Figure 3
Figure 3. Enforced interaction of αVβ3ΔRGT with kindlin-2 rescues angiogenic sprouting by ECs. (A) Schematic of the inducible heterodimerizer system used here. (Top panel) The 2 fusion proteins employed: human αV-dimerizer C and dimerizer A-FLAG-human kindlin-2. β3+/+ or β3ΔRGT immortalized lung endothelial cells were transduced with lentiviruses encoding both of these fusion proteins and an shRNA specific for mouse αV. In theory, after addition of a cell-permeable heterodimerizer (AP21967), human αV-dimerizer C should associate with dimerizer A-human kindlin-2, thereby enforcing interaction between αVβ3 and kindlin-2. (B) Cytodex beads coated with either β3+/+ or β3ΔRGT endothelial cells expressing the dimerizer system described in panel A were incubated overnight at 37°C with 250 nM AP21967 (AP) or vehicle control. Then sprout formation by the cells was evaluated using the 3-dimensional fibrin gel bead assay. Data represent means ± SEM of 4 independent experiments; *P < .05; NS, not significant (paired Student t test).

Enforced interaction of αVβ3ΔRGT with kindlin-2 rescues angiogenic sprouting by ECs. (A) Schematic of the inducible heterodimerizer system used here. (Top panel) The 2 fusion proteins employed: human αV-dimerizer C and dimerizer A-FLAG-human kindlin-2. β3+/+ or β3ΔRGT immortalized lung endothelial cells were transduced with lentiviruses encoding both of these fusion proteins and an shRNA specific for mouse αV. In theory, after addition of a cell-permeable heterodimerizer (AP21967), human αV-dimerizer C should associate with dimerizer A-human kindlin-2, thereby enforcing interaction between αVβ3 and kindlin-2. (B) Cytodex beads coated with either β3+/+ or β3ΔRGT endothelial cells expressing the dimerizer system described in panel A were incubated overnight at 37°C with 250 nM AP21967 (AP) or vehicle control. Then sprout formation by the cells was evaluated using the 3-dimensional fibrin gel bead assay. Data represent means ± SEM of 4 independent experiments; *P < .05; NS, not significant (paired Student t test).

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