|
|
|
|
|
|
|
|
|||||
| Home Help Feedback Subscriptions Archive Search Table of Contents | |||||
|
Fig. 4. Forskolin induces degradation of cyclin D3 via the ubiquitin-proteasome pathway. (A) Reh cells were pretreated with the proteasome inhibitors MG-132 (10 µM) or LLnL (100 µM) for 30 minutes before addition of forskolin (100 µM). Cells were harvested at 2 hours after addition of forskolin and total lysates were analyzed with antibodies against cyclin D3 and actin. The blot shown is the representative of four independent experiments. Lower panel, data shown represent the mean ±s.e. of the four independent experiments. The results in each experiment were quantified using a densitometer and the densitometric values of cyclin D3 were normalized with those of actin. (B) Reh cells transfected with HA-Ub vector were exposed to MG-132 (10 µM) for 30 minutes before treatment with 100 µM forskolin for 2 hours. Cells were harvested, whole cell extracts prepared, and immunoprecipitated with anti-cyclin D3 antibodies. The recovered proteins were resolved on SDS-PAGE and then subjected to immunoblotting with anti-HA and anti-cyclin D3 antibodies consecutively. The blot shown is the representative of three independent experiments. IP, immunoprecipitation; NRS, non-immune rabbit serum.
|
