FIG 1.

ME8^E2 is an E8-dependent transcriptional repressor. (A) C33A cells were transfected with 100 ng of pC18-Sp1-luc reporter plasmid, 0.5 ng pCMV-Gluc, and 30 ng of empty vector pSG5 or the indicated amounts of pSG mE8^E2, pSG mE8^E2-HA, pSG mE8^E2 d2-9, or pSG mE8^E2 d2-9-HA, plus empty vector (pSG5), to obtain equal amounts of plasmid DNA. Luciferase activities were determined 48 h posttransfection. Data are presented as ratios between firefly luciferase (Fluc) and Gaussia luciferase (Gluc) activities. Averages are derived from three independent experiments, and the error bars represent the standard error of the mean (SEM). Statistical significance was determined for mE8^E2 (black) and mE8^E2 d2-9 (blue) by one-way analysis of variance (ANOVA) and Holm-Sidak’s multiple-comparison test (ns, not significant; *, P < 0.05; **, P < 0.01). On the right, a schematic structure of the pC18-SP1-luc plasmid is shown. (B) Sequence alignment of E8 sequences. Conserved residues are shown in red. (C) Immunoblot analysis of whole-cell extracts derived from C33A cells transfected with the empty vector (V), pSG mE8^E2-HA (wt), or pSG mE8^E2 d2-9-HA (mt). E8^E2 proteins were detected with an anti-hemagglutinin (HA) antibody, and α-tubulin served as a reference protein (ref).