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) fromHelicobacter 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 totrans-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.

[1]  P. Boquet,et al.  Membrane translocation of diphtheria toxin fragment A exploits early to late endosome trafficking machinery , 1997, Molecular microbiology.

[2]  R. Rappuoli,et al.  The small GTP binding protein rab7 is essential for cellular vacuolation induced by Helicobacter pylori cytotoxin , 1997, The EMBO journal.

[3]  P. Auvinen,et al.  Endosome dynamics regulated by a Rho protein , 1996, Nature.

[4]  T. Cover,et al.  Binding and internalization of the Helicobacter pylori vacuolating cytotoxin by epithelial cells , 1996, Infection and immunity.

[5]  R. Rappuoli,et al.  Oligomeric and subunit structure of the Helicobacter pylori vacuolating cytotoxin , 1996, The Journal of cell biology.

[6]  C. Futter,et al.  Multivesicular endosomes containing internalized EGF-EGF receptor complexes mature and then fuse directly with lysosomes , 1996, The Journal of cell biology.

[7]  N. Beeching,et al.  Tuberculosis into the next century: Proceedings of a symposium held on 4 February 1995 at the Liverpool School of Tropical Medicine , 1996 .

[8]  R. Rappuoli,et al.  Lipid interaction of the 37-kDa and 58-kDa fragments of the Helicobacter pylori cytotoxin. , 1995, European journal of biochemistry.

[9]  R. Rappuoli,et al.  Helicobacter pylori cytotoxin: importance of native conformation for induction of neutralizing antibodies , 1995, Infection and immunity.

[10]  R. Rappuoli,et al.  Low pH Activates the Vacuolating Toxin of Helicobacter pylori, Which Becomes Acid and Pepsin Resistant (*) , 1995, The Journal of Biological Chemistry.

[11]  M. Blaser,et al.  Role of the Helicobacter pylori virulence factors vacuolating cytotoxin, CagA, and urease in a mouse model of disease , 1995, Infection and immunity.

[12]  W. Balch,et al.  Ricin Cytotoxicity Is Sensitive to Recycling between the Endoplasmic Reticulum and the Golgi Complex (*) , 1995, The Journal of Biological Chemistry.

[13]  S. Emr,et al.  Role for phosphatidylinositol 3-kinase in the sorting and transport of newly synthesized lysosomal enzymes in mammalian cells , 1995, The Journal of cell biology.

[14]  H. Davidson,et al.  Wortmannin causes mistargeting of procathepsin D. evidence for the involvement of a phosphatidylinositol 3-kinase in vesicular transport to lysosomes , 1995, The Journal of cell biology.

[15]  R. Rappuoli,et al.  Development of a mouse model of Helicobacter pylori infection that mimics human disease , 1995, Science.

[16]  S. Falkow,et al.  The new path to preventing ulcers , 1995, Science.

[17]  R. Rappuoli,et al.  Cellular vacuoles induced by Helicobacter pylori originate from late endosomal compartments. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[18]  R. Rappuoli,et al.  Unravelling the pathogenic role of Helicobacter pylori in peptic ulcer: potential new therapies and vaccines. , 1994, Trends in biotechnology.

[19]  K. von Figura,et al.  Proteolytic processing of cathepsin D in prelysosomal organelles. , 1994, European journal of cell biology.

[20]  R. Warnke,et al.  Helicobacter pylori infection and gastric lymphoma. , 1994, The New England journal of medicine.

[21]  S. Normark,et al.  Pathological significance and molecular characterization of the vacuolating toxin gene of Helicobacter pylori , 1994, Infection and immunity.

[22]  S. Pfeffer,et al.  Lysosome biogenesis requires Rab9 function and receptor recycling from endosomes to the trans-Golgi network , 1994, The Journal of cell biology.

[23]  M. Comanducci,et al.  Gene structure of the Helicobacter pylori cytotoxin and evidence of its key role in gastric disease , 1994, The Journal of experimental medicine.

[24]  M. Blaser,et al.  Divergence of genetic sequences for the vacuolating cytotoxin among Helicobacter pylori strains. , 1994, The Journal of biological chemistry.

[25]  R. Haas,et al.  Genetic analysis of the Helicobacter pylori vacuoiating cytotoxin: structural similarities with the IgA protease type of exported protein , 1994, Molecular microbiology.

[26]  E. Papini,et al.  Cell vacuolization induced by Helicobacter pylori: inhibition by bafilomycins A1, B1, C1 and D. , 1993, FEMS microbiology letters.

[27]  M. Blaser.  Helicobacter pylori: microbiology of a 'slow' bacterial infection. , 1993, Trends in microbiology.

[28]  D. Forman,et al.  An international association between Helicobacter pylori infection and gastric cancer , 1993 .

[29]  M. Blaser,et al.  Effects of ATPase inhibitors on the response of HeLa cells to Helicobacter pylori vacuolating toxin , 1993, Infection and immunity.

[30]  L. B. Larsen,et al.  Procathepsin D cannot autoactivate to cathepsin D at acid pH , 1993, FEBS letters.

[31]  M. Murgia,et al.  Cell penetration of diphtheria toxin. Reduction of the interchain disulfide bridge is the rate-limiting step of translocation in the cytosol. , 1993, The Journal of biological chemistry.

[32]  E. Papini,et al.  Bafilomycin A1 inhibits Helicobacter pylori‐induced vacuolization of HeLa cells , 1993, Molecular microbiology.

[33]  C. Catrenich,et al.  Character and origin of vacuoles induced in mammalian cells by the cytotoxin of Helicobacter pylori. , 1992, Journal of medical microbiology.

[34]  H. Geuze,et al.  Identification of subcellular compartments involved in biosynthetic processing of cathepsin D. , 1992, The Journal of biological chemistry.

[35]  M. Blaser,et al.  Purification and characterization of the vacuolating toxin from Helicobacter pylori. , 1992, The Journal of biological chemistry.

[36]  A. Hubbard,et al.  Sequential processing of epidermal growth factor in early and late endosomes of rat liver. , 1991, The Journal of biological chemistry.

[37]  G. Conner,et al.  Proteolytic activation of human procathepsin D. , 1991, Advances in experimental medicine and biology.

[38]  M. Zerial,et al.  Localization of low molecular weight GTP binding proteins to exocytic and endocytic compartments , 1990, Cell.

[39]  D. Cutler,et al.  Sorting during transport to the surface of PC12 cells: divergence of synaptic vesicle and secretory granule proteins , 1990, The Journal of cell biology.

[40]  D. Davies,et al.  Structure and Function of the Aspartic Proteinases , 1990, Advances in Experimental Medicine and Biology.

[41]  J. Slot,et al.  Sorting of mannose 6-phosphate receptors and lysosomal membrane proteins in endocytic vesicles , 1988, The Journal of cell biology.

[42]  Y. Goda,et al.  Selective recycling of the mannose 6-phosphate/IGF-II receptor to the trans Golgi network in vitro , 1988, Cell.

[43]  D. Morgan,et al.  Cytotoxic activity in broth-culture filtrates of Campylobacter pylori. , 1988, Journal of medical microbiology.

[44]  I. Mellman,et al.  The mannose 6-phosphate receptor and the biogenesis of lysosomes , 1988, Cell.

[45]  O. Petersen,et al.  Estimation of the amount of internalized ricin that reaches the trans- Golgi network , 1988, The Journal of cell biology.

[46]  R. Collier,et al.  [11] Diphtheria toxin: Purification and properties , 1988 .

[47]  D. Neville,et al.  Temporal separation of protein toxin translocation from processing events. , 1987, The Journal of biological chemistry.

[48]  J. Goodhouse,et al.  Mannose-6-phosphate receptors for lysosomal enzymes cycle between the Golgi complex and endosomes , 1986, The Journal of cell biology.

[49]  K. von Figura,et al.  Lysosomal enzymes and their receptors. , 1986, Annual review of biochemistry.

[50]  B. Goud,et al.  Inhibition of coated pit formation in Hep2 cells blocks the cytotoxicity of diphtheria toxin but not that of ricin toxin , 1985, The Journal of cell biology.

[51]  B. Marshall,et al.  Attempt to fulfil Koch's postulates for pyloric Campylobacter , 1985, The Medical journal of Australia.

[52]  C. Hopkins,et al.  Internalization and processing of transferrin and the transferrin receptor in human carcinoma A431 cells , 1983, The Journal of cell biology.

[53]  K. von Figura,et al.  Biosynthesis and transport of cathepsin D in cultured human fibroblasts , 1983, The Journal of cell biology.

[54]  R. Rappuoli,et al.  Rapid purification of diphtheria toxin by phenyl sepharose and DEAE-cellulose chromatography , 1983 .

[55]  F. Maxfield.  Weak bases and ionophores rapidly and reversibly raise the pH of endocytic vesicles in cultured mouse fibroblasts , 1982, The Journal of cell biology.

[56]  A. Hasilik,et al.  Biosynthesis of lysosomal enzymes in fibroblasts. Synthesis as precursors of higher molecular weight. , 1980, The Journal of biological chemistry.

[57]  P. Fraker,et al.  Protein and cell membrane iodinations with a sparingly soluble chloroamide, 1,3,4,6-tetrachloro-3a,6a-diphrenylglycoluril. , 1978, Biochemical and biophysical research communications.

[58]  E. Feytmans,et al.  ANALYTICAL STUDY OF MICROSOMES AND ISOLATED SUBCELLULAR MEMBRANES FROM RAT LIVER , 1974, The Journal of cell biology.

[59]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.