title: Glutathione redox dynamics and expression of glutathione-related genes in the developing embryo.
abstract: Embryonic development involves dramatic changes in cell proliferation and differentiation that must be highly coordinated and tightly regulated. Cellular redox balance is critical for cell fate decisions, but it is susceptible to disruption by endogenous and exogenous sources of oxidative stress. The most abundant endogenous nonprotein antioxidant defense molecule is the tripeptide glutathione (γ-glutamylcysteinylglycine, GSH), but the ontogeny of GSH concentration and redox state during early life stages is poorly understood. Here, we describe the GSH redox dynamics during embryonic and early larval development (0-5 days postfertilization) in the zebrafish (Danio rerio), a model vertebrate embryo. We measured reduced and oxidized glutathione using HPLC and calculated the whole embryo total glutathione (GSHT) concentrations and redox potentials (Eh) over 0-120 h of zebrafish development (including mature oocytes, fertilization, midblastula transition, gastrulation, somitogenesis, pharyngula, prehatch embryos, and hatched eleutheroembryos). GSHT concentration doubled between 12h postfertilization (hpf) and hatching. The GSH Eh increased, becoming more oxidizing during the first 12h, and then oscillated around -190 mV through organogenesis, followed by a rapid change, associated with hatching, to a more negative (more reducing) Eh (-220 mV). After hatching, Eh stabilized and remained steady through 120 hpf. The dynamic changes in GSH redox status and concentration defined discrete windows of development: primary organogenesis, organ differentiation, and larval growth. We identified the set of zebrafish genes involved in the synthesis, utilization, and recycling of GSH, including several novel paralogs, and measured how expression of these genes changes during development. Ontogenic changes in the expression of GSH-related genes support the hypothesis that GSH redox state is tightly regulated early in development. This study provides a foundation for understanding the redox regulation of developmental signaling and investigating the effects of oxidative stress during embryogenesis.
authors.affiliation: Biology Department, Woods Hole Oceanographic Institution, Woods Hole, MA 02543, USA; Division of Environmental Health, Department of Public Health, School of Public Health and Health Sciences, University of Massachusetts, Amherst, MA 01003, USA. Electronic address: aliciat@schoolph.umass.edu.
authors.forename: Alicia R
authors.initials: AR
authors.lastname: Timme-Laragy
authors.affiliation: Biology Department, Woods Hole Oceanographic Institution, Woods Hole, MA 02543, USA.
authors.forename: Jared V
authors.initials: JV
authors.lastname: Goldstone
authors.affiliation: Division of Pulmonary, Allergy/Immunology, Cystic Fibrosis and Sleep, Department of Pediatrics, Emory University, Atlanta, GA 30322, USA.
authors.forename: Barry R
authors.initials: BR
authors.lastname: Imhoff
authors.affiliation: Biology Department, Woods Hole Oceanographic Institution, Woods Hole, MA 02543, USA.
authors.forename: John J
authors.initials: JJ
authors.lastname: Stegeman
authors.affiliation: Biology Department, Woods Hole Oceanographic Institution, Woods Hole, MA 02543, USA.
authors.forename: Mark E
authors.initials: ME
authors.lastname: Hahn
authors.affiliation: Division of Pulmonary, Allergy/Immunology, Cystic Fibrosis and Sleep, Department of Pediatrics, Emory University, Atlanta, GA 30322, USA.
authors.forename: Jason M
authors.initials: JM
authors.lastname: Hansen
mesh_terms: D000818:Animals; D004625:Embryo, Nonmammalian; D018507:Gene Expression Regulation, Developmental; D005978:Glutathione / Q000235:genetics* / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis; D010084:Oxidation-Reduction; D015027:Zebrafish / Q000196:embryology* / Q000235:genetics / Q000378:metabolism*
chemical_list: D005978:Glutathione
keywords: Antioxidant; Embryonic development; Free radicals; GSH; GSSG; Gcl; Gclc; Gclm; Gene expression; Glutathione; Gss; Oxidative stress; PCA; ROS; Redox; Zebrafish; glutamate–cysteine ligase; glutamate–cysteine ligase catalytic subunit; glutamate–cysteine ligase modifier subunit; glutathione disulfide; glutathione synthetase; perchloric acid; reactive oxygen species; reduced glutathione