Ultrastructural localization of steroid sulphatase in cultured human fibroblasts by immunocytochemistry: A comparative study with lysosomal enzymes and the mannose 6-phosphate receptor

1988 ◽  
Vol 20 (1) ◽  
pp. 41-51 ◽  
Author(s):  
R. Willemsen ◽  
M. Kroos ◽  
A. T. Hoogeveen ◽  
J. M. Van Dongen ◽  
G. Parenti ◽  
...  
Keyword(s):  
Comparative Study ◽  
Lysosomal Enzymes ◽  
Human Fibroblasts ◽  
Ultrastructural Localization ◽  
Steroid Sulphatase ◽  
Cultured Human Fibroblasts ◽  
Mannose 6 Phosphate

scholarly journals Recognition and receptor-mediated uptake of phosphorylated high mannose-type oligosaccharides by cultured human fibroblasts.

1983 ◽  
Vol 96 (3) ◽  
pp. 915-919 ◽  
Author(s):  
M Natowicz ◽  
D W Hallett ◽  
C Frier ◽  
M Chi ◽  
P H Schlesinger ◽  
...  
Keyword(s):  
Lysosomal Enzymes ◽  
Intracellular Transport ◽  
Specific Activity ◽  
Human Fibroblasts ◽  
Lysosomal Hydrolases ◽  
Minimal Structure ◽  
Cultured Human Fibroblasts ◽  
Specific Uptake ◽  
High Mannose ◽  
Mannose 6 Phosphate

The intracellular transport of newly synthesized lysosomal hydrolases to lysosomes requires the presence of one or more phosphorylated high mannose-type oligosaccharides per enzyme. A receptor that mediates mannose-6-PO4-specific uptake of lysosomal enzymes is expressed on the surface of fibroblasts and presumably accounts for the intracellular transport of newly synthesized enzymes to the lysosome. In this study, we examined the internalization of lysosomal enzyme-derived phosphorylated oligosaccharides by cultured human fibroblasts. Oligosaccharides of known specific activity bearing a single phosphate in monoester linkage were internalized with Kuptake of 3.2 X 10(-7) M, whereas oligosaccharides bearing two phosphates in monoester linkage were internalized with a Kuptake of 3.9 X 10(-8) M. Thus, phosphorylated high mannose-type oligosaccharides appear to be the minimal structure required for recognition and uptake by the fibroblast receptor. The finding that the Kuptake for monophosphorylated oligosaccharides is 100-fold less than the reported Ki for mannose-6-phosphate indicates that the fibroblast phosphomannosyl receptor contains a binding site that recognizes features of the oligosaccharide in addition to mannose-6-phosphate.

scholarly journals Inhibition by cyanate of the processing of lysosomal enzymes

1983 ◽  
Vol 210 (3) ◽  
pp. 795-802 ◽  
Author(s):  
A Hasilik ◽  
R Pohlmann ◽  
K von Figura
Keyword(s):  
Golgi Apparatus ◽  
Lysosomal Enzyme ◽  
Cathepsin D ◽  
Lysosomal Enzymes ◽  
Intracellular Transport ◽  
Human Fibroblasts ◽  
Cultured Human Fibroblasts ◽  
High Mannose ◽  
In Cells

In cultured human fibroblasts, maturation of the lysosomal enzymes beta-hexosaminidase and cathepsin D is inhibited by 10 mM-potassium cyanate. In cells treated with cyanate the two enzymes accumulate in precursor forms. The location of the accumulated precursor is probably non-lysosomal; in fractionation experiments the precursors separate from the bulk of the beta-hexosaminidase activity. The secretion of the precursor of cathepsin D, but not that of beta-hexosaminidase precursor, is enhanced in the presence of cyanate. The secreted cathepsin D, as well as that remaining within the cells, contains mostly high-mannose oligosaccharides cleavable with endo-beta-N-acetylglucosaminidase H. After removal of cyanate, the accumulated precursor forms of the lysosomal enzymes are largely released from the pretreated cells. It is concluded that cyanate interferes with the maturation of lysosomal-enzyme precursors by perturbing their intracellular transport. Most probably cyanate affects certain functions of the Golgi apparatus.

scholarly journals Effect of monensin on intracellular transport and receptor-mediated endocytosis of lysosomal enzymes

1984 ◽  
Vol 217 (3) ◽  
pp. 649-658 ◽  
Author(s):  
R Pohlmann ◽  
S Krüger ◽  
A Hasilik ◽  
K von Figura
Keyword(s):  
Golgi Apparatus ◽  
Lysosomal Enzyme ◽  
Cathepsin D ◽  
Lysosomal Enzymes ◽  
Intracellular Transport ◽  
Human Fibroblasts ◽  
Free Medium ◽  
Intracellular Enzyme ◽  
Cultured Human Fibroblasts ◽  
Monensin A

In cultured human fibroblasts we observed that monensin, a Na+/H+-exchanging ionophore, (i) inhibits mannose 6-phosphate-sensitive endocytosis of a lysosomal enzyme, (ii) enhances secretion of the precursor of cathepsin D, while inhibiting secretion of the precursors of beta-hexosaminidase, (iii) induces secretion of mature beta-hexosaminidase and mature cathepsin D, and (iv) inhibits carbohydrate processing in and proteolytic maturation of the precursors remaining within the cells; this last effect appears to be secondary to an inhibition of the transport of the precursors. If the treated cells are transferred to a monensin-free medium, about half of the accumulated precursors are secreted, and the intracellular enzyme is converted into the mature form. Monensin blocks formation of complex oligosaccharides in lysosomal enzymes. In the presence of monensin, total phosphorylation of glycoproteins is partially inhibited, whereas the secreted glycoproteins are enriched in the phosphorylated species. The suggested inhibition by monensin of the transport within the Golgi apparatus [Tartakoff (1980) Int. Rev. Exp. Pathol. 22, 227-250] may be the cause of some of the effects observed in the present study (iv). Other effects (i, ii) are rather explained by interference by monensin with the acidification in the lysosomal and prelysosomal compartments, which appears to be necessary for the transport of endocytosed and of newly synthesized lysosomal enzymes.

Comparative study on mannose 6-phosphate residue contents of recombinant lysosomal enzymes

2014 ◽  
Vol 111 (3) ◽  
pp. 369-373 ◽  
Author(s):  
Tadayasu Togawa ◽  
Masaru Takada ◽  
Yoshiaki Aizawa ◽  
Takahiro Tsukimura ◽  
Yasunori Chiba ◽  
...  
Keyword(s):  
Comparative Study ◽  
Lysosomal Enzymes ◽  
Mannose 6 Phosphate

scholarly journals Cathepsin D precursors in clathrin-coated organelles from human fibroblasts.

1985 ◽  
Vol 101 (3) ◽  
pp. 824-829 ◽  
Author(s):  
E Schulze-Lohoff ◽  
A Hasilik ◽  
K von Figura
Keyword(s):  
Staphylococcus Aureus ◽  
Human Brain ◽  
Residence Time ◽  
Cathepsin D ◽  
Lysosomal Enzymes ◽  
Human Fibroblasts ◽  
Coated Membranes ◽  
The Mean ◽  
Mannose 6 Phosphate ◽  
And Staphylococcus Aureus

Coated vesicles were isolated from metabolically labeled human fibroblasts with the aid of affinity-purified antibodies against human brain clathrin and Staphylococcus aureus cells. The material adsorbed to the S. aureus cells was enriched in clathrin. When the S. aureus cells bearing the immunoadsorbed material were treated with 0.5% saponin, extracts containing the precursor form of cathepsin D were obtained. The extraction of the precursor was promoted in the presence of mannose 6-phosphate. Material adsorbed to S. aureus cells coated with control immunoglobulins was nearly free of clathrin and contained a small amount of the cathepsin D precursor (less than 20% of that adsorbed with anti-clathrin antibodies). The extraction of this cathepsin D precursor was independent of mannose 6-phosphate and was complete after a brief exposure to saponin. The amount of cathepsin D precursor in coated membranes varied between 0.4 and 2.5% of total precursor. Analysis of pulse chase-labeled fibroblasts revealed that cathepsin D was only transiently associated with coated membranes. The mean residence time of cathepsin D precursor in coated membranes was estimated to be 2 min. These observations support the view that coated membranes participate in the transfer of precursor forms of endogenous lysosomal enzymes to lysosomes.

scholarly journals Evidence for lysosomal enzyme recognition by human fibroblasts via a phosphorylated carbohydrate moiety

1978 ◽  
Vol 170 (3) ◽  
pp. 643-650 ◽  
Author(s):  
Kurt Ullrich ◽  
Günther Mersmann ◽  
Ernst Weber ◽  
Kurt Von Figura
Keyword(s):  
Alkaline Phosphatase ◽  
Lysosomal Enzymes ◽  
General Scheme ◽  
Human Fibroblasts ◽  
High Uptake ◽  
Surface Receptors ◽  
Phosphatase Treatment ◽  
Mannose 6 Phosphate ◽  
Phosphorylated Sugars ◽  
Adsorptive Endocytosis

Adsorptive endocytosis of five different lysosomal enzymes from various human and non-human sources was susceptible to inhibition by mannose and l-fucose, methyl α-d-mannoside, α-anomeric p-nitrophenyl glycosides of mannose and l-fucose, mannose 6-phosphate and fructose 1-phosphate. A few exceptions from this general scheme were observed for particular enzymes, particularly for β-glucuronidase from human urine. The inhibition of α-N-acetylglucosaminidase endocytosis by mannose, p-nitrophenyl α-d-mannoside and mannose 6-phosphate was shown to be competitive. The loss of endocytosis after alkaline phosphatase treatment of lysosomal enzymes supports the hypothesis that the phosphorylated sugars compete with a phosphorylated carbohydrate on the enzymes for binding to the cell-surface receptors [Kaplan, Achord & Sly (1977) Proc. Natl. Acad. Sci. U.S.A.74, 2026–2030]. Endocytosis of ‘low-uptake’ forms of α-N-acetylglucosaminidase and α-mannosidase was likewise susceptible to inhibition by sugar phosphates and by alkaline phosphatase treatment, suggesting that ‘low-uptake’ forms are either contaminated with ‘high-uptake’ forms or are internalized via the same route as ‘high-uptake’ forms. The existence of an alternative route for adsorptive endocytosis of lysosomal enzymes is indicated by the unaffected adsorptive endocytosis of rat liver β-glucuronidase in the presence of phosphorylated sugars and after treatment with alkaline phosphatase.

scholarly journals Influence of chlorate on proteoglycan biosynthesis by cultured human fibroblasts.

1988 ◽  
Vol 263 (26) ◽  
pp. 12886-12892 ◽  
Author(s):  
H Greve ◽  
Z Cully ◽  
P Blumberg ◽  
H Kresse
Keyword(s):  
Human Fibroblasts ◽  
Cultured Human Fibroblasts

scholarly journals Fatty alcohol metabolism in cultured human fibroblasts. Evidence for a fatty alcohol cycle.

1987 ◽  
Vol 262 (36) ◽  
pp. 17412-17419 ◽  
Author(s):  
W B Rizzo ◽  
D A Craft ◽  
A L Dammann ◽  
M W Phillips
Keyword(s):  
Fatty Alcohol ◽  
Human Fibroblasts ◽  
Alcohol Metabolism ◽  
Cultured Human Fibroblasts
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