The nuclear binding sites (acceptor sites) for the progesterone-receptor complex (PR) in the avian oviduct have been characterized in this laboratory as involving specific nonhistone proteins, termed acceptor proteins. Recent studies in this laboratory have revealed that these proteins must be complexed to specific DNA sequences to generate the acceptor sites. The reconstitution of hen oviduct acceptor proteins to the DNAs from E. coli, salmon, plant (corn), and plasmid pBR 322 result in protein-DNA complexes with no PR binding. In contrast, the reconstitution of these proteins to hen DNA yields a protein-DNA complex with marked PR binding. Eighty percent of the DNA from partially deproteinized chromatin (NAP) can be degraded by DNAse I digestion with no loss of acceptor site binding activity. Apparently the remaining proteins protect certainregions of the DNA from nuclease digestion. The DNA in these protected regions shows a marked enrichment of repetitive DNA sequences. It is planned to further enrich the nuclease generated acceptor protein-DNA compelxes from the multitude of other protein-DNA complexes initially on the basis of differences in physical properties. A steroid receptor affinity resin and an antiacceptor protein antibody affinity resin will also be applied for the enrichment. The final enriched deoxyribonucleoprotein with acceptor activity will de deproteinized, 32P-nick translated, and used to probe a chicken genomic library for homologous sequences. Such sequences will be unambiguously characterized by restriction endonuclease and nucleoride sequence analysis and for acceptor activity by reconstituting with the acceptor protein and measuring PR binding. The characterization of the enriched acceptor-DNA sequences will also include the location of such sequences in and around the ovalbumin gene. It is hoped that these studies will identify the DNA sequences involved in the steroid regulation of gene expression, the role these sequences perform in this function, and the distribution of these sequences in the chick genome. The biological function of specific nonhistone proteins and DNA sequences and/or regulatory gene sequences may be achieved from these studies.
