ERαΔ4, an ERα splice variant missing exon4, interacts with caveolin‐3 and mGluR2/3

Abstract

The two isoforms of the nuclear estrogen receptor, ERα and ERβ are widely expressed in the central nervous system. Although they were first described as nuclear receptors, both isoforms have also been found at the cell membrane where they mediate cell signaling. Surface biotinylation studies using neuronal and glial primary cultures label an alternatively spliced form of ERα. The 52 kDa protein, ERαΔ4, is missing exon 4 and is highly expressed in membrane fractions derived from cultured cells. In vivo, both full‐length (66 kDa) ERα and ERαΔ4 are present in membrane fractions. In response to estradiol, full‐length ERα and ERαΔ4 are initially trafficked to the membrane, and then internalized in parallel. Previous studies determined that only the full‐length ERα associates with metabotropic glutamate receptor‐1a (mGluR1a), initiating cellular signaling. The role of ERαΔ4, remained to be elucidated. Here, we report ERαΔ4 trafficking, association with mGluR2/3, and downstream signaling in female rat arcuate nucleus (ARH). Caveolin (CAV) proteins are needed for ER transport to the cell membrane, and using co‐immunoprecipitation CAV‐3 was shown to associate with ERαΔ4. CAV‐3 was necessary for ERαΔ4 trafficking to the membrane: in the ARH, microinjection of CAV‐3 siRNA reduced CAV‐3 and ERαΔ4a in membrane fractions by 50%, and 60%, respectively. Moreover, co‐immunoprecipitation revealed that ERαΔ4 associated with inhibitory mGluRs, mGluR2/3. Estrogen benzoate (EB) treatment (5 μg; s.c.; every 4 days; three cycles) reduced levels of cAMP, an effect attenuated by antagonizing mGluR2/3. Following EB treatment, membrane levels of ERαΔ4 and mGluR2/3 were reduced implying ligand‐induced internalization. These results implicate ERαΔ4 in an estradiol‐induced inhibitory cell signaling in the ARH.

Puberty enables oestradiol‐induced progesterone synthesis in female mouse hypothalamic astrocytes

“…17 Indeed, it has been shown that inhibitory E2 signalling in the hypothalamus was mediated by mGluR2/3 that was transactivated by a 52 kDa splice variant of ERα missing exon 4, the ERαΔ4. 22 In the present study, oestradiol was able to elicit an attenuated [Ca 2+ ] i in prepubertal astrocytes, perhaps due to the low-levels of full-length mERα. In these astrocytes, E2 could be signalling through an entirely different membrane-associated receptor, GPR30, whose signalling can mediate rapid E2 effects.…”

“…Conversely, inhibitory E2 signalling is mediated by mGluR2/3 activation of a Gi 17 . Indeed, it has been shown that inhibitory E2 signalling in the hypothalamus was mediated by mGluR2/3 that was transactivated by a 52 kDa splice variant of ERα missing exon 4, the ERαΔ4 22 . In the present study, oestradiol was able to elicit an attenuated [Ca 2+ ] i in prepubertal astrocytes, perhaps due to the low‐levels of full‐length mERα.…”

Estradiol-induced progesterone synthesis develops post-puberty in the rostral hypothalamus and coincides with post-pubertal changes in the steroidogenic pathway in female mouse hypothalamic astrocytes

“…Conversely, inhibitory E2 signaling is mediated by mGluR2/3 activation of a Gi (18). Indeed, it has been shown that inhibitory E2 signaling in the hypothalamus was mediated by mGluR2/3 that was transactivated by a 52 kDa splice variant of ERα missing exon 4, the ERαΔ4 (19). In the present study, estradiol was able to elicit an attenuated [Ca 2+ ] i in prepubertal astrocytes, perhaps due to the low-levels of full-length mERα.…”

“…Indeed, astrocytes derived from the adult female rodent hypothalamus respond to E2 stimulation with an increase in neuroP synthesis, and hypothalamic astrocytes harvested from neonatal female rodents or male rodents did not appear to respond to E2 with increased neuroP synthesis (12). Recently, we (Mohr et al 2019), formally tested whether systemic administration of E2 to pre-and post-pubertal ovariectomized rats induced neuroP in the hypothalamus. Only post-pubertal female rats had elevated hypothalamic neuroP levels.…”

The key action of estradiol and progesterone enables GnRH delivery during gestation in the South American plains vizcacha, Lagostomus maximus

“…In addition, the frequency of GnRH release induced by E2, but not when ER or ERwere separately activated, suggests the possible involvement of another class of ERs in this mechanism. In this way, the activation of membrane ERs, which are usually responsible for rapid and short term non-genomic modulation, such as GPR30 or nuclear ER that translocates to the cell membrane after cleavage and associates with metabotropic glutamate receptors (mGluRs), should be considered in future experiments in order to fully understand the mechanisms of GnRH delivery induction (Takeo & Sakuma 1995, Kim et al 2011a, Kim et al 2011b, Levin 2011, Prossnitz & Barton 2011, Zárate et al 2012, Wong et al 2019.…”