Employing CRISPR/Cas9 and Saccharomyces cerevisiae to understand energetic aspects of schizophrenia and antipsychotic treatment from a proteomic perspective

Abstract

Schizophrenia is a severe and debilitating disorder known to affect approximately 1% of the world population. Represents a complex syndrome thought to be of neurodevelopmental origin and is manifested through a wide range of severe symptoms. Several proteomic, pharmacological and genetic studies have been conducted to try elucidating the underlying mechanisms associated with the disorder and they reported abnormalities in mitochondrial function and energy metabolism in schizophrenia. Interestingly, the antipsychotic drug Clozapine, which has great clinical efficacy, was shown to improve glucose uptake in oligodendrocytes, indicating that in addition to rebalance neurotransmission, this drug acts on the energy metabolism of those cells which may in turn improve neuronal connectivity. Among the several proteins related to energy metabolism found altered in schizophrenia, pyruvate dehydrogenase E1 component subunit alpha stands out, as it is a mitochondrial protein that represents the primary link between glycolysis and the tricarboxylic acid cycle, by converting pyruvate into acetyl-CoA. This evidence along with reports that patients with schizophrenia present lower levels of pyruvate in some brain regions reveals that hampered overall energy metabolism is an important feature of schizophrenia. The use of yeasts to model energy metabolism dysfunction is of extreme value since human and yeast mitochondrial proteomes are so similar that yeast is very well-suited to determine the primary effects on mitochondrial energy transduction and physiology of disease-linked mutations. The use of Crispr/Cas9 technique revolutionized the field of gene editing and expression, since enables the efficient deletion and/or substitution of whole genes without the need for protein engineering. With this is mind, we propose to perform a genetic knock-out using the Crispr/Cas9 technique of the nuclear-coded gene of the pyruvate dehydrogenase E1 component subunit alpha in Saccharomyces cerevisiae and evaluate the effects of this knock-out on the cell energy metabolism before and after treatment with Clozapine. The knowledge in Crispr/Cas9 tool acquired will be of extreme value to bring this technique to our laboratory in Brazil and to implement protocols and a place for execution of future projects using Crispr/Cas9 in different cell models, such as yeast and iPSCs.