Candidate Genes
The discovery of genetic mechanisms for resistance to obesity and diabetes may illuminate new therapeutic strategies for the treatment of this global health challenge. In the current study the polygenic lean mouse model, L- line, which has been selected for low adiposity over 60 generations, was used to identify mitochondrial thiosulfate sulfurtransferase (Tst; also known as rhodanese) as a candidate obesity-resistance gene with selectively increased expression in adipocytes. To prioritize positional candidate lean genes from within major adiposity QTLs (Fob3, Fob3, Fob3b1 and Fob3b2) mRNAs whose levels were increased specifically in the adipose tissue WAT depots (3 distinct white adipose tissues depots, subcutaneous, epididymal and mesenteric) of lean mice were selected. The nuclear-encoded Tst gene, which is positioned on chromosome 15 (78,399,556- 78,405,859) within the F-line obesity QTL Fob3b2, fulfilled all of the inclusion criteria. Tst mRNA was ~7-fold higher across the three WAT depots in lean versus fat mice [F-line].In Fob3b2-M2 heterozygotes, expression of the Tst allele originating from lean mice was higher than expression of the allele originating from fat mice (Supplementary Fig. 1f,g). Given that the Fob3b2-M2 congenic line carries a ~2.8-Mbp lean-line segment around Tst, the allele-dosage studies implicate a cis-mediated mechanism underlying increased levels of Tst mRNA in lean mice.[M2 congenic developed from congenic M line in J:158707 carrying the Fob3b2 QTL between 75.25-82.93 Mb].Elevated adipose Tst expression correlated with indices of metabolic health across diverse mouse strains. Transgenic overexpression of Tst in adipocytes protected mice from diet-induced obesity and insulin-resistant diabetes. Tst-deficient mice showed markedly exacerbated diabetes, whereas pharmacological activation of TST ameliorated diabetes in mice. Mechanistically, TST selectively augmented mitochondrial function combined with degradation of reactive oxygen species and sulfide. In humans, TST mRNA expression in adipose tissue correlated positively with insulin sensitivity in adipose tissue and negatively with fat mass. Thus, the genetic identification of Tst as a beneficial regulator of adipocyte mitochondrial function may have therapeutic significance for individuals with type 2 diabetes.
The current study developed a map of regulatory elements for the Tst locus in mice and identified candidate genetic variants with increased causal likelihood. The map provided a basis for experimental validation and functional analyses of this novel candidate leanness and anti-diabetic gene. The focus of the current study was on the nuclear-encoded mitochondrial thiosulfate sulfur-transferase gene (Tst, also known by synonym Rhodanese) that the authors recently identified in a positional cloning experiment as a causal gene for the Fob3b2 QTL phenotypic effect [22, J:236030]. The genetic variants responsible for upregulation of Tst expression in the Lean mice had not yet been identified.A main objective of the study was to integrate various genomic annotations in the Tst locus to construct a map of regulatory elements of the gene and to identify and prioritise the causal genetic variants between the Fat (F-line) and Lean (L-line) lines.First, to uncover polymorphisms that might be causal for the difference in expression of the Tst gene, a classical high resolution Sanger-sequencing of the entire Tst locus in Fat and Lean lines was undertaken. Bioinformatics analysis was performed on a 7400 bp-long segment of the mouse Tst gene, including the ~ 0.5 kb upstream and downstream regions.SNPs were categorized into variation consequence types - upstream gene variant, synonymous variant, intron variant, missense variant, nonsense variant, 5 or 3 untranslated region (UTR) variant and downstream gene variant.The identified genetic variants were then evaluated and prioritised using a regulatory element map of the Tst locus that integrated broad functional information from conserved polymorphisms in other strains, association studies, transcription factor binding site motifs, chromatin modification motifs and miRNA binding sites. In the case of Tst gene the comparative analysis of various regulatory features provided strong evidence of three regulatory genome segments located in the promotor, an intron, and the 3' UTR of the locus. The study concluded that SNPs rs251994838 and rs31534689 represented the highest priority candidate genetic variants for further experimental functional validation of their causality on the phneotype.The approach, to build a detailed integrative regulatory element map can be applied for evaluating and prioritising polymorphisms within candidate regions in any trait or species of interest. Reducing and prioritising the number of potential causal polymorphisms is essential for efficient planning of further experiments to prove or support causality of candidate genetic variants.
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