EPLIN mediates linkage of the cadherin–catenin complex to F-actin and stabilizes the circumferential actin belt

Abe and Takeichi. 10.1073/pnas.0710504105.

Supporting Information

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SI Figure 6
SI Figure 7
SI Figure 8

Fig. 6. a-Catenin dependent interaction of E-cadherin with EPLIN. DLD-1 cells were transfected with control siRNA or siRNA against aE-catenin (CTNNA1). Four days after transfection, the cells were lysed and subjected to immunoprecipitation by using anti-E-cadherin antibodies. The precipitates and 2% of the lysate (Input) were subjected to Western blotting to detect each protein. EPLINb coprecipitated with E-cadherin was greatly reduced in aE-catenin-depleted cells. Detection of coprecipitated EPLINa was not sensitive enough at this time of experiments. Arrowheads indicate the band of each protein. Lower bands in the E-cadherin blots are its degradation products.

Fig. 7. Recombinant proteins used in this study. (A) CBB staining of recombinant proteins used in this study. GST-fused protein were expressed in E. coli and purified. To obtain proteins without the GST tag, the tag was removed as described in Materials and Methods. (B) Deletion series of a-catenin and EPLIN used for the experiments shown in Fig. 1C.

Fig. 8. siRNA-mediated knockdown of EPLIN and a-catenin in DLD-1 cells. DLD-1 cells were transfected with control siRNA or siRNA against EPLIN or aE-catenin (CTNNA1). Four days after transfection, the cells were lysed and subjected to Western blotting to detect each protein. a-Tubulin was blotted as a loading control. More than 90% of the targeted proteins were removed in each culture at 3-4 days after siRNA transfection.

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