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Fig. 6 | BMC Molecular and Cell Biology

Fig. 6

From: DUF3669, a “domain of unknown function” within ZNF746 and ZNF777, oligomerizes and contributes to transcriptional repression

Fig. 6

Mutation to a conserved glutamate residue in ZNF746b KRAB-A potentiates its repression activity and TRIM28 interaction. a Multiple alignment of the KRAB-A amino acid residues of ZNF746b and three ZNF746b-KRAB-A mutants, R141L, G142E and RG/LE (positions in the complete protein indicated on top and mutated residues highlighted) along with the canonical sequence of ZNF10. The conserved canonical “MLE” motif is underscored in the ZNF10 sequence. b Comparison of transcriptional repression activities of wild type KRAB-AB of ZNF746b, and the three configurations with the indicated mutated residues in KRAB-A in HAP1 wild type (blue bars), SETDB1ko (orange bars), and TRIM28ko cell lines (grey bars). The KRAB domain of ZNF10 (Z10AB) is used as positive control. Results of dual luciferase assays (see Fig. 3) presenting mean repression factor values ± STDEV of at least six biological samples from three independent experiments. Asterisks indicate results of a two-tailed unpaired t-test (* p < 0.05; ** p < 0.01; *** p < 0.001; n.s. = not significant, i.e. p > 0.05) (c) Analysis of complex formation of GST-KRAB-AB fusion proteins with TRIM28 in HeLa cells using a co-immunoprecipitation/Western blot assay with endogenous TRIM28 as bait as described in Fig. 4. Grey block arrows point to endogenous TRIM28 signals in the lysate lanes (X), and white block arrows point to the GST/GST fusion bands in the lysate lanes (X)

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