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Humanization of Drosophila Gαo to Model GNAO1 Paediatric Encephalopathies

Translational Research Center in Oncohaematology, Department of Cell Physiology and Metabolism, Faculty of Medicine, University of Geneva, 1211 Geneva, Switzerland
School of Biomedicine, Far Eastern Federal University, 690690 Vladivostok, Russia
Author to whom correspondence should be addressed.
Biomedicines 2020, 8(10), 395;
Received: 24 August 2020 / Revised: 30 September 2020 / Accepted: 2 October 2020 / Published: 6 October 2020
(This article belongs to the Section Neurobiology and Neurologic Disease)
Several hundred genes have been identified to contribute to epilepsy—the disease affecting 65 million people worldwide. One of these genes is GNAO1 encoding Gαo, the major neuronal α-subunit of heterotrimeric G proteins. An avalanche of dominant de novo mutations in GNAO1 have been recently described in paediatric epileptic patients, suffering, in addition to epilepsy, from motor dysfunction and developmental delay. Although occurring in amino acids conserved from humans to Drosophila, these mutations and their functional consequences have only been poorly analysed at the biochemical or neuronal levels. Adequate animal models to study the molecular aetiology of GNAO1 encephalopathies have also so far been lacking. As the first step towards modeling the disease in Drosophila, we here describe the humanization of the Gαo locus in the fruit fly. A two-step CRISPR/Cas9-mediated replacement was conducted, first substituting the coding exons 2–3 of Gαo with respective human GNAO1 sequences. At the next step, the remaining exons 4–7 were similarly replaced, keeping intact the gene Cyp49a1 embedded in between, as well as the non-coding exons, exon 1 and the surrounding regulatory sequences. The resulting flies, homozygous for the humanized GNAO1 loci, are viable and fertile without any visible phenotypes; their body weight, locomotion, and longevity are also normal. Human Gαo-specific antibodies confirm the endogenous-level expression of the humanized Gαo, which fully replaces the Drosophila functions. The genetic model we established will make it easy to incorporate encephalopathic GNAO1 mutations and will permit intensive investigations into the molecular aetiology of the human disease through the powerful toolkit of Drosophila genetics. View Full-Text
Keywords: paediatric encephalopathy; GNAO1; G protein; humanization; Drosophila; disease model paediatric encephalopathy; GNAO1; G protein; humanization; Drosophila; disease model
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MDPI and ACS Style

Savitsky, M.; Solis, G.P.; Kryuchkov, M.; Katanaev, V.L. Humanization of Drosophila Gαo to Model GNAO1 Paediatric Encephalopathies. Biomedicines 2020, 8, 395.

AMA Style

Savitsky M, Solis GP, Kryuchkov M, Katanaev VL. Humanization of Drosophila Gαo to Model GNAO1 Paediatric Encephalopathies. Biomedicines. 2020; 8(10):395.

Chicago/Turabian Style

Savitsky, Mikhail, Gonzalo P. Solis, Mikhail Kryuchkov, and Vladimir L. Katanaev 2020. "Humanization of Drosophila Gαo to Model GNAO1 Paediatric Encephalopathies" Biomedicines 8, no. 10: 395.

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