title: Ferredoxin 1b (Fdx1b) Is the Essential Mitochondrial Redox Partner for Cortisol Biosynthesis in Zebrafish.
abstract: Mitochondrial cytochrome P450 (CYP) enzymes rely on electron transfer from the redox partner ferredoxin 1 (FDX1) for catalytic activity. Key steps in steroidogenesis require mitochondrial CYP enzymes and FDX1. Over 30 ferredoxin mutations have been explored in vitro; however, no spontaneously occurring mutations have been identified in humans leaving the impact of FDX1 on steroidogenesis in the whole organism largely unknown. Zebrafish are an important model to study human steroidogenesis, because they have similar steroid products and endocrine tissues. This study aimed to characterize the influence of ferredoxin on steroidogenic capacity in vivo by using zebrafish. Zebrafish have duplicate ferredoxin paralogs: fdx1 and fdx1b. Although fdx1 was observed throughout development and in most tissues, fdx1b was expressed after development of the zebrafish interrenal gland (counterpart to the mammalian adrenal gland). Additionally, fdx1b was restricted to adult steroidogenic tissues, such as the interrenal, gonads, and brain, suggesting that fdx1b was interacting with steroidogenic CYP enzymes. By using transcription activator-like effector nucleases, we generated fdx1b mutant zebrafish lines. Larvae with genetic disruption of fdx1b were morphologically inconspicuous. However, steroid hormone analysis by liquid chromatography tandem mass spectrometry revealed fdx1b mutants failed to synthesize glucocorticoids. Additionally, these mutants had an up-regulation of the hypothalamus-pituitary-interrenal axis and showed altered dark-light adaptation, suggesting impaired cortisol signaling. Antisense morpholino knockdown confirmed Fdx1b is required for de novo cortisol biosynthesis. In summary, by using zebrafish, we generated a ferredoxin knockout model system, which demonstrates for the first time the impact of mitochondrial redox regulation on glucocorticoid biosynthesis in vivo.
authors.affiliation: Centre for Endocrinology, Diabetes, and Metabolism (A.G., S.P., M.W., A.Z., A.E.T., D.M.O., N.K.), School of Clinical and Experimental Medicine (F.M.), College of Medical and Dental Sciences, University of Birmingham, Birmingham B15 2TT, United Kingdom; and Department of Oncology and Metabolism (N.K.), University of Sheffield, Sheffield S10 2TG, United Kingdom.
authors.forename: Aliesha
authors.initials: A
authors.lastname: Griffin
authors.affiliation: Centre for Endocrinology, Diabetes, and Metabolism (A.G., S.P., M.W., A.Z., A.E.T., D.M.O., N.K.), School of Clinical and Experimental Medicine (F.M.), College of Medical and Dental Sciences, University of Birmingham, Birmingham B15 2TT, United Kingdom; and Department of Oncology and Metabolism (N.K.), University of Sheffield, Sheffield S10 2TG, United Kingdom.
authors.forename: Silvia
authors.initials: S
authors.lastname: Parajes
authors.affiliation: Centre for Endocrinology, Diabetes, and Metabolism (A.G., S.P., M.W., A.Z., A.E.T., D.M.O., N.K.), School of Clinical and Experimental Medicine (F.M.), College of Medical and Dental Sciences, University of Birmingham, Birmingham B15 2TT, United Kingdom; and Department of Oncology and Metabolism (N.K.), University of Sheffield, Sheffield S10 2TG, United Kingdom.
authors.forename: Meltem
authors.initials: M
authors.lastname: Weger
authors.affiliation: Centre for Endocrinology, Diabetes, and Metabolism (A.G., S.P., M.W., A.Z., A.E.T., D.M.O., N.K.), School of Clinical and Experimental Medicine (F.M.), College of Medical and Dental Sciences, University of Birmingham, Birmingham B15 2TT, United Kingdom; and Department of Oncology and Metabolism (N.K.), University of Sheffield, Sheffield S10 2TG, United Kingdom.
authors.forename: Andreas
authors.initials: A
authors.lastname: Zaucker
authors.affiliation: Centre for Endocrinology, Diabetes, and Metabolism (A.G., S.P., M.W., A.Z., A.E.T., D.M.O., N.K.), School of Clinical and Experimental Medicine (F.M.), College of Medical and Dental Sciences, University of Birmingham, Birmingham B15 2TT, United Kingdom; and Department of Oncology and Metabolism (N.K.), University of Sheffield, Sheffield S10 2TG, United Kingdom.
authors.forename: Angela E
authors.initials: AE
authors.lastname: Taylor
authors.affiliation: Centre for Endocrinology, Diabetes, and Metabolism (A.G., S.P., M.W., A.Z., A.E.T., D.M.O., N.K.), School of Clinical and Experimental Medicine (F.M.), College of Medical and Dental Sciences, University of Birmingham, Birmingham B15 2TT, United Kingdom; and Department of Oncology and Metabolism (N.K.), University of Sheffield, Sheffield S10 2TG, United Kingdom.
authors.forename: Donna M
authors.initials: DM
authors.lastname: O'Neil
authors.affiliation: Centre for Endocrinology, Diabetes, and Metabolism (A.G., S.P., M.W., A.Z., A.E.T., D.M.O., N.K.), School of Clinical and Experimental Medicine (F.M.), College of Medical and Dental Sciences, University of Birmingham, Birmingham B15 2TT, United Kingdom; and Department of Oncology and Metabolism (N.K.), University of Sheffield, Sheffield S10 2TG, United Kingdom.
authors.forename: Ferenc
authors.initials: F
authors.lastname: Müller
authors.affiliation: Centre for Endocrinology, Diabetes, and Metabolism (A.G., S.P., M.W., A.Z., A.E.T., D.M.O., N.K.), School of Clinical and Experimental Medicine (F.M.), College of Medical and Dental Sciences, University of Birmingham, Birmingham B15 2TT, United Kingdom; and Department of Oncology and Metabolism (N.K.), University of Sheffield, Sheffield S10 2TG, United Kingdom.
authors.forename: Nils
authors.initials: N
authors.lastname: Krone
mesh_terms: D000818:Animals; D001921:Brain / Q000378:metabolism; D002853:Chromatography, Liquid; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D005288:Ferredoxins / Q000235:genetics*; D018507:Gene Expression Regulation, Developmental; D055786:Gene Knockout Techniques; D005938:Glucocorticoids / Q000096:biosynthesis; D006066:Gonads / Q000378:metabolism; D006854:Hydrocortisone / Q000096:biosynthesis*; D017403:In Situ Hybridization; D007401:Interrenal Gland / Q000378:metabolism; D007814:Larva / Q000235:genetics / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism*; D010084:Oxidation-Reduction; D060888:Real-Time Polymerase Chain Reaction; D053719:Tandem Mass Spectrometry; D015027:Zebrafish / Q000235:genetics / Q000378:metabolism; D029961:Zebrafish Proteins / Q000235:genetics*
chemical_list: D005288:Ferredoxins; D005938:Glucocorticoids; D029961:Zebrafish Proteins; D003577:Cytochrome P-450 Enzyme System; D006854:Hydrocortisone