FIG 7.

Interactions between CDK2 or importin α and HCTD or DCTD detected by GST pulldown assay. HEK293 cells in 100-mm dishes were transfected with 20 μg of the indicated GST-HCTD or GST-DCTD fusion constructs. Three days posttransfection, cells were harvested and GST pulldown assay was performed. (A) Copurified proteins were resolved by SDS-PAGE and detected by either Coomassie blue staining (bottom) or Western blotting using the CDK2-specific (top) or importin α-specific (middle) antibody. Arrowheads indicate the unidentified protein that was copurified with the GST-HCTD fusion proteins. (B and C) The relative levels of CDK2 and importin α associated with the HCTD or DCTD variants were normalized to those of the GST fusion proteins in the pulldown. The ratios of CDK2 or importin α to the GST fusion proteins were compared to that of the control (GST), set to 100. Data are presented as means and SEM from three independent experiments.