Uterine RNA

As early as 1948, Jeener observed an increase in uterine RNA 24 hours following administration of estradiol to mice, without a concomitant rise in DNA concentration. This finding was soon confirmed in the rat, and with the availability of radioactively-Iabeled precursors, experiments could be designed with ever-shortening time intervals between the administration of estrogen and the analysis of the effect in vivo. Ultimately it was demonstrated by Means and Hamilton (1966) that nuclear RNA synthesis in the rat uterus is stimulated within 2 minutes after the administration of estradiol (Fig. 3). At this early time, a 40% increase in the incorporation of uridine into nuclear RNA can be demonstrated. 

In recent years, new evidence has focused attention on the role of RNA in the regulatory action of estrogen. The biological activity of RNA extracted from estrogen-stimulated rat uteri has been studied and the results reveal that several estrogen-like phenomena can be induced by the material. In this work mixed uterine RNA has been extracted by methods that do not preferentially separate species of RNA so that it is not possible to associate the reported activities with a particular nucleic acid component (Segal et ah, 1965a). These proce-... 

It should be noted that these studies pertain to the influence on a bacterial RNA polymerase of a rat uterine fraction that binds estradiol. These observations may or may not be applicable to the action of this fraction on uterine RNA polymerase, a material that has not yet been adequately purified for satisfactory quantitative studies. The binding fraction has been tested for its effect on the incorporation of UTP- C into RNA by nuclei isolated from rat uteri and was found to be inactive. Nevertheless, it is tempting to consider that the macromolecule that binds estradiol intracellularly plays a role in the control of RNA synthesis and that the steroid-protein complex within the cell is more than a transport system, and is an active regulator of cellular metabolism. 

Estrogens stimulate cellular proliferation, induce RNA and protein synthesis of uterine endometrium and the fibrous connective tissue framework for ovaries, and increase the size of the cells. This effect leads to the growth and regeneration of the endometrial layer and spinal arterioles, and increase in the number and size of endometrial glands. Under the influence of estrogen, vaginal mucosa becomes thicker, as cervical mucus becomes thinner (85,86). 

Kumar SA, Beach TA, Dickerman HW 1980. Specificity of oligodeoxynucleotide binding of mouse uterine cytosol estradiol receptors. Proc Natl Acad Sci USA 77(6) 3341-3345. Fire A, Xu S, Montgomery MK, Kostas SA, Driver SE, Mello CC 1998. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391(6669) 806-811. 

Hamilton [77] expanded Mueller s model thanks to the development of adequate techniques for separating nuclear from cytoplasmic uterine fractions. The sequence of events in the uteri of ovariectomized rats after the administration of estradiol-17 jS can be summarized as follows The injected hormones quickly pass the cellular and nuclear membranes, and within minutes after injection a small but consistent fraction of the hormone binds to the nucleus, where it is believed to stimulate the biosynthesis of nuclear RNA. It is not clear which type of RNA is stimulated first, but Hamilton believes that it is chromosomal and ribo-somal RNA. In any event, new messenger must be formed at some time in the sequence because the population of polyribosomes increases in the cytoplasm. 

Fig. 4. High-resolution autoradiogram of rat uterus 2 hrs after subcutaneous injection of 0-63 /ig H-estradiol concentration of radioactivity in cell nuclei of uterine glands and lamina propria (I /i sections, stained after development for DNA and RNA with methyl green pyronin, magnification lOOOx, exposure 42-51 days at — 15°C).

If Karlson s model is valid, it could be expected that RNA extracted from ecdysone-activated epidermis cells would reproduce the effects of the hormone on these cells. Evidence is not forthcoming on the action of such RNA on intact cells. The problems of cell penetration and degradation make the probability of technical success of these experiments very low. However, the introduction, in the uterine cavity of oophorectomized rat, of messenger RNA isolated from the uterus of oestrogen-treated castrated rats, had clear oestrogenic effects. Evidence of the activity of RNA extracted from activated epidermis cells in the synthesis of DOPA decarboxylase in acellular systems is still not clear. 

When ovariectomized immature rats w-ere fed a protein-free diet for 13 days, the concentration of uterine DNA was significantly increased, and the concentration of both protein and RNA remained unchanged, while -glucuronidase activity in the uterus was slightly depressed. Protein deprivation abolished the enhancing effect of corticosterone on i-glucu-... 

Fig. 4. Dependency of RNA synthesis on DNA, as measured by incorporation of UTP- C into RNA of uterine nuclei from castrated rats, treated with estrogen. Incubation for 10 minutes at 37°C. From Taubert and Ilan (1967).

Sucrose gradient profiles reveal that the preponderant component o each type of extract is ribosomal and that messenger RNA is small in amount. Since ribosomal RNA makes up the bulk of the mixed RNA, changes in the composition of the total following hormone treatment would be diflficult to detect by the usual methods of analysis. However, the nucleotide sequences of RNA synthesized by uterine... 

Fig. 6. Effect of 200 /tg of actinomycin D on the uterine response to 5 /ig of estradiol in adrenalectomized immature rats. Water imbibition and the in vitro incorporation over 1 hour of glycine- C (1 ftC) into protein and RNA of uteri were measured in 8-10 animals and averaged. The data are expressed as percent of the control values (wet weight 26.9 mg RNA, 85 cpm/OD 260 m/i protein, 276 cpm./mg), which are taken as 100%. The brackets indicate SE, expressed as percent of the control. From Nicolette and Mueller (1966).

Thus, the concept that estrogens may exert their stimulatory action on the uterus by activation of RNA synthesis has been developed by several lines of biochemical investigation. The evidence includes the observation that estrogen stimulation leads quickly to an increase in the rate of incorporation of precursors into RNA with the initial increase noted in the nuclear fraction of RNA, that RNA polymerase activity in uterine cells is enhanced by estrogen stimulation, that Rie nucleotide sequence of RNA synthesized by uterine nuclei is altered by estrogen, and that specific inhibitors of DNA-dependent RNA synthesis prevent the stimulatory action of estrogen. 

The biological activity of RNA produced by uterine cells under estrogen stimulation suggests that estradiol, either directly or indirectly, stimulates a new pattern of RNA biosynthesis in target cells, and that the hormone is not necessarily involved in subsequent steps leading to the transition from the nonstimulated to the stimulated state. 

Noteboom, W. D., and Gorski, J. (1963). Early effects of estrogen and puro-mycin on uterine synthesis of protein, RNA and RNA polymerase. Federation Proc. 22, 329. 

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