H. Rochefort and M. Garcia
U148 de VINSERM, Montpellier, France
[1]This work has been supported by the Institut National de la Santé et de la Recherche Médicale and the Delegation Generale à la Recherche Scientifique et Technique (France).
We have been concerned to elucidate by the mechanism of action of high pharmacological doses of androgens which induce a marked uterotrophy in the rat uterus (Lerner et al., 1966) and have tried to define the nature of the receptor protein responsible for this effect. Since the affinity of androgens for their specific androgen receptor (RA) is high (KD ~ 0.3 nM) and since high doses of androgens were needed to increase uterine wet weight, we suspect that interaction of androgens with other receptor sites is responsible for this effect. It is now known that androgens such as 5α-dihydrotestosterone (DHT), testosterone, dehydroepiandrosterone and Δ5-androstene diol bind not only to RA but also to the progesterone receptor (Rp) and the oestrogen receptor (RE) (Rochefort and Garcia, 1976; Poortman et al., 1977), and are able to induce in vitro the nuclear translocation of RE (Rochefort et al., 1972). This interaction of androgens with RE and RP is specific but of weak affinity and requires in some cases sensitized competitive experiments to demonstrate this.
This weak affinity binding also explain that the receptor sites which have been occupied by the low affinity ligands can be exchanged very easily at 0°C. An interaction of androgens on the progesterone receptor could not be involved to explain uterotrophy in this system, since progesterone, in contrast to oestradiol does not induce such an hypertrophy. After injection in vivo to immature rats of DHT, an androgen which could not be transformed into oestrogens, we have measured two kinds of uterine metabolic responses. The 3H-leucine incorporation into soluble proteins was a good marker for general protein synthesis and was found to vary parallel with uterine wet weight. The oestradiol induced protein evaluated by the double labelling technique (Katzennellenbogen and Gorski, 1972) represented a more specific response to oestrogens. The saturation and nuclear translocation of the androgen and oestrogen receptors were followed in parallel.
The results obtained indicated that physiological doses of DHT (≤ 100 pg rat) saturated and translocated RA to the nucleus. However, no response was observed neither on 3H-leucine incorporation nor on the oestradiol induced protein. At these doses, the RE was found mainly in the cytosol and no nuclear translocation could be observed. Conversely, when the RE was progressively saturated with doses ≥ 1 mg rat, a complete translocation of RE was observed which was followed by the stimulation of 3H-leucine incorporation. In addition, there was a good correlation between the amount of RE translocated by DHT at three hours and the extent of stimulation of protein synthesis. These results suggested that the occupation and translocation of RE induced by DHT could be responsible for the observed metabolic effect. However, the action of some metabolites of DHT such as androstanediol could not be excluded even though the concentration of unchanged DHT was high (~ 70%). Before concluding that the effect of an androgen was mediated by the oestrogen receptor, it was most important to show that DHT provoked the same specific response as oestradiol.
At the present time, there are two series of data which strongly support the thesis that androgens can act via the oestrogen receptor. The first evidence is that we found marked stimulation of the “oestradiol induced protein” with 10 mg DHT (Garcia and Rochefort, 1977). This approach also indicated that other proteins were stimulated by DHT which were not by oestradiol, thus suggesting that androgens at these large doses could possibly be effective on other undefined receptor sites. The second evidence was provided by Zava and McGuire (1977) who demonstrated that DHT induces the progesterone receptor in the MCF7 human breast cancer cell line.
The consequence of these results concerning the understanding of the molecular mechanism of action of steroid hormone is that the specificity of the response to a steroid hormone appears to be determined by the nature of the receptor translocated to the nucleus rather than to the nature of the hormone itself. When a RE-androgen complex is formed, the present results strongly suggest that the response is oestrogenic. Another remark can be made concerning the difference of concentration between the RE (> 50 000 sites diploid cell) and the RA (~ 1.000 sites cell). Assuming an homogeneous distribution of these receptors in all uterine cells, it is hypothesized but not proven that a low concentration of receptors would fail to induce a global hypertrophy and stimulation of protein synthesis. Finally, this effect of androgens obtained under pharmacological conditions might have practical consequences in human breast cancer in order to explain the efficiency of androgen in the treatment of this tumour (Goldenberget al., 1973) and the possible role of androgen in modulating the frequence of breast cancer (Bulbrook et al., 1962). Whether this interaction of androgens on RE is also responsible for the anti-oestrogenic effect of androgens cannot yet be ascertained.
References
Bulbrook, R. D., Hayward, J. L., Spicer, C. C. and Thomas, B. S. (1962). Lancet 2, 1238-1240.
Garcia, M. and Rochefort, H. (1977). Steroids 29, 111-126.
Goldenberg, I. S., Waters, M. N., Ravdin, R. S., Ansfield, F. J. and Segaloff, A. (1973). J. Am. med. Assoc. 223, 1267.
Katzenellenbogen, B. S. and Gorski, J. (1972). J. Moi. Chem. 247, 1299-1305. Lerner, L. J., Hilf, R., Turkheimer, A. R., Michel, I. and Engel, S. L. (1966).
Endocrinology 78, 111-124.
Poortman, J., Vroegindewey-Jie, D., Thijssen, J. H. H. and Schwartz, F. (1977).
Molec. cell. Endocr. 8, 27-34.
Rochefort, H. and Garcia, M. (1976). Steroids 28, 549-560.
Rochefort, H., Lignon, F. and Capony, F. (1972). Biochem. Biophys. Res. Commun. 47, 662-670.
Zava, D. T. and McGuire, W. L. (1977). The Endocrine Society Meeting, Chicago, Abstract 247. The Endocrine Society Bethesda, Md, p. 180.