G protein-coupled receptor 30: estrogen receptor or collaborator?

More than 800 G protein-coupled receptors (GPRs) are transcribed in the human genome (1). For many of these receptors, no endogenous ligand has been found, and, therefore, they are currently designated as orphan receptors. One such protein, GPR30, has been proposed to be an estrogen receptor (ER) (2, 3). GPR30 has been localized either to the cell plasma membrane (2) or to the endoplasmic reticulum (3) by two investigative groups, and has been proposed to mediate estrogen action as a steroid receptor. At the cellular level, the early experimental support for this idea is simply not strong. 17-Estradiol (E2) shows minimal (arguably background) binding to GPR30 and quite modest signal transduction (4). This is compared with dramatically stronger actions of physiological E2 at classical endogenous (5) or exogenous ER (6). A recent report indicates that radioactive E2 fails to bind GPR30 in a saturable or specific fashion, and E2 does not stimulate cAMP or calcium release (7). Thus, there is scant support that E2 directly binds GPR30. In addition, a putative GPR30 agonist, G1 (8), does not stimulate estrogen-like effects in the uterus or mammary gland of mice (7). It is puzzling that the receptor pharmacology is quite different, in that ICI182780 (fulvestrant; AstraZeneca Sverige AB, Södertälje Sweden), an ER antagonist, is proposed to strongly activate GPR30 (2). Importantly, it has not been demonstrated that fulvestrant actually binds GPR30. Finally, GPR30 is incapable of mediating any actions of estrogen in cells in which ER and ER are absent (5), and several laboratories have shown that E2 effects are unaltered when GPR30 has been knocked down with interfering RNA (5, 9). To test rigorously the hypothesis that GPR30 is a functional ER, Isensee et al. (10) created a GPR30-lacZ reporter mouse, described in this issue of Endocrinology. The lacZ reporter was inserted into the GPR30 gene locus, causing a partial deletion of the GPR30 coding sequence but identifying where GPR30 is expressed in the mouse. The tissue distribution showed extensive expression in the brain and endocrine organs, consistent with GPR30 production in many tissues. Isensee et al. (10) then performed extensive phenotyping of female mice. No reproductive tract or mammary gland abnormalities were seen, and the mice were fertile. Body weight and fat mass were comparable to wild-type littermates, even upon challenge with a high-fat diet. Under normal or high-fat feeding conditions, glucose tolerance testing was indistinguishable between wild-type and GPR30-lacZ mice. The authors then performed an exhaustive screen of behavior, assessment of the physiology of many organs, and blood chemistries. The only difference found between wild-type and GPR30-lacZ mice was a moderate decrease in CD4 and CD8 T lymphocyte cells in the blood of the GPR30-lacZ mice. Thus, most biological functions were not influenced by the loss of GPR30 function. Importantly, there were no similarities of the GPR30-lacZ mouse to the extraordinary phenotype of female ER knockout (KO) mice; the latter mice demonstrate a wide variety of abnormal organ development and functions (11, 12). The lack of phenotype in the GPR30-lacZ mouse nicely complements several other recent reports investigating different GPR30 KO mice. Otto et al. (13) created a complete GPR30 KO mouse by deleting exon 3. As a result, GPR30 was not expressed in any reproductive tract or mammary tissues, compared with extensive expression of this orphan receptor in these same organs from wild-type littermates. In female GPR30 KO mice, reproductive tract and mammary gland development was indistinguishable from that of wild-type littermates. In addition, fertility in the two mice was comparable, and uterine responses to injected E2 showed similar epithelial cell proliferation and gene expression. The authors concluded that GPR30 does not mediate these classical actions of E2 in female mice. In work from Wang et al. (14), the only estrogen-related phenotype of a GPR30 KO mouse reported was a decrease in thymus size. Moreover, some of the effects of E2 on thymocyte development were reported as mediated through ER . In vivo, E2 induced less cell death of F-CD4/CD8 double-negative thymocytes from GPR30 KO mice compared with wild-type littermates. However, GPR30 expression in wild-type mice was not found in the F-CD4/CD8 double-