Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.23.5 (cathepsin D)
4,130 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Phagocytic processing of heat-killed Listeria monocytogenes by peritoneal macrophages resulted in degradation of these bacteria in phagolysosomal compartments and processing of bacterial antigens for presentation to T cells by class II MHC molecules. Within 20 min of uptake by macrophages, Listeria peptide antigens were expressed on surface class II MHC molecules, capable of stimulating Listeria-specific T cells. Within this period, degradation of labeled bacteria to acid-soluble low molecular weight catabolites also commenced. Immunoelectron microscopy was used to evaluate the compartments involved in this processing. Upon uptake of the bacteria, phagosomes containing Listeria fused rapidly with both lysosomes and endosomes. Class II MHC molecules were present in a tubulo-vesicular lysosome compartment, which appeared to fuse with phagosomes, as well as in the resulting phagolysosomes containing internalized Listeria; these compartments were all positive for Lamp 1 and cathepsin D and lacked 46-kD mannose-6-phosphate receptors. In addition, class II MHC and Lamp 1 were co-localized in vesicles of the trans Golgi reticulum, where they were segregated from 46-kD mannose-6-phosphate receptors. Vesicles containing both Listeria-derived components and class II MHC molecules were also observed; some of these may represent vesicles recycling from phagolysosomes, potentially bearing processed immunogenic peptides complexed with class II MHC. These results support a central role for lysosomes and phagolysosomes in the processing of bacterial antigens for presentation to T cells. Tubulo-vesicular lysosomes appear to represent an important convergence of endocytic, phagocytic and biosynthetic pathways, where antigens may be processed to allow binding to class II MHC molecules and recycling to the cell surface.
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PMID:Class II MHC molecules are present in macrophage lysosomes and phagolysosomes that function in the phagocytic processing of Listeria monocytogenes for presentation to T cells. 140 May 90

To determine the cause of the increased content of carbohydrate-bound phosphate in tumour lysosomal hydrolases, the activity and kinetics in human hepatocellular carcinoma of two enzymes involved in the formation of mannose-6-phosphate in lysosomal hydrolases UDP-GlcNAc: lysosomal enzyme GlcNAc alpha l-phosphotransferase (GlcNAc-phosphotransferase) and phosphodiester glycosidase were studied. The activity level of the phosphotransferase with artificial and natural substrates was elevated (P less than 0.025 and P less than 0.001, respectively) in hepatoma compared to that in uninvolved tissue, while the phosphodiester glycosidase of hepatoma was at a level similar to that of the uninvolved tissue. To verify a previous observation that cathepsin D of human hepatoma contained increased GlcNAc-phosphomannose, the protease was examined for carbohydrate phosphorylation by the GlcNAc-phosphotransferase. The protease from normal human liver was much more phosphorylated than hepatoma protease, confirming the previous observation. The predominant phosphorylation of the protease occurred in one of two major heavy subunits, with some phosphorylation in one of two minor light subunits.
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PMID:Elevated carbohydrate phosphotransferase activity in human hepatoma and phosphorylation of cathepsin D. 164 48

We have investigated the distribution of newly synthesized lysosomal enzymes in endocytic compartments of normal rat kidney (NRK) cells. The mannose-6-phosphate (Man6-P) containing lysosomal enzymes could be iodinated in situ after internalization of lactoperoxidase (LPO) by fluid phase endocytosis and isolated on CI-MPR affinity columns. For EM studies, the ectodomain of the CI-MPR conjugated to colloidal gold was used as a probe specific for the phosphomannosyl marker of the newly synthesized hydrolases. In NRK cells, approximately 20-40% of the phosphorylated hydrolases present in the entire pathway were found in early endocytic structures proximal to the 18 degrees C temperature block including early endosomes. These structures were characterized by a low content of endogenous CI-MPR and were accessible to fluid phase markers internalized for 5-15 min at 37 degrees C. The bulk of the phosphorylated lysosomal enzymes was found in late endocytic structures distal to the 18 degrees C block, rich in endogenous CI-MPR and accessible to endocytic markers internalized for 30-60 min at 37 degrees C. The CI-MPR negative lysosomes were devoid of phosphorylated hydrolases. This distribution was unchanged in cells treated with Man6-P to block recapture of secreted lysosomal enzymes. However, lysosomal enzymes were no longer detected in the early endosomal elements of cells treated with cycloheximide. Immunoprecipitation of cathepsin D from early endosomes of pulse-labeled cells showed that this hydrolase is a transient component of this compartment. These data indicate that in NRK cells, the earliest point of convergence of the lysosomal biosynthetic and the endocytic pathways is the early endosome.
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PMID:Distribution of newly synthesized lysosomal enzymes in the endocytic pathway of normal rat kidney cells. 166 30

Adherent cultures of rat peritoneal macrophages secrete lysozyme and the lysosomal marker enzymes beta-glucuronidase, beta-N-acetylglucosaminidase and acid phosphatase; the levels of secreted lysosomal cathepsin D, however, were found to be insignificant. Incubation of the cells at 4 degrees C for 15 min with yeast mannan or with 50 mM mannose, methyl alpha-glucopyranoside, or N-acetylglucosamine caused the concentration of cathepsin D in the culture medium to increase 30-40-fold; mannose-6-phosphate had no effect. 125I-labeled cathepsin D was prepared and the binding constant to the macrophage cell surface was determined to be KD = 27 nM. The data suggest that cathepsin D binds to the mannose receptor of macrophages and that binding to this receptor is not in equilibrium with the bulk medium.
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PMID:Binding of cathepsin D to the mannose receptor on rat peritoneal macrophages. 193 26

Intracisternal granules (ICGs) are insoluble aggregates of pancreatic digestive enzymes and proenzymes that develop within the lumen of the rough endoplasmic reticulum of exocrine pancreatic cells, especially in guinea pigs. These ICGs are eliminated by autophagy. By morphological criteria, we identified three distinct and sequential classes of autophagic compartments, which we refer to as phagophores, Type I autophagic vacuoles, and Type II autophagic vacuoles. Lobules of guinea pig pancreas were incubated in media containing HRP for periods of 5-120 min to determine the relationship between the endocytic and autophagic pathways. Incubations with HRP of 15 min or less labeled early endosomes at the cell periphery that were not involved in autophagy of ICGs, but after these short incubations none of the autophagic compartments were HRP positive. After 30-min incubation with HRP, early endosomes at the cell periphery, late endosomes in the pericentriolar region, and, in addition, Type I autophagic vacuoles containing ICGs were all labeled by the tracer. Type II autophagic vacuoles were not labeled after 30-min incubation with HRP but were labeled after incubations of 60-120 min. Phagophores did not receive HRP even after 120 min incubations. We concluded that the autophagic and endocytic pathways converge immediately after the early endosome level and that Type I autophagic vacuoles precede Type II autophagic vacuoles on the endocytic pathway. We studied the distribution of acid phosphatase, lysosomal proteases and cation-independent-mannose-6-phosphate receptor (CI-M6PR) in the three classes of autophagic compartments by histochemical and immunocytochemical methods. Phagophores, the earliest autophagic compartment, contained none of these markers. Type I autophagic vacuoles contained acid phosphatase but, at most, only very low levels of cathepsin D and CI-M6PR. Type II autophagic vacuoles, by contrast, are enriched for acid phosphatase, cathepsin D, and other lysosomal enzymes, and they are also enriched for CI-M6PR. Moreover, soluble fragments of bovine CI-M6PR conjugated to colloidal gold particles heavily labeled Type II but not Type I autophagic vacuoles, and this labeling was specifically blocked by mannose-6-phosphate. This indicates that the lysosomal enzymes present in Type II autophagic vacuoles carry mannose-6-phosphate monoester residues. Using 3-C2, 4-dinitroanilino-3'-amino-N-methyldipropylamine (DAMP), we showed that Type II autophagic vacuoles are acidic. We interpret these findings as indicating that Type II autophagic vacuoles are a prelysosomal compartment in which the already combined endocytic and autophagic pathways meet the delivery pathway of lysosomal enzymes.
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PMID:In exocrine pancreas, the basolateral endocytic pathway converges with the autophagic pathway immediately after the early endosome. 216 50

The uncovering ratio of phosphate groups in lysosomal enzymes is defined as the percentage of phosphomonoester groups in the oligosaccharide side chains based on the sum of phosphomonoester and phosphodiester groups. Using a new procedure for the specific and complete hydrolysis of uncovered phosphomonoester groups in denatured immunoprecipitates of human cathepsin D, we show that the uncovering ratio varies between different forms of the enzyme and may be used as an indicator of the maturation of its carbohydrate side chains. The uncovering ratio in the total (cellular and secreted) cathepsin D from U937 promonocytes is greater than 95%. It is only slightly decreased in cells incubated in the presence of 1 alpha,25-dihydroxycholecalciferol, in which the rate of synthesis of cathepsin D is several times higher than in the control cells. In U937 cells and also in fibroblasts, the uncovering is nearly complete in intermediate and mature forms of the intracellular cathepsin D but less extensive in the intracellular and secreted precursor. In both cell types, incubation with 10 mM NH4Cl results in a decrease in the uncovering ratio of total cathepsin D. However, the activity of the uncovering enzyme, N-acetylglucosamine-1-phosphodiester alpha-N-acetylglucosaminidase, as determined with UDP-N-acetylglucosamine is not affected with up to 60 mM NH4Cl. Our results suggest that NH4Cl, in addition to its known effects on the acidic-pH-dependent functions of lysosomal compartments and of mannose-6-phosphate receptors, impairs the processing or transport of lysosomal enzyme precursors at, or proximally to, the site of the uncovering of their mannose-6-phosphate residues.
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PMID:Suppression of the 'uncovering' of mannose-6-phosphate residues in lysosomal enzymes in the presence of NH4Cl. 216 47

Two enzymes (N-acetylglucosamine-1-phosphotransferase and phosphodiester glycosidase) involved in formation of mannose-6-phosphate at lysosomal hydrolases were studied for the activity and kinetics in human hepatocellular carcinoma. The activity level of the phosphotransferase with an artificial substrate was elevated (p less than 0.025) in hepatoma compared to that in normal liver, while the phosphodiester glycosidase of hepatoma was in a similar level with that of control. The elevation was more remarkable with a physiological substrate, cathepsin D. (P less than 0.001). Since cathepsin D from normal liver was previously demonstrated to contain less phosphomannose compared to the hepatoma protease, the protease was investigated for carbohydrate phosphorylation by the phosphotransferase. The liver protease was much more phosphorylated than the hepatoma protease, endorsing the previous observation. The predominant phosphorylation of the protease occurred in heavy subunit.
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PMID:[Carbohydrate phosphotransferase in human hepatoma and phosphorylation of cathepsin D]. 217 73

The synthesis and secretion of pro-cathepsin D is increased by estrogens in MCF7 cells. We quantified the effect of estradiol on other lysosomal enzymes in order to investigate the mechanism of this hypersecretion. Precursors of beta-hexosaminidase, cathepsin B and beta-galactosidase, which are routed to lysosomes via the mannose-6-phosphate (Man-6-P) receptor, were secreted in much lower amounts than pro-cathepsin D, but their secretion was also increased by estradiol. The activity of acid phosphatase, which is routed to lysosomes via a different transmembrane mechanism, was not altered by estradiol. While estradiol stimulated gene expression of pro-cathepsin D, it had no effect on that of pro-cathepsin B. We conclude that estradiol stimulates the secretion of several lysosomal pro-enzymes in MCF7 cells, suggesting that a general mechanism is responsible for this derouting rather than a specific alteration of cathepsin D structure.
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PMID:Estradiol increases the secretion by MCF7 cells of several lysosomal pro-enzymes. 222 57

A significant elevation of cathepsin D activity was observed in six human hepatoma tissues as compared to 12 normal human livers. In isoelectric focusing experiments, cathepsin D purified from normal liver exhibited three different forms, with isoelectric points of 5.6, 6.1, and 6.7, while cathepsin D purified from hepatoma contained another five to six more acidic forms in addition to the forms observed in normal liver cathepsin D. When the tumor enzyme was treated with endo-beta-N-acetylglucosaminidase H followed by isoelectric focusing, the acidic components disappeared and were converted to forms identical to those of the normal liver cathepsin D. Determination of the mannose-6-phosphate content showed that hepatoma cathepsin D contains twice as much mannose-6-phosphate as normal liver cathepsin D. Peptide mapping and amino acid analysis showed that the protein moiety of cathepsin D from hepatoma is almost identical with that from normal liver. These findings indicate that the appearance of acidic variants in hepatoma cathepsin D is mainly due to changes in the oligosaccharide chains of the enzyme, which are closely associated with the increase of mannose-6-phosphate in the tumor enzyme.
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PMID:Elevated activity and increased mannose-6-phosphate in the carbohydrate moiety of cathepsin D from human hepatoma. 282 73

Cathepsin D was purified to apparently homogeneous form from normal human liver and hepatoma. The purified enzyme could not be distinguished between normal liver and hepatoma in terms of specific activity, subunit composition, antigenicity, amino acid composition and tryptic peptides. However, the hepatoma enzyme exhibited more charge heterogeneity to give multiple acidic variant forms which were devoid or much less in the normal liver enzyme. When the hepatoma enzyme was treated with endo-beta-N-acetylglucosaminidase H, the acidic variant forms disappeared and were converted into forms identical to those of normal liver. The content of mannose-6-phosphate in the hepatoma enzyme was twice as much as that in the normal liver enzyme. Thus, charge heterogeneity found in hepatoma cathepsin D is ascribed to increased phosphorylation on oligosaccharides bound to the enzyme, most probably due to cancer-associated, impaired processing in carbohydrate moiety. A significant elevation of cathepsin D activity per tissue proteins was observed in hepatoma as compared to normal liver. In contrast, true specific activity per cathepsin D protein in hepatoma was significantly lowered than that of normal liver. The lower true specific activity in hepatoma tissue may be attributed to an increased content in an inactive, large-molecular precursor form of the enzyme.
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PMID:[Tumor-associated impairment of the processing of hepatoma cathepsin D]. 283 81


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