High-affinity nucleoprotein acceptor sites for the avian oviduct progesterone receptor (PR) have been enriched by a combination of nuclease digestion and centrifugation. These enriched binding elements exhibited markedly enhanced PR binding on a per mass DNA basis compared to chromatin (20-to 25-fold) or dehistonized chromatin (4-to 5-fold). Electrophoretic analysis of the nuclease-resistant DNA showed that there is a set of DNA fragments of 100-150 base pairs that are protected from digestion. Excessive digestion resulted in smaller DNA fragments and a loss of PR binding activity. The PR binding was saturable using a crude receptor preparation and displayed a competition with the same receptor preparation that was labeled with nonradioactive progesterone. The enhanced binding was also demonstrable using highly purified receptor preparations that exhibit two classes of binding sites both of which are of high affinity and saturable as assessed by Scatchard analyses. These two high-affinity classes of binding sites are shown to be competed by unlabeled purified PR. The nuclease resistance of these nucleoprotein acceptor sites from chromatin is a property similar to the nuclear matrix binding sites suggesting a relationship between these two classes of nuclear acceptor sites.Based on the presently accepted mechanism of steroid hormone action, the interaction of a steroid receptor-hormone complex with the nucleus is an important step in the induction of specific gene products. A great deal of effort has been invested by a number of laboratories in trying to determine the relevant receptor-nuclear interactions. Despite these efforts, the exact nature of the nuclear binding sites (acceptor sites) is not completely understood. Steroid receptors have been shown to bind to DNA (1), RNA (2), chromatin (3), and ribonucleoprotein particles (4).This laboratory has been concentrating on the specific interaction ofthe chicken oviduct progesterone receptor (PR) with nuclear acceptor sites in oviduct chromatin. In contrast to the receptor binding to pure DNA sequences alone (5, 6), the binding of the oviduct PR to subfractions of chromatin containing nonhistone protein-DNA complexes, termed nucleoacidic protein or NAP, has been shown to be receptor dependent and saturable, of high affinity (7)(8)(9)(10)(11), to be receptor specific (12), and to generate patterns of bindings similar to that measured in vivo (13)(14)(15)(16). Further, the capacity of the PR to bind to nuclear acceptor sites reflects the ability of progesterone to alter RNA polymerase activity in nuclear run-off experiments (17, t) and to specifically induce the avidin gene (18). A specific subset of nonhistone proteins (acceptor proteins) has been shown to be necessary for the generation of specific nucleoprotein acceptor sites (10,11,(19)(20)(21).In other organisms, similar nuclear acceptor sites composed of tightly bound (to DNA) nonhistone proteins have been reported for the estrogen receptor/chicken oviduct system (17), for systems involving the estrog...

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During primary estrogen stimulation of chick oviduct development, estrogen withdrawal, or secondary estrogen treatment, changes in the oviduct progesterone receptor (PR) occur. The presence of estrogen appears to regulate not only PR concentration but also its biochemical activity, i.e. its capacity to bind to nuclear acceptor sites and alter RNA synthesis. This study reports that estrogen regulates the nuclear binding capacity of the PR even more rapidly than previously reported in fully developed oviducts of chicks that have been injected daily for 4 weeks with diethylstilbestrol (DES). Further, the nuclear binding capacity of the PR correlates with the ability of progesterone (P) to induce avidin protein concentrations in the oviducts in vivo. The PR concentration in the oviducts increases 2-fold within 8 h of the last injection and the decreases to a minimal value by 24 h. Injection of [3H]P into the chicks shows that the in vivo nuclear localization of the steroid increases almost 4-fold at 8 h, followed by a similar decrease to minimal values by 24 h. Cell-free nuclear binding assays, using PR isolated at various times after the last DES injection and oviduct nucleoprotein complexes, indicate that the capacity of the receptors to bind to nuclear acceptor sites is regulated by the estrogen. The enhanced nuclear binding capacity of the isolated PR increases to maximal values by 12-14 h after the last estrogen treatment and then begins to decrease to minimal values by 24 h. Similarly, the ability of P to induce in vivo avidin protein concentrations and to alter general RNA synthesis in the oviducts is reduced by 70% (of the estrogen non-withdrawn chick levels) by 24 h after the last estrogen injection. These changes over the 24-h period after the last DES treatment are not due to changes in the serum DES concentrations. The following 10-day period of estrogen withdrawal reveals a cyclic decaying pattern in the capacity of the PR for nuclear binding. The P induction of avidin and alteration of RNA polymerase II activity, using nuclear run-off experiments, also show a similar cyclic decaying pattern. By 6 days of estrogen withdrawal, the PR is incapable of any nuclear binding, and P cannot induce avidin protein concentrations in the oviducts. Serum DES concentrations over this 10-day period display only a gradual decay.(ABSTRACT TRUNCATED AT 400 WORDS)

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