title: Rapid fluorescent detection of (anti)androgens with spiggin-gfp medaka.
abstract: Widespread environmental antiandrogen contamination has been associated with negative impacts on biodiversity and human health. In particular, many pesticides are antiandrogenic, creating a need for robust and sensitive environmental monitoring. Our aim was to develop a sensitive and specific transgenic medaka (Oryzias latipes) model bearing an androgen responsive fluorescent reporter construct for whole organism-based environmental screening of pro- and antiandrogens. We analyzed the 5' regions of the androgen responsive three-spined stickleback (Gasterosteus aculeatus) spiggin genes in silico, revealing conserved blocks of sequence harboring androgen response elements. Identified putative promoters were cloned upstream of GFP. Germinal transgenesis with spg1-gfp led to stable medaka lines. GFP induction was exclusive to the kidney, the site of spiggin protein production in sticklebacks. Significant GFP expression was induced by three or four-day androgen treatment of newly hatched fry, but not by estrogens, mineralocorticoids, glucocorticoids or progestogens. The model responded dose-dependently to androgens, with highest sensitivity to 17MT (1.5 μg/L). In addition to flutamide, the biocides fenitrothion, vinclozolin and linuron significantly inhibited 17MT-induced GFP induction, validating the model for detection of antiandrogens. The spg1-gfp medaka model provides a sensitive, specific, and physiologically pertinent biosensor system for analyzing environmental androgen activity.
authors.affiliation: WatchFrog S.A., 1 rue Pierre Fontaine, 91000 Evry, France.
authors.forename: Anthony
authors.initials: A
authors.lastname: Sébillot
authors.forename: Pauliina
authors.initials: P
authors.lastname: Damdimopoulou
authors.forename: Yukiko
authors.initials: Y
authors.lastname: Ogino
authors.forename: Petra
authors.initials: P
authors.lastname: Spirhanzlova
authors.forename: Shinichi
authors.initials: S
authors.lastname: Miyagawa
authors.forename: David
authors.initials: D
authors.lastname: Du Pasquier
authors.forename: Nora
authors.initials: N
authors.lastname: Mouatassim
authors.forename: Taisen
authors.initials: T
authors.lastname: Iguchi
authors.forename: Gregory F
authors.initials: GF
authors.lastname: Lemkine
authors.forename: Barbara A
authors.initials: BA
authors.lastname: Demeneix
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Tindall
mesh_terms: D000726:Androgen Antagonists / Q000378:metabolism*; D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D003001:Cloning, Molecular; D029941:Fish Proteins / Q000378:metabolism*; D005453:Fluorescence; D049452:Green Fluorescent Proteins / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009990:Oryzias / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011944:Receptors, Androgen / Q000378:metabolism; D023701:Smegmamorpha
chemical_list: D000726:Androgen Antagonists; D029941:Fish Proteins; D011944:Receptors, Androgen; C429543:spiggin; D049452:Green Fluorescent Proteins