Early-phase redistribution of the cation-independent mannose 6-phosphate receptor by U18666A treatment in HeLa cells.
It has been shown that the treatment with 3beta-[2-(diethylamino)ethoxy] androst-5-en-17-one (U18666A) causes the accumulation of cholesterol and the cation-independent mannose 6-phosphate receptor (CIMPR) in late endosomal/lysosomal compartments in BHK cells. The present study reports on a study of the effect of U18666A on CIMPR distribution in more detail in HeLa cells. When cells were treated with U18666A for 20 h, the intense perinuclear signal for CIMPR corresponding to the trans-Golgi network (TGN) disappeared and lamp1-negative punctate signals, scattered in the perinuclear region were detected. CIMPR then began to accumulate in lamp1-positive compartments 48 h after addition of the drug. Double immunofluorescence microscopy showed that U18666A-induced mannose 6-phosphate receptor-containing compartments (U-MPRCs), which were formed in the early phase of the redistribution, contained no marker for the TGN, late endosomes or lysosomes. Approximately half of the structures contained transferrin that had been internalized for 20 min, and cathepsin D, the majority of which appeared to be its precursor form. Immunoelectron-microscopic analysis revealed that U-MPRCs are composed of multivesicular bodies, irregularly shaped structures, and vesicular structures adjacent to the multivesicular bodies. These results suggest that U18666A treatment primarily suppresses the CIMPR transport pathways to late endosomes and from transferrin-containing endosomes, both of which may be dependent on cholesterol function.
Department of Conservative Dentistry, Osaka University Faculty of Dentistry, 1-8 Yamadaoka, Suita, Osaka, Japan.
Yuji
Y
Tomiyama
Satoshi
S
Waguri
Shiro
S
Kanamori
Satoshi
S
Kametaka
Masaki
M
Wakasugi
Masahiro
M
Shibata
Shigeyuki
S
Ebisu
Yasuo
Y
Uchiyama
D000736:Androstenes / Q000494:pharmacology*; D000924:Anticholesteremic Agents / Q000494:pharmacology*; D002402:Cathepsin D / Q000378:metabolism; D002412:Cations / Q000378:metabolism; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D011992:Endosomes / Q000378:metabolism; D006367:HeLa Cells / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D016253:Microscopy, Immunoelectron; D017527:Receptor, IGF Type 2 / Q000378:metabolism*; D014168:Transferrin / Q000378:metabolism
D000736:Androstenes; D000924:Anticholesteremic Agents; D002412:Cations; D017527:Receptor, IGF Type 2; D014168:Transferrin; C006261:3-beta-(2-(diethylamino)ethoxy)androst-5-en-17-one; D002402:Cathepsin D