title: Proteolysis of enteric cell villin by Entamoeba histolytica cysteine proteinases.
abstract: Invasive microorganisms efface enteric microvilli to establish intimate contact with the apical surface of enterocytes. To understand the molecular basis of this effacement in amebic colitis, we seeded Entamoeba histolytica trophozoites on top of differentiated human Caco-2 cell layers. Western blots of detergent lysates from such cocultures showed proteolysis of the actin-bundling protein villin within 1 min of direct contact of living trophozoites with enterocytes. Mixtures of separately prepared lysates excluded detergent colysis as the cause of villin proteolysis. Caspases were not responsible as evidenced by the lack of degradation of specific substrates and the failure of a specific caspase inhibitor to prevent villin proteolysis. A crucial role for amebic cysteine proteinases was shown by prevention of villin proteolysis and associated microvillar alterations through the treatment of trophozoites before coculture with synthetic inhibitors that completely blocked amebic cysteine proteinase activity on zymograms. Moreover, trophozoites of amebic strains pSA8 and SAW760 with strongly reduced cysteine proteinase activity showed a reduced proteolysis of villin in coculture with enteric cells. Salmonella typhimurium and enteropathogenic Escherichia coli disturb microvilli without villin proteolysis, indicating that the latter is not a consequence of the disturbance of microvilli. In conclusion, villin proteolysis is an early event in the molecular cross-talk between enterocytes and amebic trophozoites, causing a disturbance of microvilli.
authors.affiliation: Laboratory of Experimental Cancerology, Ghent University Hospital, De Pintelaan 185, B-9000 Ghent, Belgium.
authors.forename: Tineke
authors.initials: T
authors.lastname: Lauwaet
authors.forename: Maria José
authors.initials: MJ
authors.lastname: Oliveira
authors.forename: Bert
authors.initials: B
authors.lastname: Callewaert
authors.forename: Georges
authors.initials: G
authors.lastname: De Bruyne
authors.forename: Xavier
authors.initials: X
authors.lastname: Saelens
authors.forename: Serge
authors.initials: S
authors.lastname: Ankri
authors.forename: Peter
authors.initials: P
authors.lastname: Vandenabeele
authors.forename: David
authors.initials: D
authors.lastname: Mirelman
authors.forename: Marc
authors.initials: M
authors.lastname: Mareel
authors.forename: Ancy
authors.initials: A
authors.lastname: Leroy
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D002448:Cell Adhesion; D018920:Coculture Techniques; D015179:Colorectal Neoplasms; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D004748:Entamoeba histolytica / Q000201:enzymology* / Q000502:physiology; D006801:Humans; D007413:Intestinal Mucosa / Q000469:parasitology*; D008840:Microfilament Proteins / Q000378:metabolism*; D008871:Microvilli / Q000469:parasitology*; D014407:Tumor Cells, Cultured; D014508:Urea / Q000378:metabolism
chemical_list: D002352:Carrier Proteins; D008840:Microfilament Proteins; C026615:villin; D014508:Urea; D003546:Cysteine Endopeptidases