Fig. 2.

PINCH^wt-Flag and PINCH^Q38A-Flag transgenes rescue the lethality of the stck null mutant. (A) Adult rescued flies do not have wing blisters or any other overt phenotypes. PINCH^wt-Flag and PINCH^Q38A-Flag adult flies appear similar to each other, and to w¹¹¹⁸ controls. (B) Western blots of adult rescued flies probed with anti-PINCH antibody show a lack of endogenous PINCH and only show the higher migrating PINCH-Flag species. Three different insertion lines are shown for both PINCH^wt-Flag and PINCH^Q38A-Flag on the X, 2nd and 3rd chromosomes. Anti-Lamin antibody is used as a loading control. (C) PINCH^wt-Flag and PINCH^Q38A-Flag transgenic protein levels are similar to levels of endogenous PINCH in wild-type flies. Bars represent the median ± range of transgenic PINCH from three different chromosomal insertions for both PINCH^wt-Flag and PINCH^Q38A-Flag rescued flies. (D) PINCH^wt-Flag transgenes rescue 84–120% and PINCH^Q38A-Flag transgenes rescue 88–107%. Bars represent the mean ± s.e.m. of the percentage rescued flies counted for the number of crosses set. C denotes the number of times the rescue cross was set. n is the total number of flies counted across all crosses. (E) Anti-Flag immunoprecipitations confirm disruption of the PINCH-ILK interaction in adult PINCH^Q38A-Flag rescued flies. PINCH^wt-Flag complexes contain both ILK and RSU-1, whereas PINCH^Q38A-Flag complexes contain only RSU-1. w¹¹¹⁸ lysate was used as a control to assess nonspecific binding. All proteins were present in the starting material.