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)

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

Mannose 6-phosphate receptors carry newly synthesized lysosomal hydrolases from the trans-Golgi network to endosomes, then return to the trans-Golgi network for another round of enzyme delivery. Wortmannin, an inhibitor of phosphatidylinositol 3-kinase, interferes with the delivery of newly synthesized lysosomal enzymes to lysosomes. We used two independent assays of mannose 6-phosphate receptor trafficking to determine the precise step that is blocked by wortmannin. Using an assay that monitors resialylation of desialylated cell surface 300-kDa mannose 6-phosphate receptors, we found that receptor endocytosis and transport to the trans-Golgi network were not inhibited by 2 microM wortmannin. In addition, this concentration of drug had no effect on the transport of the mannose 6-phosphate receptor from late endosomes to the trans-Golgi network using a system that reconstitutes this transport process in cell extracts. Under the same conditions, wortmannin significantly inhibited the generation of mature cathepsin D. In addition, the structurally unrelated phosphatidylinositol 3-kinase inhibitor, LY294002, was also without effect when added to in vitro endosome-trans-Golgi network transport reactions. These experiments demonstrate that the interruption in lysosomal enzyme targeting is most likely due to a wortmannin-sensitive process required for the export of these receptors from the trans-Golgi network, consistent with the established role of phosphatidylinositol 3-kinase in the equivalent transport process in Saccharomyces cerevisiae.
...
PMID:Phosphatidylinositol 3-kinase is not required for recycling of mannose 6-phosphate receptors from late endosomes to the trans-Golgi network. 924 39

Peptide-class II complexes are assembled in endocytic, lysosome-like compartments where newly synthesized class II molecules are targeted from the trans-Golgi network (TGN). Recent studies have implicated phosphatidylinositol 3-kinase (PI3-kinase) as an essential component in membrane trafficking from the TGN to lysosomes. Here, using subcellular fractionation, we show PI3-kinase activity associated with subcellular fractions which contain the class II peptide-loading compartment (IIPLC) in B cells. At concentrations required for inhibition of PI3-kinase activity in vivo, wortmannin blocked the processing and presentation of antigen by B cells to T cells. Treatment of B cells with wortmannin significantly limited the proteolytic degradation of invariant chain and the formation of peptide-class II complexes. Subcellular fractionation coupled with pulse-chase analyses showed that invariant chain and class II molecules trafficked to the IIPLC in wortmannin-treated cells. However, wortmannin prevented the maturation and correct targeting to the IIPLC of cathepsin D, a protease necessary for the degradation of invariant chain and assembly of processed antigen-class II complexes. These results suggest that li-class II complexes traffic to the IIPLC via a pathway that is relatively insensitive to wortmannin, but suggest a role for PI3-kinases in the trafficking of other components necessary for the assembly of processed antigen class II complexes to the IIPLC.
...
PMID:Wortmannin, a phosphatidylinositol 3-kinase inhibitor, blocks the assembly of peptide-MHC class II complexes. 941 32

Aspergillus fumigatus is the most prevalent airborne fungal pathogen responsible for fatal invasive aspergillosis in immunocompromised patients. Upon arrival in the lung alveolus, conidia of A. fumigatus are phagocytosed by alveolar macrophages, the major phagocytic cells of the lung. Engulfment and intracellular trafficking of A. fumigatus conidia in alveolar macrophages of two different origins, the murine cell line MH-S and human pulmonary alveolar macrophages, were analyzed by electron microscopy and immunofluorescence. Phagocytosis of A. fumigatus conidia required actin polymerization and phosphatidylinositol 3-kinase activity. Fusion of A. fumigatus phagosomes with early and late endosomes was shown by immunolabeling with specific markers for the transferrin receptor, early endosome antigen, and Rab7. Maturation of A. fumigatus phagolysosomes was monitored by using a fixable acidotropic probe, LysoTracker Red DND-99, and an anti-cathepsin D antibody. Bafilomycin A-induced inhibition of lysosomal acidification abolished the conidial killing by the macrophages. These data suggest that the maturation of A. fumigatus phagosomes results from fusion with the compartments of the endocytic pathway and that the killing of conidia depends on phagolysosome acidification. A model for the phagocytosis of A. fumigatus conidia by alveolar macrophages is proposed on the basis of these results.
...
PMID:Phagocytosis and intracellular fate of Aspergillus fumigatus conidia in alveolar macrophages. 1254 May 71

Small GTPase RhoA regulates signal transduction from receptors in the membrane to a variety of cellular events related to cell morphology, motility, cytoskeletal dynamics, cytokinesis, and tumour progression, but it is unclear how RhoA regulates intracellular membrane dynamics of lysosomes. We showed previously by confocal immunofluorescence microscopy that the transfection of dominant active RhoA in MM1 cells causes the dispersal translocation of lysosomes stained for cathepsin D throughout the cytoplasm. Y-27632, a selective inhibitor of p160ROCK, impeded the cellular redistribution of lysosomes and promoted reclustering of lysosomes toward the perinuclear region. Here we have further investigated whether the acidic lysosomal vesicles dispersed throughout the cytoplasm are applied to the early endosomes in the endocytic pathway, and we demonstrate that the dispersed lysosomes were accessible to endocytosed molecule such as dextran, and their acidity was not changed, as determined by increased accumulation of the acidotropic probe LysoTracker Red. Brefeldin A did not induce the tabulation of these dispersed lysosomes, but it caused early endosomes to form an extensive tubular network. The dispersed lysosomes associated with cathepsin D and LIMPII were not colocalized with early endosomes, and these vesicles were not inaccessible to the endocytosed anti-transferrin receptor antibody. Moreover, wortmannin, an inhibitor of phosphatidylinositol 3-kinase, induced a dramatic change in LIMPII-containing structures in which LIMPII-positive swollen large vacuoles were increased and small punctate structures disappeared in the cytoplasm. These swollen vacuoles were not doubly positive for LIMPII and transferrin receptor, and were not inaccessible to the internalized anti-transferrin receptor antibody. Therefore, our novel findings presented in this paper indicate that RhoA activity causes a selective translocation of lysosomes without perturbing the machinery of endocytic pathway.
...
PMID:A role for small GTPase RhoA in regulating intracellular membrane traffic of lysosomes in invasive rat hepatoma cells. 1258 97

Small GTPase Rho and its downstream effectors, ROCK family of Rho-associated serine-threonine kinases, are thought to participate in cell morphology, motility, and tumor progression through regulating the rearrangement of actin cytoskeleton. Here we present evidence that transfection of human breast cancer cells with cDNA encoding a dominant active mutant of ROCK causes dispersal of lysosomal vesicles throughout the cytoplasm without perturbing the machinery of the endocytic pathway. The intracellular distribution of lysosomes and endocytosed transferrin, an early endosomal marker, were further assessed by confocal immunofluorescence microscopy. In the active ROCK transfected cells the lysosomal proteins, cathepsin D, LIMPII, and LAMP1, were found throughout the cytoplasm in dispersed small vesicles, which were accessible to the endocytosed Texas Red-labeled transferrin. 3D-image analysis of lysosomal distribution in the active ROCK transfectants revealed abundant punctate signals in the peripheral region of the basal plasma membrane. Cells expressing vector alone did not exhibit these alterations. Wortmannin, a phosphatidylinositol 3-kinase inhibitor, induced LIMPII-positive/ transferrin negative large vacuoles in the perinuclear region, and disappearence of the dispersed small vesicular structures. To our knowledge, this is the first evidence that increasing ROCK expression contributes to selective cellular dispersion of lysosomes in invasive breast cancer cells.
...
PMID:Overexpression of ROCK in human breast cancer cells: evidence that ROCK activity mediates intracellular membrane traffic of lysosomes. 1285 12

Aquaporin-2 (AQP2) is one of the water-channel proteins expressed in principal cells of kidney collecting ducts, where it is stored in the intracellular compartment. Previous studies have demonstrated that AQP2 vesicles constitute a distinct intracellular compartment partially overlapping with early endosomes. In this report, we performed in vitro experiments using the renal epithelial cell line, Madin-Darby canine kidney (MDCK) cells, stably expressing AQP2 (MDCK-hAQP2). In nonpolarized cells, AQP2 vesicles were scattered in the cytoplasm and did not colocalize with Golgi 58K or TGN38. Small portions of AQP2 vesicles were positive for the lysosome marker cathepsin D. An early endosome antigen (EEA1) localized around AQP2 vesicles in close proximity, suggesting involvement of the endosomal system in the trafficking of AQP2. AQP2 vesicles are distinct from other recycling molecules, such as glucose transporter 4 (GLUT4) and endocytosed transferrin. In polarized MDCK-hAQP2 cells, AQP2 vesicles were localized in the subapical recycling compartment and distinct from the Golgi apparatus, trans-Golgi network, lysosome, and early endosome in the nonstimulated state. When the cells were treated with forskolin, translocation of AQP2 to the apical membrane was observed. Washout of forskolin induced retrieval of AQP2 into the cytoplasm, and AQP2 was transiently colocalized with EEA1-positive endosomes. Then, AQP2 moved from EEA1-positive endosomes to the subapical AQP2-storage compartment, which is sensitive to wortmannin and LY294002. These results suggest that AQP2 resides in a recycling compartment at the apical side in polarized MDCK-hAQP2 cells, and its retrieval uses the apical endosomal system and the phosphatidylinositol 3-kinase-dependent pathway.
...
PMID:Aquaporin-2 is retrieved to the apical storage compartment via early endosomes and phosphatidylinositol 3-kinase-dependent pathway. 1515 71

Ras expression induces increased expression and altered targeting of lysosomal proteases in multiple cell types, but the specific downstream cytoplasmic signaling pathways mediating these changes have not been identified. In this study, we compared the involvement of 3 major Ras effectors, Raf, phosphatidylinositol 3-kinase (PI3K) and Ral guanine nucleotide exchange factor (RalGEF) in the Ras-mediated alteration of lysosomal protease protein expression and targeting in rat 208F fibroblasts and rat ovarian surface epithelial (ROSE) cells. Effector domain mutants of Ras, constitutively activated variants of Raf, PI3K and RalGEF and pharmacologic inhibitors of MEK and PI3K were utilized to determine the role of these downstream pathways in mediating fibroblast transformation and lysosomal protease regulation in the fibroblasts and epithelial cells. We found that Raf activation of the ERK mitogen-activated protein kinase pathway alone was sufficient to cause morphologic and growth transformation of the fibroblasts and was necessary and sufficient to alter cathepsin L expression and targeting. In contrast, transformation and upregulation of cathepsin L expression in the epithelial cells required the activity of all 3 Ras effectors. Increased protease secretion from the epithelial cells was not observed on ectopic expression of Ras, as it was from the fibroblasts, consistent with the utilization of different signaling pathways in the 2 cell types. In neither cell type did Ras expression increase the expression, processing or secretion of 2 other major lysosomal proteases, cathepsin B and cathepsin D. Thus, Ras utilizes different effectors to mediate transformation and to deregulate cathepsin L expression and secretion in fibroblast and epithelial cells.
...
PMID:Enhanced cathepsin L expression is mediated by different Ras effector pathways in fibroblasts and epithelial cells. 1535 30

The human type III phosphatidylinositol 3-kinase, hVps34, converts phosphatidylinositol (PtdIns) to phosphatidylinositol 3-phosphate [PtdIns(3)P]. Studies using inhibitors of phosphatidylinositide 3-kinases have indicated that production of PtdIns(3)P is important for a variety of vesicle-mediated trafficking events, including endocytosis, sorting of receptors in multivesicular endosomes, and transport of lysosomal enzymes from the trans-Golgi network (TGN) to the endosomes and lysosomes. This study utilizes small interfering (si)RNA-mediated gene silencing to define the specific trafficking pathways in which hVps34 functions in human U-251 glioblastoma cells. Suppression of hVps34 expression reduced the cellular growth rate and caused a striking accumulation of large acidic phase-lucent vacuoles that contain lysosomal membrane proteins LAMP1 and LGP85. Analysis of these structures by electron microscopy suggests that they represent swollen late endosomes that have lost the capacity for inward vesiculation but retain the capacity to fuse with lysosomes. Morphological perturbation of the late endosome compartment was accompanied by a reduced rate of processing of the endosomal intermediate form of cathepsin D to the mature lysosomal form. There was also a reduction in the rate of epidermal growth factor receptor (EGFR) dephosphorylation and degradation following ligand stimulation, consistent with the retention of the EGFR on the limiting membranes of the enlarged late endosomes. By contrast, the suppression of hVps34 expression did not block trafficking of cathepsin D between the TGN and late endosomes, or endocytic uptake of fluid-phase markers, or association of a PtdIns(3)P-binding protein, EEA1, with early endosomes. LAMP1-positive vacuoles were depleted of PtdIns(3)P in the hVps34-knockdown cells, as judged by their inability to bind the PtdIns(3)P probe GFP-2xFYVE. By contrast, LAMP1-negative vesicles continued to bind GFP-2xFYVE in the knockdown cells. Overall, these findings indicate that hVps34 plays a major role in generating PtdIns(3)P for internal vesicle formation in multivesicular/late endosomes. The findings also unexpectedly suggest that other wortmannin-sensitive kinases and/or polyphosphoinositide phosphatases may be able to compensate for the loss of hVps34 and maintain PtdIns(3)P levels required for vesicular trafficking in the early endocytic pathway or the TGN.
...
PMID:Gene silencing reveals a specific function of hVps34 phosphatidylinositol 3-kinase in late versus early endosomes. 1652 86

The retromer complex mediates retrograde transport of transmembrane cargo from endosomes to the trans-Golgi network (TGN). Mammalian retromer is composed of a sorting nexin (SNX) dimer that binds to phosphatidylinositol 3-phosphate-enriched endosomal membranes and a vacuolar protein sorting (Vps) 26/29/35 trimer that participates in cargo recognition. The mammalian SNX dimer is necessary but not sufficient for recruitment of the Vps26/29/35 trimer to membranes. In this study, we demonstrate that the guanosine triphosphatase Rab7 contributes to this recruitment. The Vps26/29/35 trimer specifically binds to Rab7-guanosine triphosphate (GTP) and localizes to Rab7-containing endosomal domains. Interference with Rab7 function causes dissociation of the Vps26/29/35 trimer but not the SNX dimer from membranes. This blocks retrieval of mannose 6-phosphate receptors to the TGN and impairs cathepsin D sorting. Rab5-GTP does not bind to the Vps26/29/35 trimer, but perturbation of Rab5 function causes dissociation of both the SNX and Vps26/29/35 components from membranes through inhibition of a pathway involving phosphatidylinositol 3-kinase. These findings demonstrate that Rab5 and Rab7 act in concert to regulate retromer recruitment to endosomes.
...
PMID:Regulation of retromer recruitment to endosomes by sequential action of Rab5 and Rab7. 1898 Dec 34


1 2 Next >>

---