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. 2014 Feb 5;289(12):8390–8401. doi: 10.1074/jbc.M113.534024

FIGURE 2.

FIGURE 2.

TRIM32 interacts with the zinc finger domains of Glis2. A, FLAG-tagged TRIM32 (F.TRIM32) or F.BBS2 were transiently co-expressed in HEK 293T cells with V5-tagged Glis2 (V5.Glis2) or V5.NPHP5. Precipitation of F.TRIM32 immobilized V5.Glis2, but not V5.NPHP5. B, lysates of HEK 293T cells, transiently transfected with F.Glis2, were incubated with recombinant GST.TRIM32 or GST. F.Glis2 was pulled down by GST.TRIM32 but not by GST. Expression levels of GST.TRIM32 and GST were checked by Coomassie staining. C, truncations of Glis2, V5.Glis2 (amino acids 1–152), V5.Glis2 (amino acids 346–525), and V5.Glis2 (amino acids 146–359) were generated to map the interaction with TRIM32. F.TRIM32 interacted with the zinc finger-containing fragment of V5.Glis2, spanning amino acids 146–359. D, FLAG-tagged fragments of TRIM32 were co-expressed with V5.Glis2. Only the N-terminal fragment of TRIM32, spanning amino acids 1–256, interacted with Glis2. HC, antibody heavy chains; LC, antibody light chains; IP, immunoprecipitation; WB, Western blotting.