Human Bacterial Artificial Chromosome (BAC) Transgenesis Fully Rescues Noradrenergic Function in Dopamine beta-Hydroxylase Knockout Mice

• Type of Material: Article
• Authors: Joseph Cubells, Emory University; Jason Schroeder, Emory University; Elizabeth S. Barrie, Ohio State University; Daniel F. Manvich, Emory University; Wolfgang Sadee, Ohio State University; Tiina Berg, Emory University; Kristina Mercer, Emory University; Taylor A. Stowe, Emory University; L. Cameron Liles, Emory University; Katherine E. Squires, Emory University; Andrew Mezher, Emory University; Patrick Curtin, Emory University; Dannie L. Perdomo, Emory University; Patricia Szot, VA Puget Sound Health Care System; David Weinshenker, Emory University
• Language: English
• Date: 2016-05-05
• Publisher: Public Library of Science
• Publication Version: Final Published Version
• Copyright Statement: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose.
• License: Creative Commons Creative Commons CC0 1.0 Universal
• Final Published Version (URL): http://dx.doi.org/10.1371/journal.pone.0154864
• Title of Journal or Parent Work: PLoS ONE
• ISSN: 1932-6203
• Volume: 11
• Issue: 5
• Start Page: e0154864
• End Page: e0154864
• Grant/Funding Information: This work was supported by the National Institute of Neurological Disorders and Stroke (NS072712 to JFC; http://www.ninds.nih.gov/), the National Institute on Drug Abuse (DA027535 and DA038453 to DW and DA015040 to DLP; https://www.drugabuse.gov/), the National Institute of General Medical Sciences (GM 092655 to WS; https://www.nigms.nih.gov/), the Ohio State University (Distinguished University Fellowship to ESB; http://www.gradsch.osu.edu/fellowship-program.html), and the Department of Veterans Affairs VISN 20 (Northwest Network) Mental Illness Research, Education, and Clinical Center (MIRECC) to PS.
• Keywords: Multidisciplinary Sciences; Gene expression; NORADRENALINE; PARKINSONS-DISEASE; Neurons; Science & Technology - Other Topics; Catecholamines; EXPRESSION; BLOOD-PRESSURE; MESSENGER-RNA; GENE; PROMOTER; LACKING NOREPINEPHRINE; Mice; Phenotypes; Genome-wide association studies; NOREPINEPHRINE-DEFICIENT MICE; ALZHEIMERS-DISEASE; Oligonucleotides; Science & Technology; Biological locomotion
• Research Categories: Health Sciences, General; Biology, Genetics; Biology, Neuroscience

Abstract

Dopamine β-hydroxylase (DBH) converts dopamine (DA) to norepinephrine (NE) in noradrenergic/adrenergic cells. DBH deficiency prevents NE production and causes sympathetic failure, hypotension and ptosis in humans and mice; DBH knockout (Dbh -/-) mice reveal other NE deficiency phenotypes including embryonic lethality, delayed growth, and behavioral defects. Furthermore, a single nucleotide polymorphism (SNP) in the human DBH gene promoter (-970C>T; rs1611115) is associated with variation in serum DBH activity and with several neurological- and neuropsychiatric-related disorders, although its impact on DBH expression is controversial. Phenotypes associated with DBH deficiency are typically treated with L-3,4-dihydroxyphenylserine (DOPS), which can be converted to NE by aromatic acid decarboxylase (AADC) in the absence of DBH. In this study, we generated transgenic mice carrying a human bacterial artificial chromosome (BAC) encompassing the DBH coding locus as well as ~45 kb of upstream and ~107 kb of downstream sequence to address two issues. First, we characterized the neuroanatomical, neurochemical, physiological, and behavioral transgenic rescue of DBH deficiency by crossing the BAC onto a Dbh -/- background. Second, we compared human DBH mRNA abundance between transgenic lines carrying either a "C" or a "T" at position -970. The BAC transgene drove human DBH mRNA expression in a pattern indistinguishable from the endogenous gene, restored normal catecholamine levels to the peripheral organs and brain of Dbh -/- mice, and fully rescued embryonic lethality, delayed growth, ptosis, reduced exploratory activity, and seizure susceptibility. In some cases, transgenic rescue was superior to DOPS. However, allelic variation at the rs1611115 SNP had no impact on mRNA levels in any tissue. These results indicate that the human BAC contains all of the genetic information required for tissue-specific, functional expression of DBH and can rescue all measured Dbh deficiency phenotypes, but did not reveal an impact of the rs11115 variant on DBH expression in mice.