FIGURE 9.

PHF6 directly interacts with RBBP4 through its NoLS region. A, full-length PHF6 protein interacts with the recombinant RBBP4 protein. The pulled down RBBP4 protein band is marked with an asterisk. The band migrating higher than GST-PHF6FL is a nonspecific protein band during the protein purification. GST fusion full-length PHF6 protein is unstable and easily degradable. B, PHF6-ePHD2 domain does not interact with the RBBP4 protein. C, RBBP4 directly interacts with GST fusion PHF6 fragments (residues 145–207, lane 6; residues 152–171, lane 8). Gels were stained with Coomassie Blue. D, sequence alignment of the N-terminal residues of human Bcl11A/B, Sal1, and FOG1/2 with residues 152–171 of PHF6 protein. High similarity residues are shown on a red background. E, transcriptional repression by PHF6 protein is dependent on its NoLS region. WT, wild-type PHF6; ΔePHD2, absent ePHD2 domain from full-length PHF6; R342X, absent region 342–365 amino acid residues; ΔNoLS, absent region 157–171 amino acids from full-length PHF6. Efficiency of transfection was normalized to Renilla luciferase, as described under “Experimental Procedures.” The quantifications represent means of three independent experiments ± S.D.