Regulation of the mouse histone H2A.X gene promoter by the transcription factor E2F and CCAAT binding protein

We have molecularly cloned the genomic gene encoding the mouse histone variant H2A.X and characterized the promoter. The promoter region of the H2A.X gene was characterized by chloramphenicol acetyltransferase analysis using Balb/c 3T3 cells. Maximal promoter activity was found in the construct containing up to -282 base pairs H2A.X upstream region. Within this region, we found two sequences regulating the promoter activation; one was an E2F site and another was a CCAAT box. These sequences were also required for the DNA/protein binding activities. Thus, these activities corresponded to the promoter activities, implying that the promoter activity H2A.X gene was controlled by both the transcription factor E2F and H1TF2 through the E2F and CCAAT element. The CCAAT box binding activity was constitutive when cell cycle was progressed by release from G1 arrest, but transiently transfected chloramphenicol acetyltransferase activity slightly increased when cells entered S phase. Similarly, the level of the smallest form of E2F (free E2F) became higher when cells reentered the cell cycle, indicating that the free E2F was one capable of inducing the promoter activation. Thus, the free E2F and CCAAT DNA binding activity correlated with regulation of the promoter activity.