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. Author manuscript; available in PMC: 2008 Jun 2.
Published in final edited form as: Oncogene. 2005 Aug 25;24(36):5561–5575. doi: 10.1038/sj.onc.1208727

Figure 6.

Figure 6

HOX11 modulates multiple transcriptional networks. (A) Relative luciferase activity from cotransfection of the reporter plasmids pCRE-Luc containing multiple copies of the CRE-binding sequence (CRE), pISRE-Luc containing five copies of the ISRE-binding sequence (ISRE), and pAP1(PMA)-TA-Luc containing six tandem copies of the sequence TAATGAC resembling a HOX11-binding site [TAAT (Owens et al., 2003)] into NIH3T3 cells with HOX11 expression vectors: pcDNA3-HOX11 expressing wild-type HOX11 (HOX11); pcDNA3-HOX11 H3d, a DNA-binding deficient mutant of HOX11 that contains a deletion (from amino acid 246 to amino acid 251) of the third helix of the homeodomain (H3d); and pcDNA3-HOX11 Lys55Gln, a mutant of HOX11 in which lysine 55 in the third helix of the homeodomain, which forms a salt bridge between the protein and the phosphate groups of DNA and which may be involved in interactions with CBP/p300 coregulators, has been changed to glutamine (K55Q) (Owens et al., 2003; Shen et al., 2001). Controls included the empty pcDNA3 plasmid (Mock) as well as the pTAL-Luc luciferase reporter plasmid containing a minimum promoter (not shown). The data presented are average of at least 3 independent transfections. (B) CPB protein levels are reduced in Jurkat cells expressing MSCVhyg-HOX11 (HOX11) compared with Jurkat cells expressing the empty MSCVhyg vector (Mock). CPB was immunoprecipitated from whole cell extracts with a rabbit anti-CBP polyclonal antibody (anti-CBP NT) followed by immunoblotting with a mouse anti-CBP monoclonal antibody (anti-CBP C1). (C) Repression of transcription mediated by TCF/LEF family members was examined by transiently cotransfecting a plasmid expressing a stabilized β-catenin [ΔN87βcat (Gat et al., 1998)] from the human elongation factor 1 promoter (A. Ramezani and R.G.H., unpublished data) and the TOPflash luciferase reporter plasmid containing TCF/LEF binding sites (TCF) into 293T cells together with the HOX11 expression plasmids described in (A). The FOPflash luciferase reporter plasmid containing mutated TCF/LEF binding sites (mTCF) served as a negative control.