Detection of an altered DNA conformation at specific sites in chromatin and supercoiled DNA.
We show that bromoacetaldehyde, which reacts selectively at the N-1 and N-6 positions of unpaired adenine and at the N-3 and N-4 positions of unpaired cytosine residues reacts with chromosomal DNA in intact cells at probable regulatory sequences near active genes. A region of about 200 base pairs 5' to the chicken beta A-globin gene, which contains sites sensitive to nuclease S1, to several restriction endonucleases, and to very low levels of DNase I, also contains DNA structures that are preferentially sensitive to bromoacetaldehyde. These altered DNA structures are found at reproducible positions relative to the beta A-globin gene regardless of whether the bromoacetaldehyde is presented to intact erythrocytes, erythrocyte nuclei, or the beta A-globin gene itself carried in pBR322 as purified supercoiled DNA. The unpaired DNA 5' to the adult beta A-globin gene in adult erythrocytes is not detectable in embryonic erythrocytes that express embryonic beta-globin in contrast to adult beta A-globin. Our results suggest that well-defined regions of DNA with effectively unpaired bases occur in intact nuclei and that these structures may be important for specific recognition because they are tissue specific and are found at putative regulatory regions.
D000079:Acetaldehyde / Q000031:analogs & derivatives; D000818:Animals; D002467:Cell Nucleus / Q000032:analysis; D002645:Chickens; D002738:Chloroquine; D002843:Chromatin / Q000648:ultrastructure*; D004247:DNA / Q000097:blood*; D004262:DNA Restriction Enzymes; D004278:DNA, Superhelical / Q000097:blood*; D004912:Erythrocytes / Q000032:analysis; D007202:Indicators and Reagents; D009690:Nucleic Acid Conformation; D010957:Plasmids
D002843:Chromatin; D004278:DNA, Superhelical; D007202:Indicators and Reagents; C038235:bromoacetaldehyde; D002738:Chloroquine; D004247:DNA; D004262:DNA Restriction Enzymes; D000079:Acetaldehyde