analysis-epithelial-damage-produced-entamoeba-histolytica

Analysis of the Epithelial Damage Produced by <em>Entamoeba histolytica</em> Infection

Entamoeba histolytica is the causative agent of human amoebiasis, a major cause of diarrhea and hepatic abscess in tropical countries. Infection is ...

Center for Research and Advanced Studies of the National Polytechnic Institute

The presence in Entamoeba histolytica of a fibronectin (FN) receptor, which is antigenically related to beta1 integrin-like molecules and shows 99% homology with the intermediate subunit-2 of the Gal/GalNAc-specific lectin has been described. The E. histolytica genome has been sequenced, and its analysis shows no integrin sequences. Here we provide further evidence to demonstrate that this molecule behaves as integrin-like in its physical, structural and functional properties. The purified beta1EhFNR complex is resolved into three polypeptides of 150, 140, and 130 kDa. Transmission electron microscopy showed individual complexes consisting of oblong heads of 3 nm x 4 nm and two projecting arms 6-7 nm in length. In the absence of detergent, these complexes formed aggregates that were composed of clusters or "rosettes" of between two and six or more beta1EhFNR complexes. The physical properties of the purified beta1EhFNR complexes were: R(S)=5.8 nm, S(20)W=8.3, f/f(0)=1.4. This complex was seen in close physical association with adhesion plates and phagocytic invaginations, using confocal microscopy and the 3C10 mAb that recognizes these three subunits complex. Regulation of its surface expression is not dependent on protein synthesis; rather it is regulated by inward and outward mobilization of the molecules. The presence and antigenic similarity of putative beta1EhFNRs in different strains and species of Entamoeba was analyzed using the 3C10 mAb; this mAb recognized the complex in all E. histolytica species, however there was no recognition in E. dispar, E. invadens, and Laredo strains. Finally, evidence is provided about post-translational modifications such as tyrosine phosphorylation and glycosylation suffered by the beta1EhFNR complex.

Physical, structural, and functional properties of the beta1 integrin-like fibronectin receptor (beta1EhFNR) in Entamoeba histolytica.

Entamoeba histolytica, the protozoan responsible for human amoebiasis, causes between 30,000 and 100,000 deaths per year worldwide. Amoebiasis is characterized by intestinal epithelial damage provoking severe diarrhea. However, the molecular mechanisms by which this protozoan causes epithelial damage are poorly understood. Here, we studied the initial molecular interactions between the E. histolytica EhCPADH112 virulence complex and epithelial MDCK and Caco-2 cells. By confocal microscopy, we discovered that after contact with trophozoites or trophozoite extracts (TE), EhCPADH112 and proteins forming this complex (EhCP112 and EhADH112) co-localize with occludin and claudin-1 at tight junctions (TJ). Immunoprecipitation assays revealed interaction between EhCPADH112 and occludin, claudin-1, ZO-1 and ZO-2. Overlay assays confirmed an interaction of EhCP112 and EhADH112 with occludin and claudin-1, whereas only EhADH112 interacted also with ZO-2. We observed degradation of all mentioned TJ proteins after incubation with TE. Importantly, inhibiting proteolytic activity or blocking the complex with a specific antibody not only prevented TJ protein degradation but also epithelial barrier disruption. Furthermore, we discovered that TE treatment induces autophagy and apoptosis in MDCK cells that could contribute to the observed barrier disruption. Our results suggest a model in which epithelial damage caused by E. histolytica is initiated by the interaction of EhCP112 and EhADH112 with TJ proteins followed by their degradation. Disruption of TJs then induces increased paracellular permeability, thus facilitating the entry of more proteases and other parasite molecules leading eventually to tissue destruction.

Article	Year
Physical, structural, and functional properties of the beta1 integrin-like fibronectin receptor (beta1EhFNR) in Entamoeba histolytica. Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases\| PubMed ID: 19577014	2009
The EhCPADH112 complex of Entamoeba histolytica interacts with tight junction proteins occludin and claudin-1 to produce epithelial damage. PloS one\| PubMed ID: 23762290	2013

Rosario Javier-Reyna

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