Figure 6.

Phactr4 colocalizes with β1 integrin at the tip of lamellipodia and regulates integrin signaling. (A,B) Four hours after wounding, MEFs were fixed and stained with anti-Phactr4 (green) and anti-ITGB1 (red) antibodies and Hoechst (blue). Phactr4 colocalizes with ITGB1 at the leading edge of lamellipodium in wild type and mutant. (C) MEFs were grown on laminin and then wounded for 1 h. Western blot analysis of total lysates with the indicated antibodies is shown. (D) Quantification of protein expression based on experiments such as shown in C showed 30% increase of pFAK in mutant cells. n = 6; (*) P < 0.05. (E) ITGB1 mRNA level by quantitative RT–PCR of mRNAs isolated from MEFs or FACS-sorted ENCCs shows no significant change in RNA levels. n = 3. (F) MEFs were plated on a fibronectin-coated coverslip for 15 min and 90 min, and stained with antibody against Phactr4 (green), focal adhesion marker Vinculin (red), and nuclei marker Hoechst (blue). Phactr4 is localized to mature, but not nascent, focal adhesions in both wild-type and mutant cells.