FIGURE 10.
Treatment with cyclosporine A decreases the association of P-gp with Hsp70. Lysates from 2 × 106 cells transduced with cysless-WT and double mutant P-gp, grown in the absence and presence of 6.25 μm CsA for 18 h, were immunoprecipitated using rabbit polyclonal Hsp70 antibody (ab79852, Abcam), and the immunoprecipitate was probed with anti-P-gp antibody C219 at 1:2000 dilution. Lane 1, cysless-WT; lane 2, D164C/D805C; lane 3, cysless-WT (CsA); lane 4, D164C/D805C (CsA). Lanes 1 and 2 show associated P-gp when cells were grown in the absence of CsA, whereas lanes 3 and 4 depict these levels in the presence of CsA. The immunoblot on the right was probed with C219 antibody to show the level of P-gp in cell lysates (3 μg/lane), which were used for immunoprecipitation with Hsp70 antibody.