Gene x Lifestyle Interactions in Cardiometabolic Disease
Final Report Abstract
Since GWAS and Gene x Environment interaction studies alone fail to fully explain phenotypic variations for e.g BMI or different cardio metabolic outcomes, it is very likely that epigenetic modification may represent the mediator between genetic predisposition and our individual phenotype. To illuminate the underlying functional mechanism behind such identified associations a combined approach using in silico functional annotation and an in vitro model for Gene x Environment interactions using CRISPR Cas9 genome editing was developed. The approach was tested in a pilot study investigating the role of two genetic variants (rs2289669, rs8065082) in the SLC47A1 gene locus, encoding for the multidrug and toxin extrusion 1 transporter (MATE1), on genotype dependent metformin response in a human liver cell line (Huh7). In silico functional annotation including all SNPs in tight LD (r2>0.8) with the candidate variants further support their functional potential by their location in a predicted enhancer region in liver and close to potential transcription factor binding sites. Our studies further show a significant high methylation level of rs8065082 gene locus in Huh7 wild type cells not carrying the SNP allele, suggesting the variant could be involved in transcriptional regulation. For the in vitro model two gRNAs per SNP target were designed, cloned into a Cas9 plasmid vector and optimized for high transfection efficiency in Huh-7 cells. To generate SLC47A1/MATE1 KO (~2kb) and SNP KOs (~40bp) cells, predesigned gRNAs were used to excise the relevant genetic regions for downstream analysis. Results: Homozygous SLC47A1/MATE1 KO cells have an absence of the MATE1 protein (all P≤0.005). Functional downstream experiments, comparing clonal cell populations of SLC47A1/MATE1 KO, rs22289669 KO and rs8065082 KO with unedited control cells, indicate potential co-regulation of SLC22A1 (OCT1), another known metformin transporter, which may influence intracellular metformin accumulation. Further, rs8065082 KO cells show significant reduction of ALDH3A2 gene expression (all P≤0.002), indicating a novel relationship between rs8065082, fatty acid oxidation and the hepatic response to metformin. Conclusion: CRISPR KO of intronic SNPs in SLC47A1 reveal cis-acting regulation of SLC22A1 and ALDH3A2 gene expression; suggesting novel genetic mechanisms by which the hepatic response to metformin is determined.
Publications
- (2017) Genome-wide DNA promoter methylation and transcriptome analysis in human adipose tissue unravels novel candidate genes for obesity. Molecular metabolism 6 (1) 86–100
Keller, Maria; Hopp, Lydia; Liu, Xuanshi; Wohland, Tobias; Rohde, Kerstin; Cancello, Raffaella; Klös, Matthias; Bacos, Karl; Kern, Matthias; Eichelmann, Fabian; Dietrich, Arne; Schön, Michael R.; Gärtner, Daniel; Lohmann, Tobias; Dreßler, Miriam; Stumvoll
(See online at https://doi.org/10.1016/j.molmet.2016.11.003) - (2017) IRS1 DNA promoter methylation and expression in human adipose tissue are related to fat distribution and metabolic traits. Scientific reports 7 (1) 12369
Rohde, Kerstin; Klös, Matthias; Hopp, Lydia; Liu, Xuanshi; Keller, Maria; Stumvoll, Michael; Dietrich, Arne; Schön, Michael R.; Gärtner, Daniel; Lohmann, Tobias; Dreßler, Miriam; Kovacs, Peter; Binder, Hans; Blüher, Matthias; Böttcher, Yvonne
(See online at https://doi.org/10.1038/s41598-017-12393-5) - (2018) DNA methylation of SSPN is linked to adipose tissue distribution and glucose metabolism. The FASEB Journal 32 (12) 6898-6910
Keller, Maria; Klös, Matthias; Rohde, Kerstin; Krüger, Jacqueline; Kurze, Tabea; Dietrich, Arne; Schön, Michael R.; Gärtner, Daniel; Lohmann, Tobias; Dreßler, Miriam; Stumvoll, Michael; Blüher, Matthias; Kovacs, Peter; Böttcher, Yvonne
(See online at https://doi.org/10.1096/fj.201800528R) - (2019) The combined effects of FADS gene variation and dietary fats in obesity-related traits in a population from the far north of Sweden: the GLACIER Study. International journal of obesity (2005) 43 (4) 808–820
Chen, Yan; Estampador, Angela C.; Keller, Maria; Poveda, Alaitz; Dalla-Riva, Jonathan; Johansson, Ingegerd; Renström, Frida; Kurbasic, Azra; Franks, Paul W.; Varga, Tibor V.
(See online at https://doi.org/10.1038/s41366-018-0112-3)
