyclic forms of both isolates expressed gp30, a surface glycoprotein detectable by monoclonal antibody (MAb) 3F6 directed to gp82. Otherwise, the gp82-deficient isolates displayed a surface profile similar to that of the CL isolate and also entered epithelial HeLa cells in a manner inhibitable by MAb 3F6 and dependent on the parasite signal transduction that involved the activation of protein tyrosine kinase and Ca2+ mobilization from thapsigargin-sensitive stores. Like gp82, gp30 triggered the host cell Ca2+ response required for parasite internalization. Purified gp30 and the recombinant gp82 inhibited HeLa cell invasion of metacyclic forms of isolates 569 and 588 by ∼90 and ∼70%, respectively. A cell invasion assay performed in the presence of gastric mucin, mimicking the in vivo infection, showed an inhibition of 70 to 75% in the internalization of gp82-deficient isolates but not of the CL isolate. The recombinant gp82 exhibited an adhesive capacity toward gastric mucin much higher than that of gp30. Taken together, our findings indicate that target cell entry of metacyclic trypomastigotes can be mediated either by gp82 or gp30 but that efficient mucosal infection depends on the expression of gp82.

scholarly journals Molecular basis of mammalian cell invasion by Trypanosoma cruzi

2006 ◽  
Vol 78 (1) ◽  
pp. 87-111 ◽  
Author(s):  
Nobuko Yoshida
Keyword(s):  
Trypanosoma Cruzi ◽  
Cell Invasion ◽  
Target Cell ◽  
Protein Tyrosine Kinase ◽  
Host Cells ◽  
Target Cells ◽  
Cytokeratin 18 ◽  
Parasite Protein ◽  
Oligopeptidase B ◽  
Surface Glycoproteins

Establishment of infection by Trypanosoma cruzi, the agent of Chagas' disease, depends on a series of events involving interactions of diverse parasite molecules with host components. Here we focus on the mechanisms of target cell invasion by metacyclic trypomastigotes (MT) and mammalian tissue culture trypomastigotes (TCT). During MT or TCT internalization, signal transduction pathways are activated both in the parasite and the target cell, leading to Ca2+ mobilization. For cell adhesion, MT engage surface glycoproteins, such as gp82 and gp35/50, which are Ca2+ signal-inducing molecules. In T. cruzi isolates that enter host cells in gp82-mediated manner, parasite protein tyrosine kinase as well as phospholipase C are activated, and Ca2+ is released from I P3-sensitive stores, whereas in T. cruzi isolates that attach to target cells mainly through gp35/50, the signaling pathway involving adenylate cyclase appears to be stimulated, with Ca2+ release from acidocalciosomes. In addition, T. cruzi isolate-dependent inhibitory signals, mediated by MT-specific gp90, may be triggered both in the host cell and the parasite. The repertoire of TCT molecules implicated in cell invasion includes surface glycoproteins of gp85 family, with members containing binding sites for laminin and cytokeratin 18, enzymes such as cruzipain, trans-sialidase, and an oligopeptidase B that generates a Ca2+-agonist from a precursor molecule.

scholarly journals Infectivity of Trypanosoma cruzi strains is associated with differential expression of surface glycoproteins with differential Ca2+ signalling activity

1998 ◽  
Vol 330 (1) ◽  
pp. 505-511 ◽  
Author(s):  
C. Rita RUIZ ◽  
Silvio FAVORETO ◽  
L. Miriam DORTA ◽  
E. M. Maria OSHIRO ◽  
T. Alice FERREIRA ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Hela Cells ◽  
Strain Analysis ◽  
Inverse Correlation ◽  
Host Cells ◽  
Target Cells ◽  
Metacyclic Trypomastigotes ◽  
Cell Components ◽  
Major Surface ◽  
Surface Glycoproteins

Mammalian cell invasion assays, using metacyclic trypomastigotes of Trypanosoma cruzi G and CL strains, showed that the CL strain enters target cells in several-fold higher numbers as compared with the G strain. Analysis of expression of surface glycoproteins in metacyclic forms of the two strains by iodination, immunoprecipitation and FACS, revealed that gp90, undetectable in the CL strain, is one of the major surface molecules in the G strain, that expression of gp82 is comparable in both strains and that gp35/50 is expressed at lower levels in the CL strain. Purified gp90 and gp35/50 bound more efficiently than gp82 to cultured HeLa cells. However, the intensity of the Ca2+ response triggered in HeLa cells by gp82 was significantly higher than that induced by gp35/50 or gp90. Most of the Ca2+ signalling activity of the metacyclic extract towards HeLa cells was due to gp82 and was inhibitable by gp82-specific monoclonal antibody 3F6. Ca2+ mobilization was also triggered in metacyclic trypomastigotes by host-cell components; it was mainly gp82-mediated and more intense in the CL than in the G strain. We propose that expression of gp90 and gp35/50 at high levels impairs binding of metacyclic forms to host cells through productive gp82-mediated interaction, which leads to the target-cell and parasite Ca2+ mobilization required for invasion. Analysis of metacyclic forms of eight additional T. cruzi strains corroborated the inverse correlation between infectivity and expression of gp90 and gp35/50.

scholarly journals Developmentally regulated, phospholipase C-mediated release of the major surface glycoprotein of amastigotes of Trypanosoma cruzi.

1988 ◽  
Vol 167 (2) ◽  
pp. 300-314 ◽  
Author(s):  
N W Andrews ◽  
E S Robbins ◽  
V Ley ◽  
K S Hong ◽  
V Nussenzweig
Keyword(s):  
Trypanosoma Cruzi ◽  
Phospholipase C ◽  
Myristic Acid ◽  
Surface Glycoprotein ◽  
Developmentally Regulated ◽  
African Trypanosomes ◽  
Glycosyl Phosphatidylinositol ◽  
Major Surface Glycoprotein ◽  
Major Stage ◽  
Major Surface

The surface of amastigotes of Trypanosoma cruzi is covered by Ssp-4, a major stage-specific glycoprotein. Ssp-4 is anchored to the cell membrane by GPI. It can be metabolically labeled with [3H]myristic acid, and is converted into a hydrophilic form by treatment with the glycan-specific phospholipase C of T. brucei, or after lysis of the parasites in non-ionic detergents. The hydrophilic form of Ssp-4 is recognized by antibodies to the cross-reactive determinant of the variant surface glycoprotein of African trypanosomes. Ssp-4 is progressively shed during the intra- or extracellular development of amastigotes preceding their transformation into epi- and trypomastigotes. We show here that T. cruzi contains a phospholipase C and that most shed Ssp-4 is hydrophilic, does not contain myristic acid, and reacts with anti-CRD. These observations provide strong evidence that phospholipase C mediates the release of this glycosyl-phosphatidylinositol-anchored protein under physiological conditions, as the parasite undergoes differentiation.

scholarly journals Fibronectin-Degrading Activity of Trypanosoma cruzi Cysteine Proteinase Plays a Role in Host Cell Invasion

2014 ◽  
Vol 82 (12) ◽  
pp. 5166-5174 ◽  
Author(s):  
Fernando Yukio Maeda ◽  
Cristian Cortez ◽  
Mario Augusto Izidoro ◽  
Luiz Juliano ◽  
Nobuko Yoshida
Keyword(s):  
Trypanosoma Cruzi ◽  
Host Cell ◽  
Cell Invasion ◽  
Hela Cells ◽  
Transforming Growth Factor ◽  
Cysteine Proteinase ◽  
Mammalian Host ◽  
Cysteine Proteinase Inhibitor ◽  
Host Cell Invasion ◽  
Content Type

ABSTRACTTrypanosoma cruzi, the agent of Chagas disease, binds to diverse extracellular matrix proteins. Such an ability prevails in the parasite forms that circulate in the bloodstream and contributes to host cell invasion. Whether this also applies to the insect-stage metacyclic trypomastigotes, the developmental forms that initiate infection in the mammalian host, is not clear. UsingT. cruziCL strain metacyclic forms, we investigated whether fibronectin bound to the parasites and affected target cell invasion. Fibronectin present in cell culture medium bound to metacyclic forms and was digested by cruzipain, the majorT. cruzicysteine proteinase. G strain, with negligible cruzipain activity, displayed a minimal fibronectin-degrading effect. Binding to fibronectin was mediated by gp82, the metacyclic stage-specific surface molecule implicated in parasite internalization. When exogenous fibronectin was present at concentrations higher than cruzipain can properly digest, or fibronectin expression was stimulated by treatment of epithelial HeLa cells with transforming growth factor beta, the parasite invasion was reduced. Treatment of HeLa cells with purified recombinant cruzipain increased parasite internalization, whereas the treatment of parasites with cysteine proteinase inhibitor had the opposite effect. Metacyclic trypomastigote entry into HeLa cells was not affected by anti-β1 integrin antibody but was inhibited by anti-fibronectin antibody. Overall, our results have indicated that the cysteine proteinase ofT. cruzimetacyclic forms, through its fibronectin-degrading activity, is implicated in host cell invasion.

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