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. 2009 Apr 17;284(16):10831–10840. doi: 10.1074/jbc.M808683200

FIGURE 6.

FIGURE 6.

Analysis of Krox20-HCF interaction. This was investigated by co-immunoprecipitation of HA-tagged Krox20 with FLAG-tagged HCF-1 β-propeller. COS-7 cells were transfected with the expression plasmids indicated above each line. Cell lysates were subjected to immunoprecipitation (IP) with anti-HA antibody, and the precipitates were analyzed by Western blotting (WB) using anti-FLAG antibody. A 42 kDa band corresponding to the HCF-1 β-propeller is observed after co-transfection with wild type Krox20. Some nonspecific precipitation of the HCF-1 β-propeller occurs in the absence of Krox20 (first lane). Elimination of the N-terminal 190 amino acids of Krox20 reduces co-immunoprecipitation to background levels, whereas the internal deletion (Δ159-189) preserves the interaction. Analysis of the input (10%) indicates that the levels of HCF-1 β-propeller are identical in all cases. The Western blotting analysis of the Krox20 mutant proteins (the bands corresponding to the bona fide proteins are marked with asterisks) is shown underneath. Consistent results have been obtained in three independent experiments.