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. 2013 Dec 23;203(6):1043–1061. doi: 10.1083/jcb.201306019

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© 2013 Shahbazi et al.

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Figure 1. — CLASP2 and p120 interact at AJs. (A) WT primary mKer were grown in low confluency to allow the formation of colonies. Cells were immunostained for CLASP2, ECad, and α-tubulin in the presence or absence of calcium (NC, normal calcium; LC, low calcium). (B) WT primary mKer immunostained for CLASP2 and p120 in the presence of calcium to induce formation of AJs. (C) Plot profile of CLASP2 and p120 fluorescence intensity at AJs in the area indicated in B. (D) CLASP2 was immunoprecipitated from mKer treated with calcium, and the immunoprecipitates (IP) were analyzed for CLASP2, ECad, and p120 by immunoblotting. Rabbit IgGs were used as a control. (E) Cell surface proteins from mKer treated with calcium were labeled with biotin (bio) and purified with a streptavidin column. CLASP2 was immunoprecipitated from the purified lysate of surface proteins, and CLASP2, p120, and ECad were analyzed by immunoblotting. Rabbit IgGs were used as a control of the immunoprecipitation, and a lysate of cells without biotin was used as a control of the purification. (F) WT mKer incubated with ECad-Fc–coated microspheres for 5 h in the presence of calcium. IgG-coated microspheres were used as a control. Cells were immunostained for CLASP2, p120, and ECad. Insets are magnifications of the boxed regions. WB, Western blot. Bars: (A and B, main images) 25 µm; (F) 10 µm; (A, B, and F, insets) 5 µm.