Altered DNA conformations in the gene regulatory region of torsionally-stressed SV40 DNA.
We used mung bean nuclease to probe the SV40 genome for DNA unwinding and unpairing. Cleavage occurred at a limited number of specific sites in supercoiled, but not relaxed DNA. The number and location of cleavage sites depended upon Mg2+ concentration. Without Mg2+, cutting occurred mainly in one early denaturation region located 3' to the t antigen gene and within the T antigen gene intron. With Mg2+, cleavage occurred at a number of alternative sites in the genome. Certain Mg2+ concentrations favored cleavage in the gene regulatory region. These cleavages were mapped at single nucleotide resolution and occurred in both transcriptional enhancers and upstream from the start of major late gene transcription. The cleavages occurred between 5 bp inverted repeat sequences, consistent with the recognition of unusually small cruciform structures.
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004262:DNA Restriction Enzymes; D004278:DNA, Superhelical / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics*; D005809:Genes, Regulator; D005814:Genes, Viral; D009690:Nucleic Acid Conformation; D013539:Simian virus 40 / Q000235:genetics*; D013314:Stress, Mechanical
D004278:DNA, Superhelical; D004279:DNA, Viral; D004262:DNA Restriction Enzymes