MCAF is a transcriptional regulator. (A) Effect of MCAF on promoter activities. The full length of MCAF (0 to 3 μg) and the insertless pcDNA3 (3 to 0 μg) were introduced into HeLa cells together with the luciferase reporter (1.0 μg) and pRL-CMV (0.02 μg). The luciferase activities in combination with mock vector (1.0 μg) were normalized to 10. (B) Inhibition of an Sp1-activated transcription by TRD of MBD1 and MCAF. Full-length MCAF and Δ2 mutant deficient in association with MBD1 (Fig. 2E) were expressed in D. melanogaster SL2 cells, together with GAL4-TRD of MBD1 and Sp1. GAL4 motif-containing reporter vector (1.0 μg), pPacSp1 (1.0 μg), pAc5.1-MCAF, pAc5.1-MCAF Δ2 and its insertless mock version (1.0 μg), and pAc5.1-GAL4-TRD of MBD1 (0.5 or 1.0 μg) and its mock version (0.5 or 0 μg) were utilized. The repression level in combination with the mock (1.0 μg) was normalized to 100 (black bars), and the relative luciferase activities were corrected by an internal control, pAc5.1-pRL (0.02 μg). (C) MCAF did not enhance an E2F1-activated transcription. Full-length MCAF (0 to 3 μg) and the insertless pcDNA3 (3 to 0 μg) were introduced into the cells together with the SNRPN or VHL luciferase reporter (1.0 μg) and E2F1-expressing vector (1.0 μg).