![Figure 5. DGK-α is required for the tethering of RCP vesicles at pseudopod tips. (A–C) A2780 (A and B) or vector and p53273H-expressing H1299 cells (C) were transfected with GFP-RCP in combination with nontargeting siRNA (siRNA control [si-con]), an siRNA targeting DGK-α (si-DGKα), or siRNA targeting DGK-α in combination with siRNA-resistant DGK-α (si-DGKα/DGKα(resc)). Cells were plated onto CDM, mounted onto the stage of a confocal microscope (FV1000), and incubated in the absence (−) or presence of 2.5 µM cRGDfV. Photobleaching was performed as described in the Materials and methods section. Stills from a video (Video 8) at the indicated time points are shown. (A) The arrows indicate the direction of cell migration, whereas the circles highlight the bleaching region. Recovery curves were generated using the FV10-ASW software (Olympus), and the immobile fraction (percentile) was extracted from these. Values are means ± SEM; ***, P < 0.0001. Bar, 10 µm.](https://cdn.rupress.org/rup/content_public/journal/jcb/196/2/10.1083_jcb.201109112/6/jcb_201109112_fig5.jpeg?Expires=1771704452&Signature=2b7OXmGq2-il970Sbn6L-jR31IwrCE6Z0gYpvpCmC1vB~zYhP2RSP12NtSjwgeOruBTRtA50RezY5lts8FDc~FFrVlkCH49HZniYyNzVYuXIcYiLgQXHrmmdwMRIUWKDIbZuW6GEnwXq5ZlEZ87MZFLFgyVfNi3GEPIuSKFJ-rhFDaxGAav35W0lZ2as6bGGhiWc70PR41wWLhoUztONdodG64~2Z6ME0T7Sq~YS~vP7mqcl0OcBg~RAoFx77JgeGRX9fCy6YjoBHT8vs9yGj0DrkwMAXmqN-zF3rFrMp0S8uegzMsJ4zp-4r5lD9GOlp0C8EuuCDI~-~Rvb7RGv3w__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
DGK-α is required for the tethering of RCP vesicles at pseudopod tips. (A–C) A2780 (A and B) or vector and p53273H-expressing H1299 cells (C) were transfected with GFP-RCP in combination with nontargeting siRNA (siRNA control [si-con]), an siRNA targeting DGK-α (si-DGKα), or siRNA targeting DGK-α in combination with siRNA-resistant DGK-α (si-DGKα/DGKα(resc)). Cells were plated onto CDM, mounted onto the stage of a confocal microscope (FV1000), and incubated in the absence (−) or presence of 2.5 µM cRGDfV. Photobleaching was performed as described in the Materials and methods section. Stills from a video (Video 8) at the indicated time points are shown. (A) The arrows indicate the direction of cell migration, whereas the circles highlight the bleaching region. Recovery curves were generated using the FV10-ASW software (Olympus), and the immobile fraction (percentile) was extracted from these. Values are means ± SEM; ***, P < 0.0001. Bar, 10 µm.
