INTRACELLULAR TRAFFICKING MECHANISM OF TRANSPORTERS LOCATED ALONG THE CANALICULAR MEMBRANE OF HEPATOCYTES

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1. Construct of expression vectorWe made two types of vectors which expressed a fusion protein of human p-glycoprotein (human multidrug-resistance 1, hMDR1) and green fluorescent protein (GFP). One of them was made to express GFP-hMDR1, the other hMDR1-GFP.2. Western blottingThese vectors were transfected into HeLa cells and the cell lysate was served for Western blotting. We could detect a reaction of 220 kD using anti-GFP and C219 (anti-p-glycoprotein), suggesting that these vectors could intracellularly express fusion proteins of GFP and hMDR1.3. Localization of fusion proteinThe localization of the GFP-hMDR1 fusion protein was diffuse in various types of cells, which result was the same as GFP alone. On the other hand, hMDR1-GFP was found along the plasma membrane. Therefore, this vector, which expresses the hMDR1-GFP protein, would be very useful to investigate intracellular trafficking mechanism.