Fig. 8

The levels of CDK2m3 mutant protein and vesicles decreased in response to 3-MA treatment. A Quantification of Cdk2m3 vesicles in the 3-MA treatment. The pPCdk2m3 stable transfectants were treated with 36 μM rapamycin in the absence or presence of 5.6 mM 3-MA for 24 h and then subjected to immunofluorescence analysis using anti-HA antibody for detection. Quantitation of vesicles was performed using Imaris software. *, p < 0.05. (n = 200–300 cells/condition). B The level of Cdk2m3 protein decreased in response to 3-MA treatment. The pPCdk2m3 stable transfectants were treated with 36 μM rapamycin in the absence or presence of 5.6 mM 3-MA for 24 h and then subjected to SDS-PAGE and Western blot analysis. The blot was probed with anti-HA, anti-MLF, and anti-RAN antibodies, respectively. SDS-PAGE with Coomassie Blue staining is included as a control for equal protein loading. The band intensity from triplicate Western blots was quantified using Image J. The CDK2m3 and MLF protein levels were normalized to the loading control (Coomassie Blue-stained proteins). The ratio of CDK2m3/MLF protein levels in 3-MA-treated sample to levels in untreated sample is shown and expressed as mean ± 95% confidence intervals. **, p < 0.01. ***, p < 0.001