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 repair of DNA damage following ionizing radiation (IR). Although the GADD45 gene is transcriptionally up-regulated in response to IR, alterations in in vivo 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 regions of the gene were identified by in vivo DNase I hypersensitivity (HS) analysis. Chromatin structure and in vivo TF binding in these regions were subsequently monitored in both non-irradiated and irradiated human ML-1 cells. In non-irradiated cells expressing basal levels of GADD45, the gene exhibited a highly organized chromatin structure with distinctly positioned nucleosomes. Also identified in non-irradiated cells were DNA-protein interactions at octamer binding motifs and a CCAAT box in the promoter and at consensus binding sites for AP-1 and p53 within intron 3. Upon irradiation and a subsequent 15-fold increase in GADD45 mRNA levels, neither the chromatin structure nor the pattern of TF binding in key regulatory regions was altered. These results suggest that the GADD45 gene is poised for up-regulation and can be rapidly induced independent of gross changes in chromatin structure or TF binding.
Graunke
D M
DM
DNA Damage and Repair, Pennington Biomedical Research Center, Louisiana State University, 6400 Perkins Road, Room C2058-7, Baton Rouge, LA 70808, USA.
Fornace
A J
AJ
Pieper
R O
RO
D022762:CCAAT-Enhancer-Binding Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins / Q000378:metabolism; D003850:Deoxyribonuclease I; D051863:Host Cell Factor C1; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D007438:Introns; D009687:Nuclear Proteins / Q000378:metabolism; D009707:Nucleosomes; D050811:Octamer Transcription Factor-1; D050813:Octamer Transcription Factor-2; D016133:Polymerase Chain Reaction / Q000379:methods; D011506:Proteins / Q000235:genetics*; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D015854:Up-Regulation; D000097642:GADD45 Proteins
D022762:CCAAT-Enhancer-Binding Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; D003850:Deoxyribonuclease I; D051863:Host Cell Factor C1; D047908:Intracellular Signaling Peptides and Proteins; D009687:Nuclear Proteins; D009707:Nucleosomes; D050811:Octamer Transcription Factor-1; D050813:Octamer Transcription Factor-2; D011506:Proteins; D014157:Transcription Factors; D000097642:GADD45 Proteins; C497692:HCFC1 protein, human; C494165:POU2F1 protein, human; C058435:POU2F2 protein, human