Fig. 7. Muscarinic agonist-induced spectrin redistribution depends on transient activation of ROCK. (A) Proteolytic conversion from the full-length (*) to C-terminally cleaved, active ROCK (**) is not significantly induced in M1-CHO cells stimulated with 5 µM Oxo-M for 5 minutes. (B) Transient expression of Flag-tagged kinase-dead (KD) ROCK-II (top row, red) diminishes Oxo-M-induced membrane blebbing and redistribution of YFP-II spectrin (top row, green). Images were taken 3 minutes after activation of M1 mAChRs. Flag-tagged constitutively active (CA) ROCK-II (bottom row, red) causes intense membrane blebbing accompanied with cell rounding in the absence of M1 mAChR activation. Images were taken 0 minutes after activation of M1 mAChRs. (C) Transient expression of Myc-tagged dominant-negative Rho (RhoA N19, top row, red) has little effects on Oxo-M-induced redistribution of YFP-II spectrin to the bleb end (top row, green, arrows). Images were taken 3 minutes after activation of M1 mAChRs. Overexpression of Myc-tagged constitutively active RhoA (Rho V14, bottom row, red) itself causes neither membrane blebbing nor cell rounding. Images were taken 0 minutes after activation of M1 mAChRs. Bar, 10 µm.