TAILIEUCHUNG - Báo cáo khoa học: Role of the plasma membrane leaflets in drug uptake and multidrug resistance

The present study aimed to investigate the role played by the leaflets of the plasma membrane in the uptake of drugs into cells and in their extrusion by P-glycoprotein and multidrug resistance-associated protein 1. Drug accumulation was monitored by fluorescence resonance energy transfer from trimethylammonium-diphenyl-hexatriene (TMA-DPH) located at the outer leaflet to a rhodamine analog. | Role of the plasma membrane leaflets in drug uptake and multidrug resistance Hagar Katzir Daniella Yeheskely-Hayon Ronit Regev and Gera D. Eytan Department of Biology The Technion - Israel Institute of Technology Haifa Israel Keywords MDR1 MRP1 multidrug resistance P-glycoprotein plasma membrane Correspondence G. D. Eytan Department of Biology The Technion - Israel Institute of Technology Haifa Israel Fax 972 4 822 5153 Tel 972 4 829 3406 E-mail eytan@ Website http These authors contributed equally to this work Received 5 November 2009 revised 13 December 2009 accepted 18 December 2009 doi The present study aimed to investigate the role played by the leaflets of the plasma membrane in the uptake of drugs into cells and in their extrusion by P-glycoprotein and multidrug resistance-associated protein 1. Drug accumulation was monitored by fluorescence resonance energy transfer from trimethylammonium-diphenyl-hexatriene TMA-DPH located at the outer leaflet to a rhodamine analog. Uptake of dye into cells whose mitochondria had been inactivated was displayed as two phases of TMA-DPH fluorescence quenching. The initial phase comprised a rapid drop in fluorescence that was neither affected by cooling the cells on ice nor by activity of mitochondria or ABC transporters. This phase reflects the association of dye with the outer leaflet of the plasma membrane. The subsequent phase of TMA-DPH fluorescence quenching occurred in drugsensitive cell lines with a half-life in the range 20-40 s. The second phase of fluorescence quenching was abolished by incubation of the cells on ice and was transiently inhibited in cells with active mitochondria. Thus the second phase of fluorescence quenching reflects the accumulation of dye in the cytoplasmic leaflet of the plasma membrane presumably as a result of flipflop of dye across the plasma membrane and slow diffusion from the inner leaflet into the cells. Whereas .

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