Document Detail

Proteolytic activation of internalized cholera toxin within hepatic endosomes by cathepsin D.
MedLine Citation:
PMID:  16128808     Owner:  NLM     Status:  MEDLINE    
We have defined the in vivo and in vitro metabolic fate of internalized cholera toxin (CT) in the endosomal apparatus of rat liver. In vivo, CT was internalized and accumulated in endosomes where it underwent degradation in a pH-dependent manner. In vitro proteolysis of CT using an endosomal lysate required an acidic pH and was sensitive to pepstatin A, an inhibitor of aspartic acid proteases. By nondenaturating immunoprecipitation, the acidic CT-degrading activity was attributed to the luminal form of endosomal cathepsin D. The rate of toxin hydrolysis using an endosomal lysate or pure cathepsin D was found to be high for native CT and free CT-B subunit, and low for free CT-A subunit. On the basis of IC(50) values, competition studies revealed that CT-A and CT-B subunits share a common binding site on the cathepsin D enzyme, with native CT and free CT-B subunit displaying the highest affinity for the protease. By immunofluorescence, partial colocalization of internalized CT with cathepsin D was confirmed at early times of endocytosis in both hepatoma HepG2 and intestinal Caco-2 cells. Hydrolysates of CT generated at low pH by bovine cathepsin D displayed ADP-ribosyltransferase activity towards exogenous Gsalpha protein suggesting that CT cytotoxicity, at least in part, may be related to proteolytic events within endocytic vesicles. Together, these data identify the endocytic apparatus as a critical subcellular site for the accumulation and proteolytic degradation of endocytosed CT, and define endosomal cathepsin D an enzyme potentially responsible for CT cytotoxic activation.
Clémence Merlen; Domitille Fayol-Messaoudi; Sylvie Fabrega; Tatiana El Hage; Alain Servin; François Authier
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Publication Detail:
Type:  In Vitro; Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  The FEBS journal     Volume:  272     ISSN:  1742-464X     ISO Abbreviation:  FEBS J.     Publication Date:  2005 Sep 
Date Detail:
Created Date:  2005-08-30     Completed Date:  2005-10-13     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  101229646     Medline TA:  FEBS J     Country:  England    
Other Details:
Languages:  eng     Pagination:  4385-97     Citation Subset:  IM    
Institut National de la Santé et de la Recherche Médicale U510, Faculté de Pharmacie Paris XI, Châtenay-Malabry, France.
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MeSH Terms
Biodegradation, Environmental
Caco-2 Cells
Cathepsin D / metabolism*
Cell Line
Cholera Toxin / chemistry,  metabolism*
Endosomes / metabolism*
Hydrogen-Ion Concentration
Liver / metabolism*
Protein Subunits
Rats, Sprague-Dawley
Reg. No./Substance:
0/Protein Subunits; 9012-63-9/Cholera Toxin; EC D

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