Fig. 5. Cdc42 and RhoA activation in Caki-1 cells exposed to 1% O₂. Activation states of Cdc42 and RhoA proteins were measured by affinity precipitation with GST-PBD and GST-RBD, respectively. Cells were exposed to 1% O₂ from 0 to 6 hours, lysed and incubated with 20 µg fusion proteins. After pull-down precipitation, the GTP-bound forms of Cdc42 and RhoA proteins were analyzed by SDS-PAGE and western blot. Parallel experiments were performed to assess total amounts of Rho GTPases under these conditions. Cdc42 (A) and RhoA (B) were immunodetected with appropriate antibodies. Means ± s.e.m. from densitometric analysis of active GTPases for three independent experiments are shown. Data are expressed as the percentage of relative activity compared with values found in normoxic cells. Significant differences (P<0.05) from normoxia are indicated by asterisks (*).

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