Susan H. Socher

Estrogen-mediated cell proliferation during chick oviduct development and its modulation by progesterone

Abstract The interactions of estrogen and progesterone on mitosis were examined in the surface epithelium of the developing chick oviduct. Both of these steroid hormones can stimulate cells to divide in the unstimulated oviduct. However, progesterone treatment results in a delayed suppression of cell division in both the presence and absence of estrogen. This progesterone induced depression of estrogen-mediated cell division is observed throughout oviduct development. During oviduct development estrogen is necessary for both cell division and the differentiation of specific cell types while progesterone appears to modify the action of estrogen by blocking the progression of cells through the cell cycle.

An analysis of the binding of the chick oviduct progesterone-receptor to chromatin

The binding of progesterone-receptor complexes to chromatin from target and nontarget tissues was studied in vitro. Chromatin from both target and nontarget tissues responds in a similar manner to saly and cofactors and has the same K(D) (approx. 3.10(-9) M) for the progesterone-receptor complex. The only observed difference in the binding of the progesterone-receptor complex to target and nontarget chromatins is the difference in total number of acceptor sites. oviduct chromatin has approx. 1300 sites/pg DNA, spleen chromatin has approx. 840 sites/pg DNA, and erythrocyte chromatin has about 330 sites/pg DNA. The K(D) and number of acceptor sites for progesterone-receptor complex binding to oviduct chromatin remains the same even after extensive purification of the progesterone-receptor complex. Activation of cytosol labeled with [3H]progesterone by preincubation at 25 degrees C, analogous to that required for maximal nuclear binding, occurs if the binding studies to chromatin are performed in 0.025 M salt. The absence of an observable temperature effect when the studies are performed at 0.15 M salt is due to the activation of the receptor by salt. The dissociation of the progesterone-receptor complex from chromatin exhibits a single dissociation rate and the initial event is the appearance of free progesterone rather than a progesterone-receptor complex. Lastly, the treatment of chromatin with an antibody prepared against either single-stranded DNA or double-stranded DNA does not alter the extent of binding of the progesterone-receptor complex. Similarly, pretreatment of chromatin with a single-stranded nuclease does not inhibit the capacity of chromatin to bind the hormone-receptor complex.

Detection of casein messenger RNA in hormone-dependent mammary cancer by molecular hybridisation

THE growth and development of the normal mammary gland are regulated by the complex interaction of both peptide and steroid hormones1. Whereas some breast cancers retain the hormone-dependent characteristics of normal mammary tissue2,3, other tumours loose this responsiveness and grow autonomously. Following the pioneering work of Huggins4 on the hormonal dependence of both experimental and human breast cancer, investigators seeking a prognostic test have studied the relationship of specific oestrogen receptors to mammary cancer5,6 to the response of the tumour to endocrine ablation7,8, and to the levels of other steroid9 and peptide hormone receptors10,11. A superior marker for hormone responsiveness would, however, be a measurable product of hormone action rather than the initial binding interaction. In the mammary gland, casein synthesis has been used as a specific biochemical marker of differentiated function and hormone responsiveness. In the studies reported here, a specific complementary DNA copy (cDNA) of rat casein mRNA has been utilised to study the effects of hormones on the expression of differentiated function in chemical carcinogen-induced mammary aden-carcinomas. It is shown that casein mRNA as detected by cDNA may be a useful indicator of hormone dependence in experimental breast cancer. The cDNA probe was synthesised using as a template a 15S casein mRNA fraction purified 180-fold from a lactating RNA extract by chromatography on dT-cellulose and several precise sizing techniques12,13. The template mRNA fraction represented greater than 90% casein mRNA, as estimated by specific immunoprecipitation of the total protein synthesised in the wheat germ assay, and by a careful analysis of its kinetics of hybridisation with the cDNA probe13. The specificity of the complementary DNA copy of the 15S casein mRNA has been demonstrated previously13. This cDNA probe selectively hybridised to RNA from lactating tissue but not to rat liver poly(A)-containinig RNA and the resulting hybrid displayed a high Tm characteristic of a well basepaired duplex. In addition, the rate of hybridisation of the casein-specific cDNA to various RNA preparations was directly related to the casein mRNA activity of these same preparations determined by a cell-free translation assay.

Dysdifferentiative nature of aging: age-dependent expression of mouse mammary tumor virus and casein genes in brain and liver tissues of the C57BL/6J mouse strain

Many aspects of the aging process could be the result of cells slowly drifting away from their proper state of differentiation. This possibility has been studied by searching for an age-dependent increase in the expression of specific genes in tissues where expression of these genes would not normally be expected. In these studies, cDNA probes of specific genes are used in a DNA X NA hybridization assay to detect possible complementary RNA sequences in tissues of different-aged animals. Using this technique in past experiments, a qualitative increase in the RNA sequence complexity of mouse leukemia virus (MuLV) and a quantitative increase in the amount of alpha- and beta-globin RNA were found with increasing age in the brain and liver of the C57BL/6J mouse strain. We report here a similar age-dependent qualitative increase in the RNA sequence complexity for mouse mammary tumor virus (MMTV) but no quantitative or qualitative age-dependent change in casein RNA sequences for the same tissues and mouse strain.