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

---

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

Proteinase A, a yeast aspartyl protease that is highly homologous to the mammalian lysosomal aspartyl protease, cathepsin D, was expressed in Xenopus oocytes and its biosynthesis and post-translational modifications were characterized. While 29-45% of the proteinase A was secreted from oocytes, approximately 37% of the cell-associated proteinase A underwent proteolytic cleavage, characteristic of delivery to a lysosomal organelle. Although proteinase A is not targeted to the yeast vacuole by a mannose 6-phosphate receptor-dependent pathway, 2-5% of the proteinase A molecules expressed in oocytes bound to a Man-6-P receptor column. However, analysis of its [2-3H]mannose-labeled oligosaccharides revealed that 14-23% of these units contain phosphomannosyl residues. A hybrid molecule (H6), in which the propiece and first 12 amino acids of proteinase A were changed to the cathepsin D sequence, was also expressed in oocytes. The binding of H6 to the Man-6-P receptor was approximately 12-fold greater than observed for proteinase A. This increased level of receptor binding could be accounted for by three factors: 1) a small increase in the total amount of phosphorylated oligosaccharides, 2) an increase in the number of oligosaccharides which acquire two phosphomonoesters, and 3) the presence of a greater percentage of oligosaccharides with one phosphomonoester which exhibit high affinity binding to the Man-6-P receptor. These results demonstrate that proteinase A is recognized by UDP-GlcNAc:lysosomal enzyme N-acetylglucosaminylphosphotransferase. However, this interaction is altered by the addition of cathepsin D sequences, resulting in the generation of a higher affinity ligand for binding to the Man-6-P receptor.
...
PMID:Expression of the yeast aspartyl protease, proteinase A. Phosphorylation and binding to the mannose 6-phosphate receptor are altered by addition of cathepsin D sequences. 252 Dec 20

Immunogold double-labeling and ultrathin cryosections were used to compare the subcellular distribution of albumin, mannose 6-phosphate receptor (MPR), galactosyltransferase, and the lysosomal enzymes cathepsin D, beta-hexosaminidase, and alpha-glucosidase in Hep G2 cells. MPR and lysosomal enzymes were found throughout the stack of Golgi cisternae and in a trans-Golgi reticulum (TGR) of smooth-surfaced tubules with coated buds and vesicles. The trans-Golgi orientation of TGR was ascertained by the co-localization with galactosyltransferase. MPR was particularly abundant in TGR and CURL, the compartment of uncoupling receptors and ligands. Both TGR and CURL also contained lysosomal enzymes, but endogenous albumin was detected in TGR only. The coated buds on TGR tubules contained MPR, lysosomal enzymes, as well as albumin. MPR and lysosomal enzymes were also found in coated pits of the plasma membrane. CURL tubules seemed to give rise to smooth vesicles, often of the multivesicular body type. In CURL, the enzymes were found in the lumina of the smooth vesicles while MPR prevailed in the tubules. These observations suggest a role of CURL in transport of lysosomal enzymes to lysosomes. When the cells were treated with the lysosomotropic amine primaquine, binding of anti-MPR to the cells in culture was reduced by half. Immunocytochemistry showed that MPR accumulated in TGR, especially in coated buds. Since these buds contain endogenous albumin and lysosomal enzymes also, these data suggest that coated vesicles originating from TGR provide for a secretory route in Hep G2 cells and that this pathway is followed by the MPR system as well.
...
PMID:Possible pathways for lysosomal enzyme delivery. 293 16

An affinity-purified rabbit antibody against rat liver mannose 6-phosphate receptor (MP-R) was prepared. The antibody was directed against a 215 kd-polypeptide and it recognized both ligand-occupied and free receptor. Anti-MP-R was used for immunofluorescence and immunoelectron microscopy of cryosections from rat liver. MP-R was demonstrated in all parenchymal liver cells, but not in endothelial lining cells. MP-R labeling was found at the entire plasma membrane, in coated pits and coated vesicles, in the compartment of uncoupling receptor and ligand, and in the Golgi complex. Lysosomes showed only scarce MP-R label. In double-labeling immunoelectron microscopy, MP-R co-localized with albumin in the Golgi cisternae and in secretory vesicles with lipoprotein particles. Cathepsin D was associated with MP-R in the Golgi cisternae. This finding indicates that MP-R/cathepsin D complexes traverse the Golgi complex on their way to the lysosomes. The possible involvement of CURL in lysosomal enzyme targeting is discussed.
...
PMID:Ultrastructural localization of the mannose 6-phosphate receptor in rat liver. 632 24

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.
...
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

Previous work with the yeast Saccharomyces cerevisiae has demonstrated a role for a phosphatidylinositol-specific PI 3-kinase, the product of the VPS34 gene, in the targeting of newly synthesized proteins to the vacuole, an organelle functionally equivalent to mammalian lysosomes (Schu, P. V., K. Takegawa, M. J. Fry, J. H. Stack, M. D. Waterfield, and S. D. Emr. 1993. Science [Wash. DC]. 260:88-91). The activity of Vps34p kinase is significantly reduced by the PI 3-kinase inhibitors wortmannin, a fungal metabolite, and LY294002, a quercetin analog (Stack, J. H., and S. D. Emr. 1994. J. Biol. Chem. 269:31552-31562). We show here that at concentrations which inhibit VPS34-encoded PI 3-kinase activity, wortmannin also inhibits the processing and delivery of newly synthesized cathepsin D to lysosomes in mammalian cells with half-maximal inhibition of delivery occurring at 100 nM wortmannin. As a result of wortmannin action, newly synthesized, unprocessed cathepsin D is secreted into the media. Moreover, after accumulation in the trans-Golgi network (TGN) at 20 degrees C, cathepsin D was rapidly missorted to the secretory pathway after addition of wortmannin and shifting to 37 degrees C. At concentrations that inhibited lysosomal enzyme delivery, both wortmannin and LY294002 caused a highly specific dilation of mannose 6-phosphate receptor (M6PR)-enriched vesicles of the prelysosome compartment (PLC), which swelled to approximately 1 micron within 15 min after treatment. With increasing time, the inhibitors caused a significant yet reversible change in M6PR distribution. By 3 h of treatment, the swollen PLC vacuoles were essentially depleted of receptors and, in addition, there was a fourfold loss of receptors from the cell surface. However, M6PRs were still abundant in the TGN. These results are most consistent with the interpretation that PI 3-kinase regulates the trafficking of lysosomal enzymes by interfering with a M6PR-dependent sorting event in the TGN. Moreover, they provide evidence that trafficking of soluble hydrolases to mammalian lysosomes and yeast vacuoles rely on similar regulatory mechanisms.
...
PMID:Role for phosphatidylinositol 3-kinase in the sorting and transport of newly synthesized lysosomal enzymes in mammalian cells. 764 97

Endocytosed proteins are sorted in early endosomes to be recycled to the plasma membrane or transported further into the degradative pathway. We studied the role of endosomes acidification on the endocytic trafficking of the transferrin receptor (TfR) as a representative for the recycling pathway, the cation-dependent mannose 6-phosphate receptor (MPR) as a prototype for transport to late endosomes, and fluid-phase endocytosed HRP as a marker for transport to lysosomes. Toward this purpose, bafilomycin A1 (Baf), a specific inhibitor of the vacuolar proton pump, was used to inhibit acidification of the vacuolar system. Microspectrofluorometric measurement of the pH of fluorescein-rhodamine-conjugated transferrin (Tf)-containing endocytic compartments in living cells revealed elevated endosomal pH values (pH > 7.0) within 2 min after addition of Baf. Although recycling of endocytosed Tf to the plasma membrane continued in the presence of Baf, recycled Tf did not dissociate from its receptor, indicating failure of Fe3+ release due to a neutral endosomal pH. In the presence of Baf, the rates of internalization and recycling of Tf were reduced by a factor of 1.40 +/- 0.08 and 1.57 +/- 0.25, respectively. Consequently, little if any in TfR expression at the cell surface was measured during Baf treatment. Sorting between endocytosed TfR and MPR was analyzed by the HRP-catalyzed 3,3'-diaminobenzidine cross-linking technique, using transferrin conjugated to HRP to label the endocytic pathway of the TfR. In the absence of Baf, endocytosed surface 125I-labeled MPR was sorted from the TfR pathway starting at 10 min after uptake, reaching a plateau of 40% after 45 min. In the presence of Baf, sorting was initiated after 20 min of uptake, reaching approximately 40% after 60 min. Transport of fluid-phase endocytosed HRP to late endosomes and lysosomes was measured using cell fractionation and immunogold electron microscopy. Baf did not interfere with transport of HRP to MPR-labeled late endosomes, but nearly completely abrogated transport to cathepsin D-labeled lysosomes. From these results, we conclude that trafficking through early and late endosomes, but not to lysosomes, continued upon inactivation of the vacuolar proton pump.
...
PMID:Transport from late endosomes to lysosomes, but not sorting of integral membrane proteins in endosomes, depends on the vacuolar proton pump. 764

We evaluated levels of mannose-6-phosphate/insulin growth factor-II receptor (M6P/IGFII-R) RNA in 37 breast cancer tumors by quantitative in situ hybridization using a computer-aided image analyzer and compared them to cathepsin D RNA and protein levels in the same tissues. Breast cancer cells expressed more cathepsin D and M6P/IGFII-R RNA than fibroblasts in the same tumors. We found a significant increase of cathepsin D RNA (P = 1 x 10(-5)) and M6P/IGFII-R RNA (P = 0.02) in breast cancer cells compared to epithelial cells of benign mastopathies. There was a positive correlation (r = 0.65; P = 1 x 10(-5)) between M6P/IGFII-R and cathepsin D RNA levels measured on serial sections. This contrasted with the inverse relationship of these 2 RNA species in breast cancer cell lines where estrogen down-regulates M6P/IGFII receptor RNA levels. Moreover, in vivo we found no correlation between the M6P/IGFII-R RNA level and menopausal or estrogen receptor status, suggesting that the in vivo regulation of M6P/IGFII-R RNA differs from its in vitro regulation in cell lines. The M6P/IGFII-R RNA level was not correlated with cathepsin D status, histological grade, and tumor size but was significantly higher in lymph node-positive tumors (P = 0.047). The M6P/IGFII-R could therefore be an additional parameter to predict aggressive breast cancers, complementing cathepsin D assays and other more classical prognostic parameters.
...
PMID:Correlation between mannose-6-phosphate/IGFII receptor and cathepsin D RNA levels by in situ hybridization in benign and malignant mammary tumors. 768 51

A novel combinatorial mutagenesis strategy (shuffle mutagenesis) was developed to identify sequences in the propiece and amino lobe of cathepsin D which direct oligosaccharide phosphorylation by UDP-GlcNAc:lysosomal enzyme N-acetylglucosamine 1-phosphotransferase. Propiece restriction fragments and oligonucleotide cassettes corresponding to 13 regions of the cathepsin D and glycopepsinogen amino lobes were randomly shuffled together to generate a large library of chimeric molecules. The library was inserted into an expression vector encoding the carboxyl lobe of cathepsin D with a carboxyl-terminal myc epitope and a CD8 transmembrane extension. Transfected COS1 cells expressing the membrane-anchored forms of the cathepsin D/glycopepsinogen chimeras at the cell surface were selected with solid phase mannose 6-phosphate receptor or an antibody to the myc epitope. Plasmids were rescued in Escherichia coli and sequenced by hybridization to the original oligonucleotide cassettes. Two regions of the cathepsin D amino lobe (segments 7 and 12) were found to contribute to proper folding, surface expression, and selective phosphorylation of the carboxyl lobe oligosaccharide. Two different cathepsin D regions (the propiece and segment 5) cooperated with a previously identified recognition element in the carboxyl lobe to allow efficient phosphorylation of both the amino and carboxyl lobe oligosaccharides. Three general models for extending the catalytic reach of N-acetylglucosamine 1-phosphotransferase to widely spaced oligosaccharides are presented.
...
PMID:A novel mutagenesis strategy identifies distantly spaced amino acid sequences that are required for the phosphorylation of both the oligosaccharides of procathepsin D by N-acetylglucosamine 1-phosphotransferase. 781 70

We have compared the intracellular localization of plasma membrane proteins anchored either with a transmembrane segment or with a glycosylphosphatidylinositol moiety to estimate the effects of membrane anchor on protein segregation in the non-polarized form of the human colon cancer cell line HT-29 18. We have monitored two endogenous proteins: the carcinoembryonic antigen, a glycosylphosphatidylinositol protein and the transmembrane protein dipeptidyl peptidase IV, and two transfected proteins: the glycosylphosphatidylinositol protein Thy-1 and an engineered transmembrane form of Thy-1. Using immunocytochemistry on ultra-thin cryosections and confocal microscopy, we detected a carcinoembryonic antigen-rich vesicular compartment, excluding classical pre-lysosomal and lysosomal markers such as mannose 6-phosphate receptor, lamp-1 and cathepsin D. This compartment, where carcinoembryonic antigen accumulated, excluded the transmembrane protein dipeptidyl peptidase IV and was reduced during the polarization of the cells. Moreover, the glycosylphosphatidylinositol form of Thy-1 also accumulated in the carcinoembryonic antigen-rich compartment whereas the transmembrane form of Thy-1 was excluded. We proposed that, in the non-polarized HT-29 18 cells, accumulation of glycosylphosphatidylinositol proteins independently of transmembrane proteins reveals different intracellular fates for proteins according to their anchor in the plasma membrane.
...
PMID:GPI membrane anchor is determinant in intracellular accumulation of apical plasma membrane proteins in the non-polarized human colon cancer cell line HT-29 18. 787 37

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.
...
PMID:Proteolytic processing of cathepsin D in prelysosomal organelles. 795 14


<< Previous 1 2 3 4 5 6 7 8 Next >>

---