Progesterone receptor interaction in the 5'-flanking regulatory region of the ovalbumin gene.
Highly purified hen oviduct progesterone receptor A subunit has been found to interact preferentially with the 5'-flanking region of the chicken ovalbumin gene. Saturation kinetics showed that 2-fold less receptor was required for half-maximal binding to ovalbumin gene flanking sequences than to plasmid pBR322 sequences. Studies of selective binding were facilitated by a preferential adsorption assay involving gel electrophoretic analysis of [32P]DNA restriction fragments eluted from receptor-DNA complexes isolated on nitrocellulose filters. Approximately 10-fold difference in binding was observed at 37 degrees C for a 1.7 Kb ovalbumin DNA insert compared to binding of receptor to a 3.6 Kb plasmid pBR322 fragment. Selective receptor A binding was dramatically reduced when assayed at 4 degrees C. In addition to temperature dependence, preferential binding required the presence of glycerol or dimethyl sulfoxide during binding. Divalent cations and ionic strength had no apparent effect on the relative sequence selectivity but did alter the total DNA binding observed. Binding data from a variety of different restriction digests suggest receptor A interacts between -135 and -249 bp (upstream) from the start of ovalbumin transcription. Preferential binding to a DNA fragment containing the sequence -16 to -247 compared to an adjacent upstream fragment or plasmid DNA fragment confirm this assignment. This binding site lies within the regulatory sequence required for progesterone induction of a transfected fusion gene in primary chicken oviduct cells.
D000818:Animals; D002645:Chickens; D004247:DNA / Q000378:metabolism; D004121:Dimethyl Sulfoxide / Q000378:metabolism; D005260:Female; D005809:Genes, Regulator; D005990:Glycerol / Q000378:metabolism; D007700:Kinetics; D009994:Osmolar Concentration; D010047:Ovalbumin / Q000235:genetics*; D010057:Oviducts / Q000032:analysis; D011980:Receptors, Progesterone / Q000378:metabolism*; D013696:Temperature
D011980:Receptors, Progesterone; D010047:Ovalbumin; D004247:DNA; D005990:Glycerol; D004121:Dimethyl Sulfoxide