The two native estrogen receptor forms of 8S and 4S present in cytosol from human uterine tissues display opposite reactivities with the antiestrogen tamoxifen aziridine and the estrogen responsive element.
We have investigated the capability of the different native ER forms, present in cytosols from human uterine tissues, of reacting with the antiestrogen [3H]Tamoxifen aziridine ([3H]TA) and with the Estrogen Responsive Element (ERE). Cytosols from uterine leiomyoma (myoma) prepared in buffer containing 40 mM molybdate and protease inhibitors, labelled with [3H]estradiol and analyzed in low-salt sucrose gradient showed 8S and 4S ER forms. The same cytosols labelled with [3H]TA only showed a 4S ER form, whereas the ERE only reacted with fractions from the 8S peak. The band of ERE reaction in the EMSA assay showed a lower relative mobility than the band labelled with [3H]TA, but both bands contained immunoreactive ER of 65 kDa. Electrophoretic mobility of the [3H]TA-labelled band in that system was not affected by cytosol treatment with cross-linkers or SDS, which suggests that it is a monomeric protein. The [3H]TA-binding 4S ER form was found in all studied myoma samples, as well as in human endometrium or myometrium, but not in rat tissues. These results suggest that the 8S and 4S ER form were already present before cytosol from human uterine tissues comes into contact with the molybdate buffer. They both contain the same ER molecule of 65 kDa, either in the free form or as an oligomer. Only the ER dimers, which have been described both in the cytosolic 8S form and in the nuclear 4-5S form, react with the ERE. [3H]TA only binds to the 4S ER monomer probably because its binding site is concealed in the 8S form under these experimental conditions. The opposite reactivity of the 8S and 4S ER forms with [3H]TA and the ERE support the hypothesis that they may constitute separate entities with a different physiological role.
Navarro
D
D
Departamento de Endocrinología Celular y Molecular, Centro de Ciencias de la Salud y Facultad de Veterinaria, Universidad de Las Palmas de Gran Canaria, Canary Islands, Spain.
León
L
L
Chirino
R
R
Fernández
L
L
Pestano
J
J
Díaz-Chico
B N
BN
D000818:Animals; D001665:Binding Sites; D003600:Cytosol / Q000378:metabolism; D004717:Endometrium / Q000378:metabolism; D004965:Estrogen Antagonists / Q000378:metabolism*; D005260:Female; D006801:Humans; D066298:In Vitro Techniques; D007889:Leiomyoma / Q000378:metabolism; D008970:Molecular Weight; D009215:Myometrium / Q000378:metabolism; D009376:Neoplasms, Hormone-Dependent / Q000378:metabolism; D051381:Rats; D011960:Receptors, Estrogen / Q000737:chemistry / Q000378:metabolism*; D013629:Tamoxifen / Q000031:analogs & derivatives* / Q000378:metabolism; D014594:Uterine Neoplasms / Q000378:metabolism; D014599:Uterus / Q000378:metabolism*
D004965:Estrogen Antagonists; D011960:Receptors, Estrogen; D013629:Tamoxifen; C031859:tamoxifen aziridine