Ecdysone receptor (EcR) was expressed in vertebrate cells to study its functional properties in the absence of the heterodimerization partner Ultraspiracle (Usp) and to avoid interference with endogenous receptor isoforms. Comparison of different isoforms affords determination of receptor concentration, which was achieved either by determination of ligand binding sites by Scatchard analysis or by quantitative evaluation of specific Western blot signals, but not by normalization on transfection efficiency as determined by cotransfection with a constitutive reporter plasmid. Ligand- and DNA- binding, and transcriptional activity of EcR isoforms and the influence of Ultraspiracle (Usp) were described.
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References
Arbeitman MN, Hogness DS (2000) Molecular chaperones activate the Drosophila ecdysone receptor, an RXR heterodimer. Cell 101:67–77
Beatty J, Fauth Th, Callendar JL, Spindler-Barth M, Henrich VC (2006) Functional analysis of ecdysteroid receptor isoforms in Drosophila melanogaster in a cell culture system. Insect Mol Biol 15:785–795
Beckers C, Maroy P, Dennis R, O'Connor JD, Emmerich H (1980) The uptake and release of pon-asterone A by the Kc cell line of Drosophila melanogaster. Mol Cell Endocrinol 17:51–59
Cherbas P, Cherbas L, Lee SS, Nakanishi K (1988) 26-[125I]iodoponasterone A is a potent ecdysone and a sensitive radioligand for ecdysone receptors. Proc Natl Acad Sci USA 85:2096–2100
Cronauer MV, Braun S, Tremmel Ch, Krönke K-D, Spindler-Barth M (2007) Nuclear localization and DNA binding of ecdysone receptor and ultraspiracle. Arch Insect Biochem Physiol 65:125–133
Dela Cruz FE, Kirsch DR, Heinrich JN (2000) Transcriptional activity of Drosophila mela-nogaster ecdysone receptor isoforms and ultraspiracle in Saccharomyces cerevisiae. J Mol Endocrinol 24:183–191
Ghbeish N, Tsai CC, Schubiger M, Zhou JY, Evans RM, McKeown M (2001) The dual role of ultraspiracle, the Drosophila retinoid X receptor, in the ecdysone response. Proc Natl Acad Sci USA 98:3867–3872
Grebe M, Przibilla S, Henrich VC, Spindler-Barth M (2003) Characterization of the ligand-binding domain of the ecdysteroid receptor from Drosophila melanogaster. Biol Chem 384:105–116
Grebe M, Fauth T, Spindler-Barth M (2004) Dynamic of ligand binding to Drosophila mela-nogaster ecdysteroid receptor. Insect Biochem Mol Biol 34:981–989
Greb-Markiewicz B, Fauth T, Spindler-Barth M (2005) Ligand binding is without effect on complex formation of the ligand binding domain of the ecdysone receptor (EcR). Arch Insect Biochem Physiol 59:1–11
Gwozdz T, Dutko-Gwozdz J, Nieva C, Betanska K, Orlowski M, Kowalska A, Dobrucki J, Spindler-Barth M, Spindler K-D, Ozyhar A (2007) EcR and Usp, components of the ecdys-teroid nuclear receptor complex, exhibit differential distribution of molecular determinants directing subcellular trafficking. Cell Signal 19:490–503
Haelens A, Tanner T, Denayer S, Callewaert L, Claessens F (2007) The hinge region regulates DNA binding, nuclear translocation, and transactivation of the androgen receptor. Cancer Res 67:4514–4523
Halling BP, Yuhas DA, Eldridge RR, Gilbey SN, Deutsch VA, Herron JD (1999) Expression and purification of the hormone binding domain of the Drosophila ecdysone and ultraspiracle receptors. Protein Expr Purif 17:373–386
Henrich VC (2005) The ecdysteroid receptor. In: Comprehensive Molecular Insect Science (eds. L.J. Gilbert, K. Iatrou, and S.S. Gill), Oxford, Elsevier/Pergamon. Vo l 3, 243–282
Henrich VC, Burns E, Yelverton DP, Christensen E, Weinberger C (2003) Juvenile hormone potentiates ecdysone receptor-dependent transcription in a mammalian cell culture system. Insect Biochem Mol Biol 33:1239–1244
Hu X, Cherbas L, Cherbas P (2003) Transcription activation by the ecdysone receptor (EcR/USP): identification of activation functions. Mol Endocrinol 17:716–731
Lezzi M, Bergman T, Henrich VC, Vogtli M, Fromel C, Grebe M, Przibilla S, Spindler-Barth M (2002) Ligand-induced heterodimerization between the ligand binding domains of the ecdys-one receptor. Eur J Biochem 269:3237–3245
Maroy P, Dennis R, Beckers C, Sage BA, O'Connor JD (1978) Demonstration of an ecdysteroid receptor in a cultured cell line of Drosophila melanogaster. Proc Natl Acad Sci USA 75:6035–6038
Mouillet JF, Henrich VC, Lezzi M, Vögtli M (2001) Differential control of gene activity by iso-forms A, B1 and B2 of the Drosophila ecdysone receptor. Eur J Biochem 268:1811–1819
Nieva C, Gwozdz T, Dutko-Gwozdz J, Wiedenmann J, Spindler-Barth M, Wieczorek E, Dobrucki J, Dus D, Henrich V, Ozyhar A, Spindler K-D (2005) Ultraspiracle promotes the nuclear localization of ecdysteroid receptor in mammalian cells. Biol Chem 386:463–470
Nieva C, Spindler-Barth M, Spindler K-D (2007) Influence of hormone on intracellular localization of the Drosophila melanogaster ecdysteroid receptor (EcR). Cell Signal 19:2582–2587
Nieva C, Spindler-Barth M, Azoitei A, Spindler K-D (2008) Impact of heterodimerization on intracellular localization of the ecdysteroid receptor (EcR). Arch Insect Biochem Physiol 68:40–49
Ogura T, Minakuchi C, Nakagawa Y, Smagghe G, Miyagawa H (2005) Molecular cloning, expression analysis and functional confirmation of ecdysone receptor and ultraspiracle from the Colorado potato beetle Leptinotarsa decemlineata. FEBS J 272:4114–4128
Spindler-Barth M, Spindler K-D (2003) Ecdysteroid receptors (EcR/USP). In: Encyclopedia of Hormones and Related Cell Regulators (eds. H.L. Henry and A.W. Norman), Academic Press, New-York, London, pp 466–470.
Yao TP, Forman BM, Jiang Z, Cherbas L, Chen JD, McKeown M, Cherbas P, Evans RM (1993) Functional ecdysone receptor is the product of EcR and Ultraspiracle genes. Nature 366:476–479
Yund MA, King DS, Fristrom JW (1978) Ecdysteroid receptors in imaginal discs of Drosophila melanogaster. Proc Natl Acad Sci USA 75:6039–6043
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Azoitei, A., Ruff, H., Tremmel, C., Braun, S., Spindler-Barth, M. (2009). Functional Analysis of Ecdysteroid Receptor from Drosophila melanogaster “In Vitro”. In: Smagghe, G. (eds) Ecdysone: Structures and Functions. Springer, Dordrecht. https://doi.org/10.1007/978-1-4020-9112-4_15
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