Effect of estrogen on gene expression in the chick oviduct. V. Changes in the number of RNA polymerase binding and initiation sites in chromatin

Abstract

Estrogen administration to chicks results in an increase in the chromatin template activity of oviduct target tissue as assayed under standard in vitro assay conditions. However, the results obtained by the simple measurement of template activity may be a complicated function of the number of available RNA polymerase initiation sites, the rate of RNA chain elongation, and the rate of reinitiation. In the present study, we have measured separately the change in both the number of chain initiations as well as the rate of RNA chain propagation under conditions in which reinitiation was eliminated. Chromatin prepared from either estrogen-treated or control oviducts both supported an RNA chain elongation rate of six nucleotides per s and a chain size of approximately 700 nucleotides. Thus, both the elongation rate and size of the average product remained relatively constant following estrogen stimulation. In contrast, within 8 hours after a single injection of estrogen to unstimulated chicks, the concentration of RNA polymerase needed to saturate chromatin binding sites was increased to 150% in comparison to control values, and by 24 hours the level of polymerase bound to chromatin was twice that of the untreated control chick chromatin. With daily injections of estrogen, polymerase binding continued to rise. Coincident with the over-all increase in chromatin-bound polymerase was an increase in rifampicin-insensitive initiation sites and newly synthesized RNA chains. Unstimulated chick oviduct chromatin initiated 10,000 RNA chains/pg of DNA, while 24 hours of steroid treatment increased the number of initiated chains to 34,000 chains. These data demonstrate that the estrogen-induced increase in chromatin transcriptive activity was due to an increased number of polymerase binding and initiation sites on the chromatin template without a detectable change in the rate of RNA chain elongation.