TISSUE-SPECIFIC FACTORS ADDITIVELY INCREASE THE PROBABILITY OF THE ALL-OR-NONE FORMATION OF A HYPERSENSITIVE SITE

DNase I-hypersensitive sites lack a canonical nucleosome and have bind ing sites for various transcription factors, To understand how the hyp ersensitivity is generated and maintained, we studied the chicken eryt hroid-specific beta A/epsilon globin gene enhancer, a region where bot h tissue-specific and ubiquitous transcription factors can bind, Const ructions containing mutations of this enhancer were stably introduced into a chicken erythroid cell line. We found that the hypersensitivity was determined primarily by the erythroid factors and that their bind ing additively increased the accessibility, The fraction of accessible sites in clonal cell lines was quantitated using restriction endonucl eases; these data implied that the formation of each hypersensitive si te was an all-or-none phenomenon. Use of DNase I and micrococcal nucle ase probes further indicated that the size of the hypersensitive site was influeuced by the binding of transcription factors which then dete rmined the length of the nucleosome-free gap. Our data are consistent with a model in which hypersensitive sites are generated stochasticall y: mutations that reduce the number of bound factors reduce the probab ility that these factors will prevail over a nucleosome; thus, the fra ction of sites in the population that are accessible is also diminishe d.