Action of calpain on the basic estrogen receptor molecule of porcine uterus.
Basic estrogen receptor (ER) molecule (vero-ER) of the cytosol of porcine uterus was purified 1,200-fold after successive chromatographies on phenyl-Sepharose, hydroxylapatite, and DEAE-cellulose, followed by Sephadex G-150 gel filtration. The purified vero-ER was completely free from endogenous protease and ER-binding factor. The action of Ca2+-dependent cysteine proteinase (calpain) on vero-ER was studied by utilizing the purified receptor and calpains from porcine uterus (endogenous calpain), porcine kidney, and human erythrocytes. Proteolysis of vero-ER was followed by monitoring the disappearance of the binding capability of vero-ER with "8S" ER-forming factor. Vero-ER was proteolyzed by both the endogenous and the exogenous calpains in the presence of Ca2+. The calpains did not attack vero-ER in the absence of Ca2+. The results indicated the absolute requirement by calpain for Ca2+ for the limited hydrolysis of vero-ER. Uterine cytosol was shown to contain, in parallel with calpain, a protease which does not require Ca2+ for the limited proteolysis of vero-ER. The strongly hydrophobic domain of vero-ER, recently shown to be indispensable for the nuclear translocation of vero-ER (Murayama, A. & Fukai, F. (1983) FEBS Lett. 158, 255), was preferentially destroyed by both the Ca2+-requiring and -nonrequiring enzymes. It was assumed that calpain might intervene in the estrogen action by diminishing irreversibly the amount of the cytoplasmic ER capable of translocating into the nucleus.
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002154:Calpain; D003600:Cytosol / Q000378:metabolism; D010450:Endopeptidases / Q000494:pharmacology*; D005260:Female; D007700:Kinetics; D011960:Receptors, Estrogen / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D013552:Swine; D014599:Uterus / Q000187:drug effects / Q000378:metabolism*
D011960:Receptors, Estrogen; D010450:Endopeptidases; D002154:Calpain; D002118:Calcium