Transport of glutathione conjugates into secretory vesicles is mediated by the multidrug-resistance protein 1.
Intracellular glutathione-conjugate transport was evaluated in the human small cell lung carcinoma cell line GLC4 with low multidrug resistance protein (MRP1) expression and its 300x doxorubicin-resistant, MRP1-over-expressing, GLC4-Adr subline. Transport of non-toxic concentrations of monochlorobimane and 5-chloro-methyl fluorescein diacetate was evaluated using fluorescence microscopy. After exposure to these compounds, fluorescence was observed especially in intracellular vesicles in GLC4-Adr. Immunotransmission electron microscopy showed that MRP1 was present in the vesicle membranes and plasma membrane, while inside the vesicles the glutathione conjugate of 1-chloro-2,4-dinitrobenzene could be detected. Experiments with brefeldin A, which induces arrest in vesicle release from the Golgi complex, indicated that these vesicles may originate from the trans-Golgi network. In GLC4-Adr cells, doxorubicin also was transported in vesicles, with an arrest in vesicle release from the Golgi complex. Our results indicate that MRP1 functions as a glutathione-conjugate transporter not only at the plasma membrane but also in intracellular secretory vesicles.
Van Luyn
M J
MJ
Department of Cell Biology, University Hospital Groningen, The Netherlands.
Müller
M
M
Renes
J
J
Meijer
C
C
Scheper
R J
RJ
Nienhuis
E F
EF
Mulder
N H
NH
Jansen
P L
PL
De Vries
E G
EG
D018528:ATP-Binding Cassette Transporters / Q000502:physiology*; D015415:Biomarkers; D018288:Carcinoma, Small Cell / Q000378:metabolism*; D002462:Cell Membrane; D003594:Cytoplasmic Granules / Q000378:metabolism*; D018432:Drug Resistance, Multiple; D005452:Fluoresceins / Q000378:metabolism; D005456:Fluorescent Dyes; D005978:Glutathione / Q000378:metabolism*; D006801:Humans; D008175:Lung Neoplasms / Q000378:metabolism*; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D009363:Neoplasm Proteins / Q000494:pharmacology; D011720:Pyrazoles / Q000378:metabolism; D014407:Tumor Cells, Cultured
D018528:ATP-Binding Cassette Transporters; D015415:Biomarkers; D005452:Fluoresceins; D005456:Fluorescent Dyes; D005978:Glutathione; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D009363:Neoplasm Proteins; D011720:Pyrazoles; C069306:5-chloromethylfluorescein; C059597:monochlorobimane