spacer gif spacer gif spacer gif spacer gif spacer gif
QUICK SEARCH:   [advanced]


spacer gif
     Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents    

Right arrow Help viewing high resolution images
Right arrow Return to article
(Downloading may take up to 30 seconds.
If the slide opens in your browser, select File -> Save As to save it.)
Click on image to view larger version.


Figure 7


Fig. 7. The effect of Kank protein on ß-catenin-dependent transcription and nuclear localization. (A) The effect of Kank protein on ß-catenin-dependent transcription examined by TOPFLASH reporter gene assay. SW480 cells were transfected with the reporter gene plasmid (TOPFLASH) together with vectors expressing wild-type and mutant Kank proteins. The pCMV Tag-2B cloning vector (Vector) was used to show the background level of ß-catenin-dependent transcription. The constructs for Kank-L-WT, NESm and NLSm are the same as shown in Fig. 4. The constructs for Kank-L with a function-less NLS1 (NLS1m) and Kank-S were also included. Luciferase activity was normalized against Renilla luciferase activity. The results were obtained from triplicate transfections. (B) The efficiency of Kank RNAi. The level of endogenous Kank protein was examined by western blotting (top) and quantified (bottom). (C) The requirement of endogenous Kank protein for ß-catenin-dependent transcription examined by TOPFLASH reporter gene assay. The reporter gene plasmid (TOPFLASH) with or without ß-catenin-S33Y or esiRNA against Kank were transfected into HEK293T cells. Luciferase activity was normalized against Renilla luciferase activity. The results were obtained from triplicate transfections. (D) Representative images of NIH3T3 cells transiently transfected with the constructs expressing ß-catenin-S33Y alone or ß-catenin-S33Y with FLAG-tagged wild-type Kank-L or FLAG-tagged Kank-L with mutant NLS (NLSm) or NES (NESm) motifs. ß-catenin was visualized with an FITC-conjugated antibody and Kank was visualized with a rhodamine-conjugated antibody. The nucleus was stained with DAPI. (E) Quantification of the Kank-positive cells according to the localization of ß-catenin protein. The asterisks indicate P<0.05 by t test. (F) Binding between FLAG-tagged Kank and ß-catenin. All constructs used were the same as those used in (D). IP, immunoprecipitation. Bar, 20 µm.





Right arrow Return to article