FIG 6.

NGF does not induce translocation of TRPV2 to the plasma membrane in PC12 cells. (A) PC12 cells were cultured in the absence or presence of NGF (100 ng/ml) for 24 h. Surface proteins were biotinylated in intact cells at 37°C. Cells were then lysed, and biotinylated proteins were captured with streptavidin = agarose. Captured proteins were resolved by SDS-PAGE and detected by Western blotting with TRPV2 and Na/K ATPase antibodies. Na/K ATPase was used as a positive control. Surface proteins represent the biotinylated fraction, and input represents 5% of the total extracted protein. (B) Cell surface biotinylation experiment performed as described above for panel A. TRPV3 and Na/K ATPase antibodies were used for Western blot analysis. (C) PC12 cells were cultured in the absence or presence of NGF (100 ng/ml) for 24 h, followed by fixation and staining for TRPV2 (green) and Rab7 (red). Bar, 10 μm.