NGFN-PLUS
Syntenic regions for atherosclerosis in mice and humans
| Coordinator: | Ronny Baber | |
| Institution: | Universität Leipzig | |
| Homepage: | http://www.uniklinikum-leipzig.de |
Comparative genetic analyses are based on the hypothesis that disease-susceptibility genes are shared between species. One major focus of our work was related to comparative and functional studies of a genetic locus of atherosclerosis predisposition and sterol metabolism at ABCG5/8 in mice and men. We used next generation sequencing (454-GS FLX) for complete resequencing of ABCG5 und ABCG8 in mice and men. For the resequencing of mice we used DNA from 3 different inbred strains (C57BL/6, BALB/c and FVB) where the locus was previously identified. We found a total of 81 variants (63 in abcg5 and 18 in abcg8) and constructed a total of 10 DNA-fragments carrying those SNPs. These DNA-fragments are now being checked for their impact on the expression of abcg5/8 in reporter gene assays.
Complete sequencing of ABCG5/8 in humans was performed with DNA of probands from the LIFE/Leipzig Herzstudie and one patient with sitosterolemia. Studies of two basic ABCG8 haplotypes, which influenced sterol levels and predisposition to atherosclerosis resulted in identification of 14 additional variants, which were in strong linkage disequilibrium with the initially identified variants. For these SNPs reporter gene vectors have been constructed for expression analyses in a cell model. The causal mutation of the sitosterolemia patient could be identified as W361X leading to an early stop codon in exon 7 of ABCG8. Additionally 9 yet unknown nonsynonymous mutations have been found in our experimental setup. The next step would be a screening of the LIFE/Leipzig Herzstudie for these mutations.
The genome-wide association study in cooperation with members of this consortium resulted in a second variant located in the blood-group gene ABO. In the past reporting-period mice with the human blood group A und 0 have been generated. The transgenic over-expression of ABO was tissue specific for the 7 different mouse strains. To investigate effects of ABO on atherosclerosis, we started backcrossing these strains onto the atherosclerosis susceptible LDLR-/- background. Finally, comparative genetic studies of loci identified by eQTL mapping in human peripheral blood cells from patients with different degree of coronary disease, combining GWE and GWA in >2000 subjects are well advanced.
Further Coordinators:
Complete sequencing of ABCG5/8 in humans was performed with DNA of probands from the LIFE/Leipzig Herzstudie and one patient with sitosterolemia. Studies of two basic ABCG8 haplotypes, which influenced sterol levels and predisposition to atherosclerosis resulted in identification of 14 additional variants, which were in strong linkage disequilibrium with the initially identified variants. For these SNPs reporter gene vectors have been constructed for expression analyses in a cell model. The causal mutation of the sitosterolemia patient could be identified as W361X leading to an early stop codon in exon 7 of ABCG8. Additionally 9 yet unknown nonsynonymous mutations have been found in our experimental setup. The next step would be a screening of the LIFE/Leipzig Herzstudie for these mutations.
The genome-wide association study in cooperation with members of this consortium resulted in a second variant located in the blood-group gene ABO. In the past reporting-period mice with the human blood group A und 0 have been generated. The transgenic over-expression of ABO was tissue specific for the 7 different mouse strains. To investigate effects of ABO on atherosclerosis, we started backcrossing these strains onto the atherosclerosis susceptible LDLR-/- background. Finally, comparative genetic studies of loci identified by eQTL mapping in human peripheral blood cells from patients with different degree of coronary disease, combining GWE and GWA in >2000 subjects are well advanced.
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