FIGURE 3:

Stabilization of CREB-H after mutation of the phosphodegron region. (a) Cells transfected with wt or mutants were treated −/+ cycloheximide 100 μg/ml (+Cyclo), samples were harvested at the intervals indicated (minutes) and then analyzed by Western blotting, and detection used fluorochrome-coupled secondary antibodies as described in Materials and Methods. (b) Relative levels of each of the mutant proteins were calculated. For each mutant, the abundance at the beginning of treatment is normalized to 100. Quantitative estimates of half-lives were obtained using GraphPad Prism software with curve fitting and one-phase decay parameters.