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)

Autolysosomes were isolated from rat livers treated with leupeptin by a combination of differential and Percoll density gradient centrifugation techniques. The purified autolysosome fraction was verified by morphological analysis to be highly purified and to contain contaminants which were scarcely detectable. The enrichment of the lysosomal enzyme activities in the purified autolysosomes over the homogenate was 12-, 14-, 22-, and 24-fold for beta-glucuronidase, acid phosphatase, beta-N-acetylglucosaminidase, and cathepsin D, respectively. Measurement of the activity of the marker enzymes for various subcellular organelles also proved that the purified autolysosome fraction was essentially free from contamination by other organelles. When the autolysosomes isolated from rat livers treated with leupeptin for 1 h were disrupted by osmolysis, acid hydrolases were easily solubilized. Acid phosphatase, however, became membrane bound in the autolysosomes prepared at longer periods of time after the leupeptin treatment. The autolysosomes exhibited enhanced permeability of the membranes after a short duration of time after the leupeptin treatment (30 and 60 min) and became stabilized later. These changes in the properties of the autolysosomes with time after the leupeptin treatment may be interpreted as meaning that progressive rearrangement of the lysosomal constituents occurred within the autolysosomes with time after the genesis.
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PMID:Isolation and characterization of autolysosomes which appeared in rat liver after leupeptin treatment. 711 55

Chronic alcohol intoxication led to an increase in activity of alcohol dehydrogenase and to decrease -- of aldehyde dehydrogenase and the microsomal ethanol oxidizing system (MEOS) with simultaneous activation of cytochrome P-450 in liver tissue of rats during ontogenesis. Ethanol, which did not affect the enzymatic status of lysosomes within ontogenesis (alpha- and beta-glucosidases, alpha- and beta-galactosidases, alpha-mannosidase, beta-N-acetylglucosaminidase, beta-xylosidase, beta-glucuronidase, beta-N-acetyl galactosaminidase acid RNAase, arylsulfatases A and B, cathepsin D), activated the majority of hydrolases in both embryonal and postnatal periods of development. Distinct increase in lipoperoxidation was detected under conditions of chronic alcohol intoxication.
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PMID:[Enzyme characteristics of the rat liver in ontogeny in chronic alcohol intoxication]. 720 88

Insulin-like growth factor-II (IGF-II) and lysosomal enzymes bearing the mannose 6-phosphate (Man6P) recognition marker, bind to two distinct binding sites of the IGF-II/M6P receptor. The two classes of ligands reciprocally modulate the binding of the other class of ligand to the receptor [Kiess, W., Thomas, C. L., Greenstein, L., Lee, L., Sklar, M. M., Rechler, M. M., Sahagian, G. G. & Nissley, S. P. (1989) J. Biol. Chem. 264, 4710-4714]. We asked whether or not overexpression of pro-IGF-II by cells in culture leads to missorting of lysosomal enzymes. Human embryonal kidney fibroblasts were transfected with the full-length human IGF-II cDNA or a control cDNA. Solution hybridization/RNase protection experiments using a human IGF-II riboprobe showed that two transfectants expressed large quantities of IGF-II mRNA, whereas the non-transfected cells did not. The analysis of conditioned media revealed that these cells secrete approximately 0.15 micrograms and 1.0 micrograms immunoreactive IGF-II/ml and 22 x 10(6) cells and 24 x 10(6) cells within 24 hours. Immunoreactive IGF-II was shown by Western blotting to represent 17-kDa pro-IGF-II. The amount of the lysosomal enzyme, beta-hexosaminidase, was approximately twofold increased in the conditioned media from pro-IGF-II overexpressing cells compared with control media, as shown by Western-blot analysis and immunoprecipitation of media extracts of metabolically labeled cells. The synthesis rate of beta-hexosaminidase was not affected by pro-IGF-II overexpression. In addition, the basal amount of another newly synthesized lysosomal enzyme, the cathepsin D precursor, was also twofold higher in pro-IGF-II overexpressing cells than in control cells. In contrast, the surface binding and cellular uptake rate of a Man6P-containing neoglycoprotein did not differ between the cell lines. The results indicate that the overexpression of pro-IGF-II doubles the secretion and/or reduces the re-uptake of beta-hexosaminidase and cathepsin D to approximately 20% of the total synthesized enzymes in human embryonal kidney fibroblasts compared to control cells. We hypothesize that, in cells synthesizing high amounts of pro-IGF-II, the growth factor may modulate the targeting of a portion of lysosomal enzymes, mainly by partially enhancing the secretion of newly synthesized enzymes and, in addition, possibly by affecting the re-uptake mechanism.
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PMID:Does the overexpression of pro-insulin-like growth factor-II in transfected human embryonic kidney fibroblasts increase the secretion of lysosomal enzymes? 755 47

In HL-60 cells the most abundant isoenzymes of beta-N-acetyl-hexosaminidase are A (alpha beta) and S (alpha alpha). Sub-cellular fractionation of HL-60 cells by differential centrifugation showed that both A and S forms were present in the lysosomal and post-lysosomal (soluble) fractions in approximately equal abundance. Ion-exchange chromatography showed that a fraction enriched with plasma membranes had the A form, and a form of beta-N-acetylhexosaminidase less acidic than A, but there was no S. Analysis of the alpha-subunits of beta-N-acetylhexosaminidases A and S using Western blotting and immuno-detection with antisera raised to synthetic peptides showed that mature alpha-subunits were present in both A and S isolated from the lysosomal fraction. This observation establishes that the alpha alpha-dimer of beta-N-acetyl-hexosaminidase (S) can be transported to lysosomes in HL-60 cells whereas there is good evidence that this does not take place in fibroblasts. HL-60 cells were not stimulated to secrete lysosomal enzymes by incubating them with NH4Cl and, unlike fibroblasts, are unlikely to use mannose-6-phosphate mediated transport of beta-N-acetylhexosaminidases to lysosomes. Comparison of the sequence of the beta-N-acetylhexosaminidase alpha-subunit with a 43 amino acid sequence of cathepsin D, though to function in the mannose-6-phosphate independent targeting of this enzyme to lysosomes, showed alignment in a region towards the C-terminus in which 21% of the residues were identical with the interposition of a one amino acid gap.
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PMID:Beta-N-acetylhexosaminidases A and S have similar sub-cellular distributions in HL-60 cells. 772 24

MHC class II (MHC-II) molecules bind fragments of exogenous Ags in an intracellular endocytotic compartment. In view of divergent data on the MHC-II distribution in different cell lines, it was of interest to localize MHC-II molecules in a natural and the most potent APC type, the dendritic cell (DC). By using immunogold labeling of ultrathin cryosections of cultured mouse spleen DC, we found that MHC-II molecules were present abundantly at the plasma membrane and in intracellular compartments containing internal membrane vesicles and/or membrane sheets. The majority of these compartments was situated late in the endocytotic route, as demonstrated by the late appearance (after a lag of 30 min) of internalized exogenous tracer. These compartments contained the lysosomal enzymes cathepsin D and beta-hexosaminidase, but lacked the late endosomal marker cation-dependent mannose-6-phosphate receptor. We conclude that most of the intracellular MHC-II molecules in cultured spleen DC reside in a compartment with (pre)lysosomal characteristics, resembling the so-called MHC-II-enriched compartments (MIIC), originally described in B cells. We also investigated whether the presence of MHC-II molecules in endocytotic compartments was related to the kinetics of Ag processing and presentation by these cells. Pulse-chase endocytosis experiments with hen egg lysozyme (HEL) as a model Ag showed that activated spleen DC were able to efficiently process and present this Ag to an HEL-specific T hybridoma cell line. However, presentation started only after a lag of 2 h and was maximal after 6 h. The difference in time between the arrival of Ag in proteolytic endocytotic compartments, in particular MIIC, and effective Ag presentation is discussed in the context of DC maturation.
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PMID:MHC class II compartments and the kinetics of antigen presentation in activated mouse spleen dendritic cells. 775 23

The way in which the human epididymis modifies spermatozoa during their sojourn in this structure might be clarified by knowledge of the nature of its secretions. We have examined the presence of several lysosomal hydrolases in human epididymal tissue and fluids, and their synthesis and secretion by monolayer cultures. Tissues were obtained from men undergoing orchidectomy for prostatic carcinoma. The enzymes cathepsin D and acid alpha-glucosidase were localised in the lysosomes of epithelial cells from the corpus epididymidis, by an immunocytochemical technique. Cathepsin D was also found in epithelial cells of the efferent ducts within lysosomes, apical vesicles and multivesicular bodies. No immunolocalisation of acid glucosidase in the efferent ducts or on the microvilli of the corpus was demonstrable. Cathepsin D, beta-hexosaminidase (N-acetylglucosaminidase) and alpha-glucosidase were measurable in the luminal fluid from the human corpus epididymidis; beta-hexosaminidase was secreted into the culture medium by confluent monolayers of epididymal and efferent duct cells. Immunoprecipitation of cell extracts and culture medium of these cultures incubated with 35S-methionine revealed that the precursors of cathepsin D and beta-hexosaminidase were synthesized and secreted by such monolayers. Thus, active lytic enzymes are secreted by the human epididymis and could modify sperm membranes.
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PMID:Immunocytochemical localisation of some lysosomal hydrolases, their presence in luminal fluid and their directional secretion by human epididymal cells in culture. 778 Oct 38

To determine their value as markers of the clinical stage of human immunodeficiency virus (HIV) disease, plasma activities of lysosomal glycosidases were determined in the plasma of 97 HIV-infected patients: molecular forms of cathepsin D were characterized by Western blot to examine the mode of enzyme release. In patients with Centers for Disease Control and Prevention stage II and III of HIV disease, plasma activity of beta-hexosaminidase was significantly increased. In patients with stage III infection, alpha-mannosidase activity was also significantly increased and cathepsin D was elevated and present only in its premature form. Thus, determination of plasma activities of beta-hexosaminidase and alpha-mannosidase in HIV-positive persons may be useful for distinguishing the clinical stage of disease. The elevation of precursors of cathepsin D in patients with stage III HIV disease indicates that secretion of lysosomal enzymes rather than leakage of enzymes from damaged cells is markedly elevated.
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PMID:Elevated levels of activities of beta-hexosaminidase and alpha-mannosidase in human immunodeficiency virus-infected patients. 787 17

The initial steps of proteolytic processing of newly synthesized cathepsin D were investigated in prelysosomal membranes, which were defined by their contents of 300 kDa mannose 6-phosphate receptor (MPR 300). MPR 300-containing vesicles were immuno-isolated from a postmitochondrial supernatant of HepG2 cells using a peptide-specific antibody directed against the 15 C-terminal amino acids of the cytoplasmic domain of MPR 300. In the immunoisolated fraction, MPR 300 was enriched 11.5-fold over [35S]polypeptides, 29-fold over the lysosomal marker beta-hexosaminidase and 4.5-fold over the trans Golgi marker galactosyltransferase, when referred to the postmitochondrial supernatant. MPR 300-containing vesicles harbored, on average, 12% of the cathepsin D precursor from the postmitochondrial supernatant, suggesting that segregation of MPR 300 and cathepsin D occurs rapidly in prelysosomal organelles. Detection of low, but significant amount of mature cathepsin D in the immunoisolated fraction suggests that proteolytic processing is initiated in MPR 300-containing vesicles or in tightly associated prelysosomal vesicles, which are distinct from mature lysosomes.
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PMID:Proteolytic processing of cathepsin D in prelysosomal organelles. 795 14

The secretion of pro-cathepsin D (pro-cath-D) in some human metastatic breast cancer cells (MCF7, MDA/MB231), contrary to normal mammary cells, is not increased by ammonium chloride treatment, indicating a mannose-6-phosphate-independent sorting to lysosomes. By studying a variety of cell lines and lysosomal enzymes, we show that secretion of newly synthesized pro-cath-D was not mediated by the 46-kDa mannose-6-phosphate receptor (MPR) and that its resistance to NH4Cl for secretion was specific to cath-D and not to other lysosomal enzymes. This resistance appeared to be correlated with the basal hypersecretion of pro-cath-D, but not with its overexpression. By contrast, pro-cath-D secretion was increased by NH4Cl in fibroblasts and nontumoral epithelial mammary cells, suggesting a specificity for cancer cells. Immunofluorescence staining showed that pro-cath-D, but neither cathepsin B nor beta-hexosaminidase, accumulated in intracytoplasmic vesicles of cells treated with ammonium chloride. In pulse--chase experiments and by subcellular fractionation on Percoll gradient, cath-D was found to be sorted into dense lysosomes whether cells were treated or not by NH4Cl. Treatment of cells with NH4Cl, however, inhibited processing and maturation of pro-cath-D, which was also observed in light vesicles in the absence of NH4Cl. Part of pro-cath-D, but not processed enzyme, was also found to be membrane associated in saponin-permeabilized cells. We conclude that in breast cancer cells, the MPR-independent pathway of pro-cath-D to lysosome is predominant compared to normal cells and other lysosomal enzymes. This alternative pathway should therefore be considered, in addition to MPR, to explain pro-cath-D sorting and activation in breast cancer cells.
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PMID:Specific mannose-6-phosphate receptor-independent sorting of pro-cathepsin D in breast cancer cells. 795 63

We have compared the intracellular transport and subcellular distribution of MHC class II-invariant chain complexes in a wild-type HLA-DR3 homozygous cell line and a mutant cell line, T2.DR3. The latter has a defect in antigen processing and accumulates HLA-DR3 molecules associated with an invariant chain-derived peptide (CLIP) rather than the normal complement of peptides derived from endocytosed proteins. We find that in the wild-type cells, CLIP is transiently associated with HLA-DR3 molecules, suggesting that the peptide is a normal class II-associated intermediate generated during proteolysis of the invariant chain. In the mutant cell line proteolysis of the invariant chain is less efficient, and HLA-DR3/CLIP complexes are generated much more slowly. Examination of the mutant cell line by immunoelectronmicroscopy shows that class II-invariant chain complexes accumulate intracellularly in large acidic vesicles which contain lysosomal markers, including beta-hexosaminidase, cathepsin D, and the lysosomal membrane protein CD63. The markers in these vesicles are identical to those seen in the class II-containing vesicles (MIICs) seen in the wild-type cells but the morphology is drastically different. The vesicles in the mutant cells are endocytic, as measured by the internalization of BSA-gold conjugates. The implication of these findings for antigen processing in general and the nature of the mutation in particular are discussed.
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PMID:Transport and intracellular distribution of MHC class II molecules and associated invariant chain in normal and antigen-processing mutant cell lines. 820 55


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