title: Characterization of evolutionary trend in squamate estrogen receptor sensitivity.
abstract: Steroid hormones are a key regulator of reproductive biology in vertebrates, and are largely regulated via nuclear receptor families. Estrogen signaling is regulated by two estrogen receptor (ER) subtypes alpha and beta in the nucleus. In order to understand the role of estrogen in vertebrates, these ER from various species have been isolated and were functionally analyzed using luciferase reporter gene assays. Interestingly, species difference in estrogen sensitivity has been noted in the past, and it was reported that snake ER displayed highest estrogen sensitivity. Here, we isolated additional ER from three lizards: chameleon (Bradypodion pumilum), skink (Plestiodon finitimus), and gecko (Gekko japonicus). We have performed functional characterization of these ERs using reporter gene assay system, and found high estrogen sensitivity in all three species. Furthermore, comparison with results from other tetrapod ER revealed a seemingly uniform gradual pattern of ligand sensitivity evolution. In silico 3D homology modeling of the ligand-binding domain revealed structural variation at three sites, helix 2, and juncture between helices 8 and 9, and caudal region of helix 10/11. Docking simulations indicated that predicted ligand-receptor interaction also correlated with the reporter assay results, and overall squamates displayed highest stabilized interactions. The assay system and homology modeling system provides tool for in-depth comparative analysis of estrogen function, and provides insight toward the evolution of ER among vertebrates.
authors.affiliation: Department of Basic Biology, Faculty of Life Science, SOKENDAI (Graduate University for Advanced Studies), 5-1 Higashiyama, Myodaiji, Okazaki, Aichi 444-8787, Japan. Electronic address: ryoheiy@nibb.ac.jp.
authors.forename: Ryohei
authors.initials: R
authors.lastname: Yatsu
authors.affiliation: Graduate School of Life Science and Department of Biological Sciences, Hokkaido University, Sapporo, Hokkaido 060-0810, Japan. Electronic address: ykatsu@sci.hokudai.ac.jp.
authors.forename: Yoshinao
authors.initials: Y
authors.lastname: Katsu
authors.affiliation: Department of Obstetrics and Gynecology, Medical University of South Carolina, and Marine Biomedicine and Environmental Science Center, Hollings Marine Laboratory, Charleston, SC 29412, USA. Electronic address: kohno@musc.edu.
authors.forename: Satomi
authors.initials: S
authors.lastname: Kohno
authors.affiliation: Okazaki Institute for Integrative Bioscience, National Institute for Basic Biology, National Institutes of Natural Sciences, 5-1 Higashiyama, Myodaiji, Okazaki, Aichi 444-8787, Japan. Electronic address: takeshim@nibb.ac.jp.
authors.forename: Takeshi
authors.initials: T
authors.lastname: Mizutani
authors.affiliation: Department of Basic Biology, Faculty of Life Science, SOKENDAI (Graduate University for Advanced Studies), 5-1 Higashiyama, Myodaiji, Okazaki, Aichi 444-8787, Japan; Okazaki Institute for Integrative Bioscience, National Institute for Basic Biology, National Institutes of Natural Sciences, 5-1 Higashiyama, Myodaiji, Okazaki, Aichi 444-8787, Japan. Electronic address: ogino@nibb.ac.jp.
authors.forename: Yukiko
authors.initials: Y
authors.lastname: Ogino
authors.affiliation: Department of Veterinary Medicine, Faculty of Agriculture, Tottori University, Koyama, Tottori 680-8553, Japan. Electronic address: ohta1022@gmail.com.
authors.forename: Yasuhiko
authors.initials: Y
authors.lastname: Ohta
authors.affiliation: Department of Paraclinical Sciences, University of Pretoria, Private Bag 04, Onderstepoort 0110, South Africa. Electronic address: jan.myburgh@up.ac.za.
authors.forename: Jan
authors.initials: J
authors.lastname: Myburgh
authors.affiliation: Department of Botany & Zoology, University of Stellenbosch, Stellenbosch 7600, South Africa. Electronic address: jhvw@sun.ac.za.
authors.forename: Johannes H
authors.initials: JH
authors.lastname: van Wyk
authors.affiliation: Department of Obstetrics and Gynecology, Medical University of South Carolina, and Marine Biomedicine and Environmental Science Center, Hollings Marine Laboratory, Charleston, SC 29412, USA.
authors.forename: Louis J
authors.initials: LJ
authors.lastname: Guillette
authors.affiliation: Department of Basic Biology, Faculty of Life Science, SOKENDAI (Graduate University for Advanced Studies), 5-1 Higashiyama, Myodaiji, Okazaki, Aichi 444-8787, Japan; Okazaki Institute for Integrative Bioscience, National Institute for Basic Biology, National Institutes of Natural Sciences, 5-1 Higashiyama, Myodaiji, Okazaki, Aichi 444-8787, Japan. Electronic address: miyagawa@nibb.ac.jp.
authors.forename: Shinichi
authors.initials: S
authors.lastname: Miyagawa
authors.affiliation: Department of Basic Biology, Faculty of Life Science, SOKENDAI (Graduate University for Advanced Studies), 5-1 Higashiyama, Myodaiji, Okazaki, Aichi 444-8787, Japan; Okazaki Institute for Integrative Bioscience, National Institute for Basic Biology, National Institutes of Natural Sciences, 5-1 Higashiyama, Myodaiji, Okazaki, Aichi 444-8787, Japan. Electronic address: taisen@nibb.ac.jp.
authors.forename: Taisen
authors.initials: T
authors.lastname: Iguchi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D005075:Biological Evolution; D003001:Cloning, Molecular; D003198:Computer Simulation; D047628:Estrogen Receptor alpha / Q000737:chemistry / Q000378:metabolism; D047629:Estrogen Receptor beta / Q000378:metabolism; D008024:Ligands; D008116:Lizards / Q000378:metabolism*; D008958:Models, Molecular; D000072417:Protein Domains; D011960:Receptors, Estrogen / Q000737:chemistry / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D047628:Estrogen Receptor alpha; D047629:Estrogen Receptor beta; D008024:Ligands; D011960:Receptors, Estrogen
keywords: Estrogen receptor; Estrogen sensitivity; Evolution of estrogen receptor; Squamates