An Estrogen Receptor-α Knock-In Mutation Provides Evidence of Ligand-Independent Signaling and Allows Modulation of Ligand-Induced Pathways in Vivo

Abstract

Estrogen-nonresponsive estrogen receptor-alpha (ERalpha) knock-in (ENERKI) mice were generated to distinguish between ligand-induced and ligand-independent ER-alpha actions in vivo. These mice have a mutation [glycine 525 to leucine (G525L)] in the ligand-binding domain of ERalpha, which significantly reduces ERalpha interaction with and response to endogenous estrogens, whereas not affecting growth factor activation of ligand-independent pathways. ENERKI mice had hypoplastic uterine tissues and rudimentary mammary gland ductal trees. Females were infertile due to anovulation, and their ovaries contained hemorrhagic cystic follicles because of chronically elevated levels of LH. The ENERKI phenotype confirmed that ligand-induced activation of ERalpha is crucial in the female reproductive tract and mammary gland development. Growth factor treatments induced uterine epithelial proliferation in ovariectomized ENERKI females, directly demonstrating that ERalpha ligand-independent pathways were active. In addition, the synthetic ERalpha selective agonist propyl pyrazole triol (PPT) and ER agonist diethylstilbestrol (DES) were still able to activate ligand-induced G525L ERalpha pathways in vitro. PPT treatments initiated at puberty stimulated ENERKI uterine development, whereas neonatal treatments were needed to restore mammary gland ductal elongation, indicating that neonatal ligand-induced ERalpha activation may prime mammary ducts to become more responsive to estrogens in adult tissues. This is a useful model for in vivo evaluation of ligand-induced ERalpha pathways and temporal patterns of response. DES did not stimulate an ENERKI uterotrophic response. Because ERbeta may modulate ERalpha activation and have an antiproliferative function in the uterus, we hypothesize that ENERKI animals were particularly sensitive to DES-induced inhibition of ERalpha due to up-regulated uterine ERbeta levels.

Developmental Phenotype of a Membrane Only Estrogen Receptor α (MOER) Mouse

“…Our model begins to sort this out in that membrane-initiated signaling by E2 is evident but is not sufficient by itself for the normal organ development investigated here. A requirement of E2 binding to ER␣ for normal reproductive tract and mammary gland development and function in female mice has also recently been established (21).…”

Estrogen receptor α AF-2 mutation results in antagonist reversal and reveals tissue selective function of estrogen receptor modulators

“…This result is different from those in a similar mouse model in which the ERα AF-2 is mutated: the ENERKI mouse. The ERα in the ENERKI mouse is mutated in the ligand-binding pocket (G525L), is unable to bind E2, and exhibits reduced E2-dependent transcription activation while maintaining helix 12 structure (29). In the ENERKI homozygous female, IGF-1 induced the proliferation of uterine endometrial epithelial cells without E2, as reflected by an increase in the general cell cycle marker Ki67.…”

Oestrogens and spermatogenesis

“…4. Because ERa is present in ArKO ductules, ERa is activated in a ligand-independent pathway (Power et al 1991;Salomonsson et al 1994;Weis et al 1996;Wang et al 2001;O'Malley 2005;McDevitt et al 2007;Sinkevicius et al 2008Sinkevicius et al , 2009). 5.…”