A novel vector for transgenesis in the rat CNS

T. Peter Lopez, Kurt Giles, Brittany Dugger, Abby Oehler, Carlo Condello, Zuzana Krejciova, Julian A. Castaneda, George A. Carlson, Stanley B. Prusiner

Research output: Contribution to journalArticle

Abstract

The larger brain of the rat enables a much greater repertoire of complex behaviors than mice, likely making rats preferential for investigating neurodegeneration. Because molecular tools for specific expression of transgenes in the rat brain are sparse, we chose Prnp encoding the prion protein (PrP) to develop a novel vector to drive transgene expression in the rat brain. We compared the rat Prnp sequence with mouse and Syrian hamster Prnp sequences, identifying conserved genetic elements and hypothesizing that these elements would be able to drive neuronal transgene expression. We investigated this by generating a vector termed RaPrnp that encompasses portions of the rat Prnp gene. Importantly, we replaced the rat Prnp open reading frame (ORF) with a cloning site for rapid and seamless In-Fusion cloning. To validate the in vivo neuronal specificity of the RaPrnp vector in rats, we generated stable RaPrnp-LacZ/enhanced green fluorescent protein (EGFP) transgenic (Tg) rat lines, which led to robust LacZ activity and high EGFP fluorescence in the central nervous system of embryos and adult animals. Next, we restored the rat Prnp ORF and generated multiple Tg(RaPrnp-PrP) lines, demonstrating that overexpression of Prnp accelerates the onset of scrapie. While the incubation time in wild-type (WT) rats was 175 ± 3 days post inoculation (dpi), one line, Tg2919, overexpressed RaPrPC at 4.4-fold and exhibited a reduced incubation time of 149 ± 2 dpi. The second line, Tg2922, overexpressed RaPrPC at 9.7-fold compared with WT animals and had an incubation time of 112 ± 0 dpi. Tg2922 rats inoculated with rat RML showed extensive vacuolation of the brainstem in contrast to WT and Tg2919 animals in which vacuolation was most prominent in the hippocampus and striatum as well as the motor and sensory cortices. It is possible that construction of Tg rats with modified phenotypes will prove more advantageous than mice for neurodegeneration studies.

Original languageEnglish (US)
Number of pages1
JournalActa neuropathologica communications
Volume5
Issue number1
DOIs
StatePublished - Nov 21 2017

Fingerprint

Gene Transfer Techniques
Transgenes
Transgenic Rats
Wild Animals
Open Reading Frames
Organism Cloning
Brain
Scrapie
Conserved Sequence
Mesocricetus
Brain Stem
Hippocampus
Embryonic Structures
Central Nervous System

Keywords

  • Neurodegenerative disease
  • Prnp
  • Scrapie
  • Vector

ASJC Scopus subject areas

  • Pathology and Forensic Medicine
  • Clinical Neurology
  • Cellular and Molecular Neuroscience

Cite this

Lopez, T. P., Giles, K., Dugger, B., Oehler, A., Condello, C., Krejciova, Z., ... Prusiner, S. B. (2017). A novel vector for transgenesis in the rat CNS. Acta neuropathologica communications, 5(1). https://doi.org/10.1186/s40478-017-0484-y

A novel vector for transgenesis in the rat CNS. / Lopez, T. Peter; Giles, Kurt; Dugger, Brittany; Oehler, Abby; Condello, Carlo; Krejciova, Zuzana; Castaneda, Julian A.; Carlson, George A.; Prusiner, Stanley B.

In: Acta neuropathologica communications, Vol. 5, No. 1, 21.11.2017.

Research output: Contribution to journalArticle

Lopez, TP, Giles, K, Dugger, B, Oehler, A, Condello, C, Krejciova, Z, Castaneda, JA, Carlson, GA & Prusiner, SB 2017, 'A novel vector for transgenesis in the rat CNS', Acta neuropathologica communications, vol. 5, no. 1. https://doi.org/10.1186/s40478-017-0484-y
Lopez, T. Peter ; Giles, Kurt ; Dugger, Brittany ; Oehler, Abby ; Condello, Carlo ; Krejciova, Zuzana ; Castaneda, Julian A. ; Carlson, George A. ; Prusiner, Stanley B. / A novel vector for transgenesis in the rat CNS. In: Acta neuropathologica communications. 2017 ; Vol. 5, No. 1.
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