Evidence for a Specific Estradiol Binding Site on Rat Pituitary Membranes

Abstract

Using adenohypophyses from normal female rats, we demonstrate that estradiol binds pituitary membranes to one homogeneous population of sites with high affinity [dissociation constant (Kd) = 0.041 +/- 0.014 nM; n = 6] and low capacity [maximum binding (Bmax) = 13.6 +/- 5.6 fmol/mg protein]. The binding is thermolabile. Association experiments show that the best experimental conditions are an overnight incubation at 0 C. When the amount of proteins is increased more than 0.3 mg/ml of membrane suspension, binding is rapidly nonlinear. The presence of 0.5 M leupeptin does not improve the binding. Extensive washing of the membranes does not decrease the amount of sites, indicating that the binding is not loosely attached to the membranes. Parenthetically, it should be noted that the membrane fraction was devoid of the cytosolic enzyme marker, lactate dehydrogenase. Binding is specific for estrogenic compounds. When 100% specific binding was determined in the presence of 10(-6) M diethylstilbestrol, 17 beta-estradiol, estrone, and estriol displaced total binding by 110, 80, and 75%, respectively. Neither 4-OH-tamoxifen nor dihydrotestosterone, progesterone, or cortisol displaced the binding. Taken together, these data argue in favor of the presence of specific membrane recognition sites for estradiol in the rat pituitary.

Induction of phosphoinositide-mediated signal transduction pathway by 17 beta-oestradiol in rat vaginal epithelial cells

“…A second messenger pathway is possible if the hormone receptors are associated with membranes or alternatively if the hormone is able to affect directly the membrane-associated signal-transducing enzymes during its passage through the membrane. The presence of specific binding sites for steroid hormones on the membrane of endometrial cells (Pietras & Szego 1977), rat pituitary (Bression et al 1986, Pappas et al 1995 has been demonstrated. Furthermore, Grove & Korach (1987) and Freter et al (1988) have shown enhanced phosphatidyl inositol (PI) lipid metabolism in mouse uteri and breast cancer cells.…”

The integrated action of oestrogen receptor isoforms and sites with progesterone receptor in the gonadotrope modulates LH secretion: evidence from tamoxifen-treated ovariectomized rats

“…Rat gonadotropes express both isoforms of ER, the predominant ERa (Lindzey et al 2006) and ERb at nuclear level (Kuiper et al 1997, Mitchner et al 1998, Vaillant et al 2002, Sánchez-Criado et al 2005a. In addition, an ER which appears to be the same protein as nuclear ERa is expressed at the plasma membrane level (Bression et al 1986, Razandi et al 1999, Toran-Allerand et al 1999, Schmidt et al 2000, Kelly & Levin 2001, Simoncini & Genazzani 2003, Toran-Allerand 2004, Levin 2005. Integration of E 2 effects at different ER pools in the gonadotrope (nuclear ERa and ERb and membrane ERa) determines the cellular actions of E 2 on basal and preovulatory LH secretion and hence on ovulation (Fink 1988(Fink , 2000.…”

Cell surface expression of the 70-kD component of Ku, a DNA-binding nuclear autoantigen.

“…Examples include the originspecific DNA-binding protein of Bacillus subtilis (13) and the SV-40 large T antigen (14). The only known example of such proteins in eukaryotic cells may be the steroid receptors, which bind DNA to exert their action, and are thought to be associated with the plasma membrane (15)(16)(17). The function ofthe 70-kD protein is not known.…”