The effect of salt extraction on the structure of transcriptionally active genes; evidence for a DNAseI-sensitive structure which could be dependent on chromatin structure at levels higher than the 30 nm fibre.
The procedure developed by Lawson and Cole (Biochemistry, 1979, 18 2161-2166) for removing lysine-rich histones from nuclei at low pH also quantitatively extracts proteins HMG14 and 17. The effect of this low pH extraction on the DNAseI-sensitive structures of active genes in avian red blood cells has been investigated. No major perturbation of a developmentally regulated DNAseI hypersensitive site in the beta-globin domain and at the 5' end of the alpha D gene was seen. The overall DNAseI-sensitive conformation of the beta A-globin gene (relative to the ovalbumin gene) is minimally affected by pH3 salt extraction, but there is some loss of sensitivity of the alpha D gene. Removal of HMG proteins at neutral pH had no effect on the sensitivity of active genes in erythroid or fibroblast nuclei. These results, together with those carried out on DNAseI sensitivity and HMG binding to monomer nucleosomes, indicate that there is a major structural feature of active genes responsible for DNAseI-sensitivity which is independent of HMG proteins or nucleosome core particle structure but may be dependent on higher order chromatin structures.
D000818:Animals; D002642:Chick Embryo; D002843:Chromatin / Q000032:analysis*; D004247:DNA / Q000032:analysis*; D003850:Deoxyribonuclease I / Q000494:pharmacology*; D005914:Globins / Q000235:genetics*; D006609:High Mobility Group Proteins / Q000032:analysis / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D009693:Nucleic Acid Hybridization; D009707:Nucleosomes / Q000378:metabolism; D010957:Plasmids; D012965:Sodium Chloride / Q000494:pharmacology; D014158:Transcription, Genetic
D002843:Chromatin; D006609:High Mobility Group Proteins; D009707:Nucleosomes; D012965:Sodium Chloride; D005914:Globins; D004247:DNA; D003850:Deoxyribonuclease I