The subcellular localization of soluble and membrane-bound lysosomal enzymes in I-cell fibroblasts and in fibroblasts from control individuals

Dongen, J.M. van; Willemsen, Rob; Visser, W.J.; Hoogeveen, A.T.; Sips, Herman J.; Barranger, John A.; Tager, J. M.; Reuser, A. J. J.; Galjaard, H.

doi:10.1016/0304-3991(86)90043-4

Cited by 15 publications

(18 citation statements)

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“…Recently the major ex creted protein (MEP) from transformed mouse fibroblasts has been identified to be a high Mr inactive (acid activatable) precursor of cathepsin L [46][47][48][49][50], Lysosomal enzymes are processed and transported to the lysosomes via receptordependent pathways such as the mannose-6-phosphate pathway [51], This intracellular sorting has been extensively studied in fibro blasts for certain lysosomal enzymes not in cluding cathepsins B and L. Defects in these transport systems have been identified in fibroblasts from patients with lysosomal storage diseases [52] and in some murine tumor cell lines [53,54], In both cases the defect results in increased secretion of lyso somal enzymes. The defect may also result in plasma membrane association, e.g., in I-cell fibroblasts [3-hexosaminidase was found to be localized in small vesicles associated with the plasma membrane in addition to being secreted [55].…”

Section: Lysosomal Cysteine Proteinasesmentioning

confidence: 99%

Plasma Membrane-Associated Cysteine Proteinases in Human and Animal Tumors

Sloane¹,

Rozhin

Hatfield

et al. 1987

Pathobiology

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The ability of tumor cells to invade into and through normal tissue during the metastatic cascade has been attributed to tumor-associated degradative enzymes including proteinases of the metallo, serine and cysteine classes. Work from several laboratories has established that the cysteine proteinases cathepsins L and B are released from tumor cells, primarily as latent precursor forms. In addition, a cathepsin B-like cysteine proteinase has been shown to be associated with the plasma membrane fraction of several animal and human tumors. This form of the enzyme retains activity under physiologic (or pathologic) conditions including at neutral pH and in the presence of low Mr inhibitors. Since we have established that cathepsin B can degrade the basement membrane attachment glycoprotein laminin, we speculate that plasma membrane-associated cathepsin B may participate in focal dissolution of the basement membrane during tumor cell extravasation.

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Section: Lysosomal Cysteine Proteinasesmentioning

confidence: 99%

Plasma Membrane-Associated Cysteine Proteinases in Human and Animal Tumors

Sloane¹,

Rozhin

Hatfield

et al. 1987

Pathobiology

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“…Western blots were prepared from 12.5% polyacrylamide gels and probed as described. 20 After electrophoresis, proteins were transferred to nitrocellulose (Bio-Rad, Hercules, CA) and immunostaining was performed with the following antibodies: ␣85, anti-GLB1 antibody, previously characterized; 7,15 Alf1, anti-EBP antibody [the identification of EBP antigenic and hydrophilic peptides was used to obtain the following EBP epitope: NH2-VGSPSAQDEASPLS-COOH (91 to 104 amino acids) that was also previously used. 9 This anti-EBP peptide, conjugated with bovine serum albumin and immunized into rabbits by Igtech (Perdifumo, Italy), has been previously reported in a study on a galactosialidosis patient, who showed a reduced EBP amount and in a study on G M1 -gangliosidosis.…”

Section: Western Blotsmentioning

confidence: 99%

Primary and Secondary Elastin-Binding Protein Defect Leads to Impaired Elastogenesis in Fibroblasts from GM1-Gangliosidosis Patients

Caciotti

Donati

Bardelli

et al. 2005

The American Journal of Pathology

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G M1 -gangliosidosis is a lysosomal storage disorder caused by acid ␤-galactosidase deficiency. Aside from the lysosomal ␤-galactosidase enzyme, the ␤-galactosidase gene also encodes the elastin-binding protein (EBP), deficiency in which impairs elastogenesis. Using expression studies and Western blots of COS-1 cells, we identified and characterized four new and two known ␤-galactosidase gene mutations detected in G M1 -gangliosidosis patients with infantile, juvenile, or adult forms of disease. We then focused on impaired elastogenesis detected in fibroblasts from patients with infantile and juvenile disease. The juvenile patient showed connective-tissue abnormalities, unusual urinary keratan sulfate excretion, and an EBP reduction, despite mutations affecting only ␤-galactosidase. Because galactosugar-bearing moieties may alter EBP function and impair elastogenesis, we assessed infantile and juvenile patients for the source of altered elastogenesis. We confirmed that the infantile patient's impaired elastogenesis arose from a primary EBP defect, according to molecular analysis. We examined the juvenile's fibroblasts by immunohistochemistry, addition of keratanase, soluble/insoluble elastin assay, and radiolabeling of tropoelastin. These experiments revealed that the juvenile's impaired elastogenesis likely arose from secondary EBP deficiency caused by keratan sulfate accumulation. Thus, impaired elastogenesis in G M1 -gangliosidosis can arise from primary or secondary EBP defects in fibroblasts from infantile and juvenile patients, respectively. (Am J Pathol 2005, 167:1689 -1698)

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“…The next day, the cells were processed according to van Dongen et al (1985), using the antisera mentioned above and FITC-conjugated antirabbit IgG antibodies (Sigma).…”

Section: Immunofluorescence Western Blotting and Imm Unoprecipita Tionmentioning

confidence: 99%

Characterization of human lysosomal neuraminidase defines the molecular basis of the metabolic storage disorder sialidosis.

Bonten¹,

Spoel²,

Fornerod³

et al. 1996

Genes Dev.

278

240

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Neuraminidases (sialidases) have an essential role in the removal of terminal sialic acid residues from sialoglycoconjugates and are distributed widely in nature. The human lysosomal enzyme occurs in complex with 13-galactosidase and protective protein/cathepsin A (PPCA), and is deficient in two genetic disorders: sialidosis, caused by a structural defect in the neuraminidase gene, and galactosialidosis, in which the loss of neuraminidase activity is secondary to a deficiency of PPCA. We identified a full-length cDNA clone in the dbEST data base, of which the predicted amino acid sequence has extensive homology to other mammalian and bacterial neuraminidases, including the F(Y)RIP domain and "Asp-boxes." In situ hybridization localized the human neuraminidase gene to chromosome band 6p21, a region known to contain the HLA locus. Transient expression of the cDNA in deficient human fibroblasts showed that the enzyme is compartmentalized in lysosomes and restored neuraminidase activity in a PPCA-dependent manner. The authenticity of the cDNA was verified by the identification of three independent mutations in the open reading frame of the mRNA from clinically distinct sialidosis patients. Coexpression of the mutant cDNAs with PPCA failed to generate neuraminidase activity, confirming the inactivating effect of the mutations. These results establish the molecular basis of sialidosis in these patients, and clearly identify the cDNA-encoded protein as lysosomal neuraminidase.

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The subcellular localization of soluble and membrane-bound lysosomal enzymes in I-cell fibroblasts and in fibroblasts from control individuals

Cited by 15 publications

References 0 publications

Plasma Membrane-Associated Cysteine Proteinases in Human and Animal Tumors

Plasma Membrane-Associated Cysteine Proteinases in Human and Animal Tumors

Primary and Secondary Elastin-Binding Protein Defect Leads to Impaired Elastogenesis in Fibroblasts from GM1-Gangliosidosis Patients

Characterization of human lysosomal neuraminidase defines the molecular basis of the metabolic storage disorder sialidosis.

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