Fig. 4. EPHA2 regulates adherens junction stability via modulation of RhoA activity. (A) Activated Rho and Rac GTPases in MCF-10A, MCF-10A-EPHA2, or MCF-10A-C cells in response to ephrin-A1 stimulation were measured by GST-Rhotekin binding domain and GST-Pak binding domain pull-down assays, respectively. Total levels of Rho and Rac proteins were assay by western blot analysis. ^*P<0.05. (B-E) MCF-10A, MCF-10A-C, MCF-10A-EPHA2 or MCF-10A-EPHA2 treated with ROCK kinase inhibitor, Y27632, were subjected to Ca²⁺ depletion for 8 hours, followed by detection of E-cadherin. (F-I) MCF-10A and MCF-10A-C cells expressing a constitutively active Rho (Q63L) or control β-galactosidase (LacZ) were assayed for adherens junction stability by Ca²⁺-depletion assay. (J) The expression and activity of Q63L in control and MCF-10A-C cells were confirmed by Rhotekin pull-down assays and western blot analysis.