Document Detail

Wortmannin and Li+ specifically inhibit clathrin-independent endocytic internalization of bulk fluid.
MedLine Citation:
PMID:  8797088     Owner:  NLM     Status:  MEDLINE    
Incubation of a human fibrosarcoma cell line HT-1080 in Li(+)-containing medium inhibited internalization of a fluid marker, horseradish peroxidase (HRP), by more than 80%. The ion inhibited the activity enhanced by Ca2+ or phorbol 12-myristate 13-acetate. We also found that wortmannin (WT), a potent inhibitor of phosphoinositide (PI) 3-kinase (PI 3-k), inhibited the non-stimulated and the two stimulated types of endocytosis to the same extent as Li+. In contrast, neither WT nor Li+ influenced the early internalization of transferrin (Tfn), EGF or platelet-derived growth factor. Neither targeting to early endosomes nor recycling of the once-internalized Tfn was influenced. When the cytoplasmic pH was lowered by chasing cells that had been preincubated with 25 mM NH4Cl in an amiloride-containing Na(+)-free medium, more than 90% of internalization of Tfn in HT-1080 cells was inhibited, while that of HRP was reduced by only 35%. In contrast, WT reduced the uptake of HRP by KB cells by 34%, while 60% of the activity was inhibited by the treatment for cytoplasmic acidification. Comparison of other cells i.e., A-549 and a human diploid cell line Miyajima, indicated that cells showing higher sensitivity to WT were less sensitive to low cytoplasmic pH. These results suggest that, in all the cells studied, bulk fluid is internalized either via a clathrin-independent/PI 3-k-dependent route or via a clathrin-dependent/PI 3-k-independent one, though the ratio varied among them. We also found that internalization of a mAb directed toward the 116 (100)-kDa subunit of vacuolar ATPase [OSW2; Sato and Toyama (1994) J. Cell Biol. 127, 39-53] in the fluid phase was inhibited by WT, but the antibody was still internalized in a surface-bound form. Regardless of the treatment with WT, most of the antibody was transported to endosomes that were associated with Tfn receptor. These results suggest that both internalization routes are targeted to the same early endosomal compartments.
S B Sato; T Taguchi; S Yamashina; S Toyama
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Journal of biochemistry     Volume:  119     ISSN:  0021-924X     ISO Abbreviation:  J. Biochem.     Publication Date:  1996 May 
Date Detail:
Created Date:  1997-04-01     Completed Date:  1997-04-01     Revised Date:  2007-12-19    
Medline Journal Info:
Nlm Unique ID:  0376600     Medline TA:  J Biochem     Country:  JAPAN    
Other Details:
Languages:  eng     Pagination:  887-97     Citation Subset:  IM    
PRESTO, Research Development Corporation of Japan, Kyoto.
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MeSH Terms
1-Phosphatidylinositol 3-Kinase
Ammonium Chloride / pharmacology
Androstadienes / pharmacology*
Antibodies, Monoclonal / metabolism
Calcium / pharmacology
Cell Line
Clathrin / physiology*
Endocytosis / drug effects*,  physiology
Endosomes / physiology
Enzyme Inhibitors / pharmacology
Epidermal Growth Factor / metabolism
Horseradish Peroxidase / metabolism
Hydrogen-Ion Concentration
Lithium / pharmacology*
Microscopy, Electron
Phosphotransferases (Alcohol Group Acceptor) / antagonists & inhibitors*
Platelet-Derived Growth Factor / metabolism
Proton-Translocating ATPases / immunology,  metabolism
Tetradecanoylphorbol Acetate / pharmacology
Transferrin / metabolism
Tumor Cells, Cultured
Reg. No./Substance:
0/Androstadienes; 0/Antibodies, Monoclonal; 0/Clathrin; 0/Enzyme Inhibitors; 0/Platelet-Derived Growth Factor; 11096-37-0/Transferrin; 12125-02-9/Ammonium Chloride; 16561-29-8/Tetradecanoylphorbol Acetate; 19545-26-7/wortmannin; 62229-50-9/Epidermal Growth Factor; 7439-93-2/Lithium; 7440-70-2/Calcium; EC 1.11.1.-/Horseradish Peroxidase; EC 2.7.1.-/Phosphotransferases (Alcohol Group Acceptor); EC 3-Kinase; EC ATPases

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