Figure 6. DKK2-induced Cdc42 activation requires LRP6-mediated APC/Asef2 signaling.
(A–C) HUVECs stably expressing eGFP or DKK2 were transiently transfected with control, LRP5-specific, or LRP6-specific siRNA. After 60 hours, Cdc42 activity was measured (A). Cells were plated on Matrigel-coated plates at a density of 1.5 × 105 cells/well and incubated for 18 hours. Microphotographs were taken and capillary-like networks were quantified with Image-Pro Plus software (B and C). (D) Coimmunoprecipitation of APC with Asef2. Lysates were prepared from cells overexpressing eGFP or DKK2, immunoprecipitated with anti-APC antibody, and probed with anti-Asef2 or anti-APC antibodies. (E) eGFP- or DKK2-expressing cells were transiently transfected with control or LRP6-specific siRNA. After 60 hours, cell lysates were subjected to coimmunoprecipitation. (F–H) eGFP- or DKK2-expressing cells were transiently transfected with control, APC-specific, or Asef2-specific siRNA. After 60 hours, Cdc42 activity was measured (F). The cells were plated on Matrigel-coated plates at a density of 1.5 × 105 cells/well and incubated for 18 hours. Capillary-like networks were quantified with Image-Pro Plus software (G and H). Data represent mean ± SD. ***P < 0.001.