TY - JOUR
T1 - Detection of P-glycoprotein in the Golgi apparatus of drug-untreated human melanoma cells
AU - Molinari, Agnese
AU - Calcabrini, Annarica
AU - Meschini, Stefania
AU - Stringaro, Annarita
AU - Del Bufalo, Donatella
AU - Cianfriglia, Maurizio
AU - Arancia, Giuseppe
PY - 1998
Y1 - 1998
N2 - The intracellular location of the MDRI gene product, known as P- glycoprotein (P-gp), has been detected by flow cytometry in 3 stabilized human melanoma cell lines which had never undergone cytotoxic drug treatment and did not express P-gp on the plasma membrane. In addition, MDRI mRNA expression was revealed by RT-PCR in the same cell lines. Immunofluorescence microscopy, performed by using the same 2 monoclonal antibodies (MM4.17 and MRK-16) as employed in the flow-cytometric analysis, revealed the presence of P-gp intracytoplasmically, in a well-defined perinuclear region. Double immunofluorescence labelling and immunoelectron microscopy strongly suggested the location of the transporter molecule in the Golgi apparatus. The same observations have been obtained on a primary culture from a metastasis of human melanoma. Analysis of the expression of another membrane transport protein, the multidrug-resistance-related protein (MRPI), showed that it was present in the cytoplasm of all the melanoma cell lines examined. MRPI also showed Golgi-like localization. The study by laser scanning confocal microscopy on the intracellular localization of the anti-tumoral agent doxorubicin (DOX) during the drug-uptake and -efflux phases, indicated the Golgi apparatus as a preferential accumulation site for the anthracyclinic antibiotic. P-gp function modulators (verapamil and cyclosporin A) were able to modify DOX intracytoplasmic distribution and to increase drug intracellular concentration and cytotoxic effect in melanoma cells. On the contrary, MRPI modulators (probenecid and genistein) did not significantly influence either DOX efflux and distribution or the sensitivity of melanoma cells to the cytotoxic drug.
AB - The intracellular location of the MDRI gene product, known as P- glycoprotein (P-gp), has been detected by flow cytometry in 3 stabilized human melanoma cell lines which had never undergone cytotoxic drug treatment and did not express P-gp on the plasma membrane. In addition, MDRI mRNA expression was revealed by RT-PCR in the same cell lines. Immunofluorescence microscopy, performed by using the same 2 monoclonal antibodies (MM4.17 and MRK-16) as employed in the flow-cytometric analysis, revealed the presence of P-gp intracytoplasmically, in a well-defined perinuclear region. Double immunofluorescence labelling and immunoelectron microscopy strongly suggested the location of the transporter molecule in the Golgi apparatus. The same observations have been obtained on a primary culture from a metastasis of human melanoma. Analysis of the expression of another membrane transport protein, the multidrug-resistance-related protein (MRPI), showed that it was present in the cytoplasm of all the melanoma cell lines examined. MRPI also showed Golgi-like localization. The study by laser scanning confocal microscopy on the intracellular localization of the anti-tumoral agent doxorubicin (DOX) during the drug-uptake and -efflux phases, indicated the Golgi apparatus as a preferential accumulation site for the anthracyclinic antibiotic. P-gp function modulators (verapamil and cyclosporin A) were able to modify DOX intracytoplasmic distribution and to increase drug intracellular concentration and cytotoxic effect in melanoma cells. On the contrary, MRPI modulators (probenecid and genistein) did not significantly influence either DOX efflux and distribution or the sensitivity of melanoma cells to the cytotoxic drug.
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U2 - 10.1002/(SICI)1097-0215(19980316)75:6<885::AID-IJC11>3.0.CO;2-2
DO - 10.1002/(SICI)1097-0215(19980316)75:6<885::AID-IJC11>3.0.CO;2-2
M3 - Article
C2 - 9506534
AN - SCOPUS:0031941406
VL - 75
SP - 885
EP - 893
JO - International Journal of Cancer
JF - International Journal of Cancer
SN - 0020-7136
IS - 6
ER -