Structural and functional promoter analyses of epilepsy-relevant genes as basis for targeted expression interference

Abstract

<strong>Background:</strong> Epilepsies are a group of brain disorders characterized by recurrent seizures. A fraction of those has a well defined genetic aetiology including monogenic epilepsies, and others are the result of acquired factors such as head trauma. Interestingly, the fact that (i) monogenic epilepsies can present with a wide variety of clinical manifestations and that (ii) the risk of epilepsy increases after an acquired factor in individuals with a familial history of epilepsy strongly suggests a genetic susceptibility. GWAS showed that most trait-associated SNPs lie in non-coding genomic regions. Thus, they may operate by altering gene expression. Functional characterization of non-coding SNPs is challenging. Hippocampal biopsies from patients, in combination with in vitro reporter assays are of great value to determine the functionality of regulatory SNPs in the context of epilepsy. A new technology based on CRISPR can manipulate gene expression by targeting non-coding regulatory elements and be used therefore to identify, validate, and functionally elucidate vulnerability loci in the genome. <br /> <strong>Aims:</strong> (1) Investigate if candidate SNPs in non-coding genomic regions of genes previously associated with epilepsy alter gene expression in hippocampi of temporal lobe epilepsy (TLE) patients indicating genetic susceptibility, (2) reveal the molecular promoter control mechanism that confers distinct allelic expression and (3) engineer a CRISPR toolbox to manipulate the promoter activity of the epileptogenesis relevant calcium channel cacna1h, as a way to further understand the genomic architecture and its importance on gene expression. <br /> <strong>Results and conclusion:</strong> The first publication demonstrated that promoter haplotypes of ALDH5A1 confer differential gene expression in the hippocampi of TLE patients and that the promoter might be under regulation of the transcription factor Egr1. The second publication demonstrated that a GWAS-derived SNP in the SCN1A gene, confers differential expression in hippocampi of TLE patients and that the SNP lies in an intronic region that contains promoter-like activity. In the third publication, we showed that CRISPR modulates the promoter activity of cacna1h in a very specific manner, resulting in changes in gene expression. This work contributes to a better understanding of the importance of genetic regulatory regions in epilepsy and encourages the use of CRISPR methods in the field.