Effect of helicobacter pylori vacuolating toxin on maturation and extracellular release of procathepsin D and on epidermal growth factor degradation.
The effect of vacuolating toxin (VacA) from Helicobacter pylori on endosomal and lysosomal functions was studied by following procathepsin D maturation and epidermal growth factor (EGF) degradation in HeLa cells exposed to the toxin. VacA inhibited the conversion of procathepsin D (53 kDa) into both the intermediate (47 kDa) and the mature (31 kDa) form. Nonprocessed cathepsin D was partly retained inside cells and partly secreted in the extracellular medium via the constitutive secretion pathway. Intracellular degradation of EGF was also inhibited by VacA with a similar dose-response curve. VacA did not alter endocytosis, cell surface recycling, and retrograde transport from plasma membrane to trans-Golgi network and endoplasmic reticulum, as estimated by using transferrin, diphtheria toxin, and ricin as tracers. Subcellular fractionation of intoxicated cells showed that procathepsin D and nondegraded EGF accumulate in lysosomes. Measurements of intracellular acidification with fluorescein isothiocyanate-dextran revealed a partial neutralization of the lumen of endosomes and lysosomes, sufficient to account for both mistargeting of procathepsin D outside the cell and the decreased activity of lysosomal proteases.
Satin
B
B
Centro CNR Biomembrane and Dipartimento di Scienze Biomediche, Universita' di Padova, Via G. Colombo 3, 35121 Padova, Italy.
Norais
N
N
Telford
J
J
Rappuoli
R
R
Murgia
M
M
Montecucco
C
C
Papini
E
E
D001426:Bacterial Proteins / Q000633:toxicity*; D001427:Bacterial Toxins / Q000633:toxicity; D002402:Cathepsin D / Q000096:biosynthesis / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D004167:Diphtheria Toxin / Q000493:pharmacokinetics; D004705:Endocytosis / Q000187:drug effects; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism; D004792:Enzyme Precursors / Q000378:metabolism*; D004815:Epidermal Growth Factor / Q000378:metabolism*; D006056:Golgi Apparatus / Q000187:drug effects / Q000378:metabolism; D006367:HeLa Cells; D016480:Helicobacter pylori; D006801:Humans; D007700:Kinetics; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects*; D012276:Ricin / Q000493:pharmacokinetics; D014168:Transferrin / Q000378:metabolism
D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004167:Diphtheria Toxin; D004792:Enzyme Precursors; D014168:Transferrin; C074689:VacA protein, Helicobacter pylori; D004815:Epidermal Growth Factor; D012276:Ricin; C058345:procathepsin D; D002402:Cathepsin D