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. 2000 Sep;20(17):6612–6625. doi: 10.1128/mcb.20.17.6612-6625.2000

FIG. 4.

FIG. 4

Effect of MEKK-1 signaling on transcriptional repression and T3R interactions with SMRT. (A) Inhibition by MEKK-1 of the two-hybrid interaction between SMRT and T3Rα. The effect of various signal transducers on the two-hybrid interaction between SMRT and T3Rα was tested as described in the legend to Fig. 1C by comparing an empty expression vector (None), a v-ErbB expression plasmid (v-ErbB), a full-length MEKK-1 expression plasmid (MEKK1), or a dominant negative MEKK-1 (residues 817 to 1493) expression plasmid (MEKK1 DN), as indicated below the panel. (B) Effects of MEKK-1 and MEK-1 on T3Rα-mediated repression of a TRE-luc reporter. The effect of different signal transducers on the ability of T3Rα to repress a TRE-driven promoter was tested as described in the legend to Fig. 1A. Expression plasmids for full-length MEKK-1, dominant negative MEKK-1, MEK-1, or dominant negative MEK-1 (MEK1 DN) were compared to an equivalent empty expression vector (None), as indicated below the panel. TRE reporter activity was assayed only in the absence of T3. (Inset) Effect of dominant negative MEKK-1 on the ability of v-ErbB to counteract T3R repression. Various combinations of expression plasmids for T3Rα, v-ErbB, and dominant negative MEKK-1 were transfected into CV-1 cells, as indicated below the inset panel, and the relative luciferase activity was determined. Fold repression was calculated relative to the basal levels of reporter gene expression in the absence of T3Rα. (C) Effects of EGF receptor and MEKK-1 signaling on transcriptional repression by PLZF. Various amounts of a GAL4DBD-PLZF fusion construct were introduced into CV-1 cells together with the GAL4–17-mer luciferase reporter and expression plasmids for v-ErbB or full-length MEKK-1. Relative luciferase activity was determined by normalization to an internal pCH110 β-galactosidase control; fold repression was calculated relative to the basal level of luciferase expression observed in the absence of the PLZF construct. The average and standard deviation of duplicate experiments are presented.