Figure 3
Effects of EPAC1 Activators on EPAC1 and Rap1 Activation. (A) U2OS cells transfected with EPAC1 were preincubated for 10 min with 100 μM of EPAC1 activators, PWO577, SY007 or D-007. Active EPAC1 was then isolated from cells by immunoprecipitation with the activation selective EPAC1 (DE3) antibody and detected by immunoblotting with the same antibody, as described in “Materials and methods”. Significant increases in active EPAC1 levels relative to D-007-treated cells are indicated; **p < 0.01 and ***p < 0.001 or ns, for no significant difference (n = 5–7). (B) U2OS cells transfected with EPAC1 were preincubated for 10 min with 100 μM of EPAC1 activators, PWO577, SY007 or D-007. Active Rap1 was then isolated from cell lysates, immunoblotted and compared to total Rap1 levels, as described in “Materials and methods”. Data are presented as means ± SEM (n = 5–7). Significant increases in active Rap1 levels in comparison to D-007-treated cells treated cells are indicated, p < 0.001 (n = 5–7).
