Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.23.5 (cathepsin D)
 4,130 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Macrophages actively internalize macromolecules into endosomal vesicles containing proteases. The plant toxin, ricin A chain delivered into this pathway by receptor-mediated endocytosis, was found to be exquisitely sensitive to cleavage by these proteases. Proteolytic fragments of ricin A chain were generated within cells as early as 2-3 min after internalization. Toxin proteolysis was initiated in early endosomal vesicles, and transport to lysosomes was not required. As endosomes transit the cell, their lumenal pH drops from neutral to acidic. Previous studies in macrophages had suggested that endosomal proteolysis is dependent on vesicle acidification. Isolated endosomal vesicles containing ricin A chain catalyzed the cleavage of this protein in vitro; however, proteolysis was observed at both neutral and acidic pH. Experiments using isolated endosomes demonstrated that both cysteine and aspartyl proteases were responsible for the cleavage of ricin A chain. The cysteine protease, cathepsin B, catalyzed toxin proteolysis in endosomes between pH 4.5 and 7.0 while aspartyl protease activity was maximal below pH 5.5. Radiolabeling the lumenal contents of macrophage endosomes confirmed that both the cysteine protease, cathepsin B, and the aspartyl protease, cathepsin D, were present in these vesicles. These proteases were not present on the plasma membrane but were found in early endosomes indicating they are derived from an intracellular source. The presence of proteases with different pH optima in early endosomes suggests that processing in these vesicles may be regulated by changes in endosomal pH. This result represents an important difference in protein processing in endosomes versus lysosomes and provides new insights into the function of endosomal proteases.
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PMID:Proteolytic cleavage of ricin A chain in endosomal vesicles. Evidence for the action of endosomal proteases at both neutral and acidic pH. 193 30

Chloroquine-resistant (CQr) clones (CQ-21 and CQ-22) have been isolated from mutagenized hamster lung V79 cells by exposing the cells to a high dose of chloroquine. CQ-21 and CQ-22 showed about 3-fold higher resistance to chloroquine than the parental V79 cells, and they showed specific cross-resistance to another amine, NH4Cl, which is also concentrated in lysosomes. CQr clone showed no cross-resistance to other unrelated agents. Chloroquine-induced inhibition of [125I]ricin internalization was observed in both cell lines at neutral pH, but the inhibition of uptake was less in the variant. Also, the degradation of endogenous protein was slowed in the mutant; further, treatment of cells with 30 micrograms/ml of chloroquine inhibited the degradation of endogenous proteins in the parental V79, but not in CQ-22 cells. Similar levels of acid phosphatase, beta-glucuronidase and cathepsin D were observed in V79 and CQ-22 cells, but the level of cathepsin B was lower in the mutant. Electron microscopy showed an increased number of electron-dense bodies, possibly autophagosomes/lysosomes, in the mutant cells grown for 4 days with 5 micrograms/ml of chloroquine. Similar aberrant structures were observed in the parental V79 cells treated for only 3 h with 5 micrograms/ml of chloroquine.
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PMID:Isolation of chloroquine-resistant Chinese hamster V79 cell variants that are also resistant to ammonium chloride. 665 68

Ricin A-chain is delivered into macrophages via receptor-mediated endocytosis. We have found that following uptake via the mannose receptor, ricin A-chain is rapidly cleaved by endosomal proteases. Inhibition of endosomal proteases such as cathepsin D and B leads to the accumulation of toxin inside the cell. Inhibition of cathepsin D reduces ricin A-chain cytotoxicity, while blocking cathepsin B enhances cytotoxicity. Similar results were obtained using fibroblasts transfected with the mannose receptor. Our data strongly suggest that the activation or membrane translocation of ricin A-chain is dependent upon the action of specific proteases.
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PMID:Endosomal proteolysis precedes ricin A-chain toxicity in macrophages. 827 7

In the human carcinoma cell line HEp-2, endosomes are multivesicular bodies (MVBs) which gradually mature and eventually fuse with other mature endosomes and/or with preexisting lysosomes. Selective inhibition of the vacuolar H(+)-ATPase with bafilomycin A1 (Baf) did not influence endocytic uptake and recycling of the protein toxin ricin. Moreover, quantification of immunogold labeling on ultracryosections revealed that the frequency by which MVBs containing internalized cationized gold (Cat.Au) also labeled for the cation-independent mannose-phosphate receptor (MPR) was the same with and without Baf, suggesting that formation and maturation of MVBs were unchanged in the presence of Baf. However, degradation of ricin was strongly reduced by bafilomycin, and this reduction was not only due to increased pH in lysosomes. Thus, quantitative lmmunogold labeling showed that the Baf-induced alkalinization reduced the transfer of internalized Cat.Au to lysosomes, as defined by their content of human lysosome-associated membrane protein 1 (h-lamp-1) and cathepsin D. Accordingly, although low vacuolar pH does not seem to be required for transport to MPR-containing endosomes, it seems to be important for a late fusion step along the endocytic pathway.
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PMID:Inhibition of the vacuolar H(+)-ATPase with bafilomycin reduces delivery of internalized molecules from mature multivesicular endosomes to lysosomes in HEp-2 cells. 874 Dec 16

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.
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PMID:Effect of helicobacter pylori vacuolating toxin on maturation and extracellular release of procathepsin D and on epidermal growth factor degradation. 931 9