Early Estrogen Action: Stimulation of the Metabolism of High Molecular Weight and Ribosomal RNAs

Abstract

Samples of RNA, isolated from uteri of ovariectomized adult rats treated with estrogen, have been analyzed on sucrose gradients. Treatment with estrogen either for 20 min or 2 hr increased the specific activity of all classes of uterine RNA, but produced no significant alteration in the distribution of radioactivity in the gradients, when animals received [3H] One of the earliest effects of estrogen on the metabolism of the uterus of the immature or ovariectomized adult rat is an acceleration of the rate of RNA synthesis (for review, see ref. 1). Furthermore, the ability of estrogen to elicit pronounced physiological responses in the uterus appears to be mediated, at least in part, by this modification of RNA metabolism (2). Attempts have been made to isolate rat uterine RNA and to compare the rapidly labeled RNA formed in the organ stimulated by estrogen with that formed in the absence of the hormone, but the results obtained by different investigators (3-5) have been difficult to interpret. One reason for this is that the methods used in the past to extract total, and particularly nuclear, RNA from the uterus have generally resulted in the isolation of partially degraded and/or contaminated RNA samples, as indicated either by an examination of the procedures used or by the sucrose gradient analyses reported. In investigations designed to measure the effect of estrogen on the metabolism of RNA in the uterus of the ovariectomized adult rat, we have developed a procedure for the isolation of RNA from this organ that routinely yields an undegraded and uncontaminated product. The procedure is similar to that described by Joel and Hagerman (5). We now report the results of sucrose gradient analyses of isolated uterine RNA labeled in vivo. We conclude that estrogen very early in its action in the uterus stimulates the synthesis of all major classes of RNA. In addition, RNA samples isolated from hormone-treated animals showed a smaller percentage of labeled, rapidly sedimenting RNA species (faster than 28 S) 157 than RNA from rats not given estrogen. This observation indicates that the rate or efficiency of processing of precursor RNA species of high molecular weight is also increased during the early action of the hormone. MATERIALS AND METHODSAdult Sprague-Dawley rats of uniform weight (160-180 g) and age, ovariectomized 3-4 weeks before experimentation, were used.RNA was routinely extracted from the combined uteri of four animals by a modification of the sodium dodecyl sulfatehot phenol procedure (6, 7). The rats were killed by cervical dislocation and their uteri were rapidly removed and dropped directly into ethanol-dry ice. The frozen tissue was then broken into small pieces, blotted, and homogenized in a mixture of 4 ml of aqueous phase (0.05 M sodium acetate, 0.05 M NaCl, 0.5% bentonite, and 0.001% polyvinyl sulfate, pH 5.1) and 4 ml of water-saturated phenol containing 0.1% 8-hydroxyquinoline at 20C. Homogenization was with a Polytron (PT 10) tissue disintegrator (Kinematica Gmbh., Lucerne, Switzerlan...

Clomiphene and Tamoxifen Action in the Rat Uterus

“…There have been several reports of an increase in high-molecular-weight RNA early in the uterine response (Luck & Hamilton, 1972;Knowler & Smellie, 1973) and these results are supported by the observations at the same time of an enhancement of the activity of RNA polymerase B, the enzyme believed to be involved in the synthesis of heavy nuclear RNA (Glasser, Chytil & Spelsberg, 1972;Borthwick & Smellie, 1975). Later effects of oestradiol on ribosomal RNA synthesis occur 2-4 h after hormone treatment (Hamilton, Widnell & Tata, 1968;Knowler & Smellie, 1971), times at which the activity of RNA polymerase A is also enhanced, this being the enzyme involved in rRNA synthesis (Glasser et al 1972;Borthwick & Smellie, 1975).…”

The In Vitro Synthesis of RNA within the Rat Nodose Ganglion Following Vagotomy

“…This suggests that the synthesis of cytoplasmic rRNA from its precursors is not normally 100% efficient and that following trauma or stimulation cells may be able to call upon this reserve of unutilised precursor. The existence of this rapid method of control over the synthesis of ribosomal RNA has been reported in a number of other cell types (Luck and Hamilton, 1972;Cooper, 1972;Rizzo and Webb, 1972;Vaughan, 1972).…”

EARLY INCREASES IN RIBONUCLEIC ACID POLYMERASE ACTIVITIES OF DIMETHYLBENZANTHRACENE-INDUCED MAMMARY TUMOUR NUCLEI IN RESPONSE TO OESTRADIOL-17β AND TAMOXIFEN

“…increases RNA synthesis (Hamilton, Teng, Means & Luck, 1971 ;Knowler & Smellie, 1971). This increase appears to be dependent upon an early rise in synthesis of DNA-like RNA without which the stimulation of total RNA synthesis cannot occur (Knowler & Smelile, 1971;Luck & Hamilton, 1972;Borthwick «fe Smellie, 1975).…”