Razandi M, Pedram A, Greene G L, Levin E R
Department of Medicine, University of California, Irvine 92717, USA.
Mol Endocrinol. 1999 Feb;13(2):307-19. doi: 10.1210/mend.13.2.0239.
The existence of a putative membrane estrogen receptor (ER) has been supported by studies accomplished over the past 20 yr. However, the origin and functions of this receptor are not well defined. To study the membrane receptor, we transiently transfected cDNAs for ERalpha or ERbeta into Chinese hamster ovary (CHO) cells. Transfection of ERalpha resulted in a single transcript by Northern blot, specific binding of labeled 17beta-estradiol (E2), and expression of ER in both nuclear and membrane cell fractions. Competitive binding studies in both compartments revealed near identical dissociation constants (K(d)S) of 0.283 and 0.287 nM, respectively, but the membrane receptor number was only 3% as great as the nuclear receptor density. Transfection of ERbeta3 also yielded a single transcript and nuclear and membrane receptors with respective Kd values of 1.23 and 1.14 nM; the membrane receptor number was only 2% compared with expressed nuclear receptors. Estradiol binding to CHO-ERalpha or CHO-ERbeta activated Galphaq and G(alpha)s proteins in the membrane and rapidly stimulated corresponding inositol phosphate production and adenylate cyclase activity. Binding by 17-beta-E2 to either expressed receptor comparably enhanced the nuclear incorporation of thymidine, critically dependent upon the activation of the mitogen-activated protein kinase, ERK (extracellular regulated kinase). In contrast, c-Jun N-terminal kinase activity was stimulated by 17-beta-E2 in ERbeta-expressing CHO, but was inhibited in CHO-ERalpha cells. In summary, membrane and nuclear ER can be derived from a single transcript and have near-identical affinities for 17-beta-E2, but there are considerably more nuclear than membrane receptors. This is also the first report that cells can express a membrane ERbeta. Both membrane ERs activate G proteins, ERK, and cell proliferation, but there is novel differential regulation of c-Jun kinase activity by ERbeta and ERalpha.
过去20年完成的研究支持了假定的膜雌激素受体（ER）的存在。然而，该受体的起源和功能尚未明确界定。为了研究膜受体，我们将ERα或ERβ的cDNA瞬时转染到中国仓鼠卵巢（CHO）细胞中。通过Northern印迹法，ERα的转染产生了单一转录本，标记的17β-雌二醇（E2）发生特异性结合，并且ER在细胞核和细胞膜组分中均有表达。两个区室中的竞争性结合研究分别显示解离常数（K(d)S）近乎相同，分别为0.283和0.287 nM，但膜受体数量仅为核受体密度的3%。ERβ3的转染也产生了单一转录本以及核受体和膜受体，其Kd值分别为1.23和1.14 nM；与表达的核受体相比，膜受体数量仅为2%。雌二醇与CHO-ERα或CHO-ERβ的结合激活了膜中的Gαq和G(α)s蛋白，并迅速刺激了相应的肌醇磷酸生成和腺苷酸环化酶活性。17-β-E2与任一表达受体的结合同等程度地增强了胸苷的核掺入，这严重依赖于丝裂原活化蛋白激酶ERK（细胞外调节激酶）的激活。相比之下，17-β-E2在表达ERβ的CHO中刺激了c-Jun N末端激酶活性，但在CHO-ERα细胞中受到抑制。总之，膜ER和核ER可源自单一转录本，并且对17-β-E2具有近乎相同的亲和力，但核受体比膜受体多得多。这也是关于细胞可表达膜ERβ的首次报道。两种膜ER均激活G蛋白、ERK和细胞增殖，但ERβ和ERα对c-Jun激酶活性存在新的差异调节。
