MMSET mediates repression of a model target gene. (A) Constructs used in the reporter assay are fusions of the respective MMSET fragments to the DNA-binding domain of Gal4 (Gal-DBD, residues 1-147). The MMSET domains present in each fusion fragment are shown on the top of the figure. (B) Plasmids (125 ng) expressing the Gal4 DNA binding domain or Gal4 fusions of the MMSET domains were transfected into the Gal4-tk-luciferase reporter cell line in 24-well dishes. A representative triplicate experiment showing luciferase activity normalized to protein concentration is shown. (C) MMSET-II coimmunoprecipitates with MMSET-I and II. FLAG-tagged MMSET-II was cotransfected in HEK293T cells either with HA-MMSET-I or HA-MMSET-II. Total cell lysates (WCL) were immunoprecipitated with anti-FLAG antibody and subjected to immunoblotting with anti-HA antibody. Immunoprecipitation with nonspecific IgG is shown as a control. (D) Gal4-MMSET-I or Gal4-MMSET-II plasmids (125 ng) were transfected into the Gal4 reporter cell line along with 250 ng of MMSET-I or MMSET-II expression plasmid as indicated. A representative triplicate experiment showing luciferase activity normalized to protein concentration is shown. Error bars represent SD.