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Author Notes:

E-mail: dweinshenker@genetics.emory.edu

Conceived and designed the experiments: JFC DW.

Performed the experiments: JPS ESB DFM WS TB KM TAS LCL KES AM PC DLP PS.

Analyzed the data: JFC JPS ESB DFM WS TAS LCL KES KM PC DLP PS DW.

Contributed reagents/materials/analysis tools: ESB WS TB KM PS DW.

Wrote the paper: JFC ESB WS DFM DLP PS JPS DW.

We thank Ellen Hess and Doug Bernhard for assistance with the HPLC experiments, and Dr. Michael J. Kuhar, Principal Investigator for T32 DA015040, which supported DLP for her role in the project.

The authors have declared that no competing interests exist.

The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Subjects:

Research Funding:

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:

  • Science & Technology
  • Multidisciplinary Sciences
  • Science & Technology - Other Topics
  • NOREPINEPHRINE-DEFICIENT MICE
  • MESSENGER-RNA
  • LACKING NOREPINEPHRINE
  • PARKINSONS-DISEASE
  • ALZHEIMERS-DISEASE
  • BLOOD-PRESSURE
  • EXPRESSION
  • GENE
  • NORADRENALINE
  • PROMOTER
  • Mice
  • Neurons
  • Biological locomotion
  • Gene expression
  • Phenotypes
  • Catecholamines
  • Genome-wide association studies
  • Oligonucleotides

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

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Journal Title:

PLoS ONE

Volume:

Volume 11, Number 5

Publisher:

, Pages e0154864-e0154864

Type of Work:

Article | Final Publisher PDF

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.

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This is an Open Access work distributed under the terms of the Creative Commons Universal : Public Domain Dedication License (http://creativecommons.org/publicdomain/zero/1.0/).

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