FIG. 4.

Mtgr1 is a corepressor for TCF4. (A) Expression of β-catenin disrupts the interaction between Mtgr1 or Mtg16 and TCF4. GAL-Mtgr1 or GAL-Mtg16 and HA-TCF4 were transfected into Cos7 cells with or without a β-catenin expression plasmid. Immunoprecipitation (IP) with anti-GAL was followed by anti-HA (αHA) immunoblotting to detect TCF4 (top panel). Anti-GAL immunoblot analysis was used to detect the expression of GAL-Mtgr1 in the immune complexes, while anti-HA immunoblotting confirmed that equal amounts of TCF4 were present in whole-cell lysates (WCL). The bottom panel shows an immunoblot with anti-β-catenin (αβ-cat), which detects both endogenous and overexpressed protein. (B) Mtgr1 impairs β-catenin-induced activation of a TCF reporter plasmid in a dose-dependent fashion. NIH 3T3 cells were transfected with the TOP-FLASH or FOP-FLASH (containing mutant TCF sites) reporter and either TCF4 alone, TCF4 plus β-catenin, or TCF4 plus β-catenin and either 100 or 250 ng of an Mtgr1 expression plasmid, as indicated. A plasmid expressing cytomegalovirus-secreted alkaline phosphatase was used to normalize for transfection efficiency. Numbers below the graphs indicate the amount of DNA in micrograms.