Figure 5

Representative immunoblots showing activation of RhoA in acetylcholine (ACh)-stimulated bronchi obtained from the sensitized control (Control) and repeatedly ovalbumin (OA)-challenged (Challenged) mice. Isolated bronchial tissues were incubated for 10 min in the absence (-) or presence (+) of 10^-3 M ACh (see Methods). Tissues were then rapidly lysed, GTP-bound active form of RhoA was pulled down with GST-tagged Rho binding domain of rhotekin, and RhoA was visualized by Western blotting. The respective blot of total RhoA in each sample is also shown. The GTP-bound RhoA in ACh-stimulated bronchi was markedly increased in the OA-challenged mice.