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Gene expression profiling of teloHAEC cells with CRISPR-Cas9-mediated deletion of candidate enhancer regions

ABSTRACT: Functional consequences of genetic variation in the non-coding human genome are difficult to ascertain despite demonstrated associations to common, complex disease traits. To elucidate properties of functional non-coding SNPs with effects in human endothelial cells (EC), we utilized our previous molecular Quantitative Trait Locus (molQTL) analysis for transcription factor binding, chromatin accessibility, and H3K27 acetylation to nominate a set of likely functional non-coding SNPs. Together with information from Genome-Wide Association Studies (GWAS) for vascular disease traits, we tested the ability of more than 34,000 genetic variants to perturb enhancer function in ECs using the highly multiplexed reporter assay. Based on these results, CRISPR-mediated deletion followed by mRNA-Seq was carried out for SNP-containing regulatory elements near SMAD3 and LDAH (C2orf43) in order to functionally verify the target genes of the regulatory elements.

ORGANISM(S): Homo sapiens

PROVIDER: GSE191253 | GEO | 2021/12/21

REPOSITORIES: GEO

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Project description:Genetics of gene expression (eQTLs or expression QTLs) has proved an indispensable tool for understanding biological pathways and pathomechanisms of trait associated SNPs. However, power of most genome-wide eQTL studies is still limited. We performed a large eQTL study in peripheral blood mononuclear cells of 2,112 individuals increasing the power to detect trans-effects genome-wide. Going beyond univariate SNP-transcript associations, we analyse relations of eQTLs to biological pathways, polygenetic effects of expression regulation, trans-clusters, and enrichment of co-localised functional elements. We found eQTLs for about 85% of analysed genes, 18% of genes were trans-regulated. Local eSNPs were enriched up to a distance of 5 MB to the transcript challenging typically implemented ranges of cis-regulations. Pathway enrichment within regulated genes of GWAS-related eSNPs supported functional relevance of identified eQTLs. We demonstrate that nearest genes of GWAS-SNPs might often be misleading functional candidates. We identified novel trans-clusters of potential functional relevance for GWAS-SNPs of several phenotypes including obesity-related traits, HDL-cholesterol levels, and haematological phenotypes. We used chromatin immunoprecipitation data for demonstrating biological effects. We show for strongly heritable transcripts that a considerable gap still exists between total heritability resulting from all trans-chromosomes and explained variance of all identified trans-eSNPs. In contrast, the vast majority of most cis-heritability of these genes is already explained. Dissection of co-localised functional elements indicated a prominent role of SNPs in loci of pseudogenes and non-coding RNAs for the regulation of coding genes. In summary, our study substantially increases the catalogue of human eQTLs and improves our understanding of the complex genetic regulation of gene-expression, pathways and disease related processes. Gene expression from human blood mononuclear cells from individuals of the Leipzig LIFE Heart Study was analyzed applying Illumina HT-12 v4 Expression BeadChips. After preprocessing, 28,295 expression probes for 2,112 individuals remained for analysis. In a population-based analysis, we identified associations between DNA variants and RNA expression levels and characterized these findings.

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Gene expression profiling of teloHAEC cells with CRISPR-Cas9-mediated deletion of candidate enhancer regions

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