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

• 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
• Journal Title: PLoS ONE
• Volume: Volume 11, Number 5
• Publisher: Public Library of Science | 2016-05-05, Pages e0154864-e0154864
• Type of Work: Article | Final Publisher PDF
• Publisher DOI: http://doi.org/10.1371/journal.pone.0154864
• Subjects: Biology, Genetics; Biology, Neuroscience; Health Sciences, General
• 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

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.