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. Author manuscript; available in PMC: 2009 Nov 20.
Published in final edited form as: Curr Biol. 2006 Dec 19;16(24):2466–2472. doi: 10.1016/j.cub.2006.10.056

Figure 3. Effect of RhoE on ROCK I kinase activity during genotoxic stress.

Figure 3

(A) Inhibition of RhoE expression enhances ROCK I kinase activity in wt-p53 expressing cells. U2OS cells stably expressing scrambled-RhoE-shRNA and RhoE-shRNA were treated with CPT for 12 or 24 hr. Cell lysates were subjected to immunoprecipitation (IP) with 2 μg of ROCK I antibody and western blotted to examine the levels of ROCK I and RhoE. The IP pallets were assayed for Rho kinase activity using MYPT1 as the substrate. The assay mixture was then subjected to SDS-PAGE. The gel was dried and autoradiographed. The band corresponding to MYPT1 was excised and quantified by scintillation counting. Error bars represent the standard deviation from three individual experiments.

(B) Effects of RhoE over-expression on ROCK I kinase activity in 293ET cells containing dysfunctional p53. 293ET cells were transfected with control and RhoE expression vector for 24 hr. The cells were then treated with 600 nM CPT for 24 and 32 hr, immunoprecipitated, and analyzed as described in (A).