FIG. 8.

Transcriptional repression by HP1 proteins is mediated by the CSD. Plasmids expressing G4DBD fusions were cotransfected into NIH 3T3 cells with a fixed amount of a luciferase reporter carrying five copies of a synthetic GAL4 UAS in front of a minimal herpes simplex virus TK promoter. The amounts of transfected plasmids (0.5, 1.0, and 5.0 μg) expressing the G4DBD fusion proteins are shown as triangles on the y axis. The pcDNAlacZ plasmid was included to normalize transfection efficiencies. The portions of human HP1α, mouse M31, or KAP-1 fused to G4DBD are schematically drawn as in Fig. 2 and indicated by amino acid numbers in the construct names. In KAP-1, H denotes the HP1BD and Br denotes the bromodomain. Three independent transfections were carried out, and the average value for fold repression, along with the standard error, is given. Fold repression was calculated from the basal luciferase activity obtained from cells transfected without a G4DBD plasmid. A parallel series of transient transfections was carried out with a luciferase reporter lacking the GAL4 UAS sites, and this nonspecific fold repression was subtracted from the values obtained for transfections with the GAL4 UAS site-containing luciferase reporter. NLS, nuclear localization signal.