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. 1999 Jun;10(6):1851–1857. doi: 10.1091/mbc.10.6.1851

Figure 1.

Figure 1

A new assay to measure Rho activation. (A) Cos7 cells were transfected with expression vectors encoding myc-tagged versions of either wt, activated (L63), or inactive (N19) RhoA. In addition, two activated RhoA mutants with additional point mutations in the effector loop were used: L63A37 and L63G39. Cell lysates were incubated with GST-ROK, and GST-ROK–associating proteins were analyzed by Western blotting using 9E10 monoclonal anti-myc antibody. GST-ROK binds to wt and L63 Rho but not to N19, L63G39, or L63A37. (B) Cos cells were transfected with myc-tagged versions of wt, activated (V12/V14), or inactive (N17/N19) Rho, Rac, or Cdc42. Binding to GST-ROK was then assessed as in A. GST-ROK discriminates between activated and inactive Rho and Cdc42, but Rac is recovered independent of its activation state.