Evidence for two kinds of chromatin binding sites for the estradiol-receptor complex

Abstract

Evidence for two types of chromatin binding sites for the estradiol-receptor complex was found in target nuclei using micrococcal nuclease digestion. Nuclei from rat uteri, which had been incubated for 1 hr at 37°C in Dulbecco's modified Eagle's medium containing 10 −8 M 3H-estradiol, produced monomer (nu 1) and short oligomer nubodies which contained significantly higher specific activities of 3H-estradiol/nu-body than the bulk chromatin. The specific activities decreased from nu 1 to nu 2 to nu 3, and more extensive digestion produced nu-bodies with lower specific activities. 3H-estradiol-containing nu-bodies sedimented slightly faster than bulk particles. The bound estradiol could be released from the nu-bodies with 400 mM KCl, producing a 5.3S estradiol-receptor complex similar to the “activated” form of receptor released by salt from uterine nuclei of steroid-treated animals. Concomitant with the appearance of 3H-estradiol-enriched nu-bodies, micrococcal nuclease also released from the chromatin an estradiol-receptor complex which sedimented at 6.9S in low salt and 5.3S in high salt. This moiety may be bound to a small fragment of DNA since receptor released from chromatin by DNAase I sedimented at 6.0S rather than at 6.9S. It is possible that the binding of estradiol to chromatin, which is known to turn on specific gene transcription in vivo, induces or reflects a local conformational alteration which we detect as an enhanced sensitivity to nuclease, and which is recognized intracellularly as a start signal for transcription.