Presetting of chromatin structure and transcription factor binding poise the human GADD45 gene for rapid transcriptional up-regulation

GADD45 has been suggested to coordinate cell cycle regulation with the repa ir of DNA damage following ionizing radiation (IR). Although the GADD45 gen e is transcriptionally up-regulated in response to IR, alterations in in vi vo transcription factor (TF) binding or chromatin structure associated with up-regulation have not been defined, To understand how chromatin structure might influence TF binding and GADD45 up-regulation, key regulatory region s of the gene were identified by in vivo DNase I hypersensitivity (HS) anal ysis. Chromatin structure and in vivo TF binding in these regions were subs equently monitored in both non-irradiated and irradiated human ML-1 cells. In non-irradiated cells expressing bas;al levels of GADD45, the gene exhibi ted a highly organized chromatin structure with distinctly positioned nucle osomes, Also identified in non-irradiated cells were DNA-protein interactio ns at octamer binding motifs and a CCAAT box in the promoter and at consens us binding sites for AP-1 and p53 within intron 3, Upon irradiation and a s ubsequent 15-fold increase in GADD45 mRNA levels, neither the chromatin str ucture nor the pattern of TF binding in key regulatory regions was altered. These results suggest that the GADD45 gene is poised far up-regulation and can be rapidly induced independent of gross changes in chromatin structure or TF binding.