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Linking the FTO obesity rs1421085 variant circuitry to cellular, metabolic, and organismal phenotypes in vivo.

ABSTRACT: Variants in FTO have the strongest association with obesity; however, it is still unclear how those noncoding variants mechanistically affect whole-body physiology. We engineered a deletion of the rs1421085 conserved cis-regulatory module (CRM) in mice and confirmed in vivo that the CRM modulates Irx3 and Irx5 gene expression and mitochondrial function in adipocytes. The CRM affects molecular and cellular phenotypes in an adipose depot-dependent manner and affects organismal phenotypes that are relevant for obesity, including decreased high-fat diet-induced weight gain, decreased whole-body fat mass, and decreased skin fat thickness. Last, we connected the CRM to a genetically determined effect on steroid patterns in males that was dependent on nutritional challenge and conserved across mice and humans. Together, our data establish cross-species conservation of the rs1421085 regulatory circuitry at the molecular, cellular, metabolic, and organismal level, revealing previously unknown contextual dependence of the variant's action.

SUBMITTER: Laber S

PROVIDER: S-EPMC8294759 | biostudies-literature | 2021 Jul

REPOSITORIES: biostudies-literature

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Linking the FTO obesity rs1421085 variant circuitry to cellular, metabolic, and organismal phenotypes in vivo.

Laber Samantha S Forcisi Sara S Bentley Liz L Petzold Julia J Moritz Franco F Smirnov Kirill S KS Al Sadat Loubna L Williamson Iain I Strobel Sophie S Agnew Thomas T Sengupta Shahana S Nicol Tom T Grallert Harald H Heier Margit M Honecker Julius J Mianne Joffrey J Teboul Lydia L Dumbell Rebecca R Long Helen H Simon Michelle M Lindgren Cecilia C Bickmore Wendy A WA Hauner Hans H Schmitt-Kopplin Philippe P Claussnitzer Melina M Cox Roger D RD

Science advances 20210721 30

Variants in FTO have the strongest association with obesity; however, it is still unclear how those noncoding variants mechanistically affect whole-body physiology. We engineered a deletion of the rs1421085 conserved cis-regulatory module (CRM) in mice and confirmed in vivo that the CRM modulates Irx3 and Irx5 gene expression and mitochondrial function in adipocytes. The CRM affects molecular and cellular phenotypes in an adipose depot-dependent manner and affects organismal phenot ...[more]

PMID: 34290091

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Project description:BackgroundGenomewide association studies can be used to identify disease-relevant genomic regions, but interpretation of the data is challenging. The FTO region harbors the strongest genetic association with obesity, yet the mechanistic basis of this association remains elusive.MethodsWe examined epigenomic data, allelic activity, motif conservation, regulator expression, and gene coexpression patterns, with the aim of dissecting the regulatory circuitry and mechanistic basis of the association between the FTO region and obesity. We validated our predictions with the use of directed perturbations in samples from patients and from mice and with endogenous CRISPR-Cas9 genome editing in samples from patients.ResultsOur data indicate that the FTO allele associated with obesity represses mitochondrial thermogenesis in adipocyte precursor cells in a tissue-autonomous manner. The rs1421085 T-to-C single-nucleotide variant disrupts a conserved motif for the ARID5B repressor, which leads to derepression of a potent preadipocyte enhancer and a doubling of IRX3 and IRX5 expression during early adipocyte differentiation. This results in a cell-autonomous developmental shift from energy-dissipating beige (brite) adipocytes to energy-storing white adipocytes, with a reduction in mitochondrial thermogenesis by a factor of 5, as well as an increase in lipid storage. Inhibition of Irx3 in adipose tissue in mice reduced body weight and increased energy dissipation without a change in physical activity or appetite. Knockdown of IRX3 or IRX5 in primary adipocytes from participants with the risk allele restored thermogenesis, increasing it by a factor of 7, and overexpression of these genes had the opposite effect in adipocytes from nonrisk-allele carriers. Repair of the ARID5B motif by CRISPR-Cas9 editing of rs1421085 in primary adipocytes from a patient with the risk allele restored IRX3 and IRX5 repression, activated browning expression programs, and restored thermogenesis, increasing it by a factor of 7.ConclusionsOur results point to a pathway for adipocyte thermogenesis regulation involving ARID5B, rs1421085, IRX3, and IRX5, which, when manipulated, had pronounced pro-obesity and anti-obesity effects. (Funded by the German Research Center for Environmental Health and others.).

Project description:Introduction: White adipocytes store lipids, have a large lipid droplet and few mitochondria. Brown and beige adipocytes, which produce heat, are characterized by high expression of uncoupling protein (UCP) 1, multilocular lipid droplets, and large amounts of mitochondria. The rs1421085 T-to-C single-nucleotide polymorphism (SNP) of the human FTO gene interrupts a conserved motif for ARID5B repressor, resulting in adipocyte type shift from beige to white. Methods: We obtained abdominal subcutaneous adipose tissue from donors carrying FTO rs1421085 TT (risk-free) or CC (obesity-risk) genotypes, isolated and differentiated their preadipocytes into beige adipocytes (driven by the PPARγ agonist rosiglitazone for 14 days), and activated them with dibutyryl-cAMP for 4 hours. Then, either the same culture conditions were applied for additional 14 days (active beige adipocytes) or it was replaced by a white differentiation medium (inactive beige adipocytes). White adipocytes were differentiated by their medium for 28 days. Results and Discussion: RNA-sequencing was performed to investigate the gene expression pattern of adipocytes carrying different FTO alleles and found that active beige adipocytes had higher brown adipocyte content and browning capacity compared to white or inactive beige ones when the cells were obtained from risk-free TT but not from obesity-risk CC genotype carriers. Active beige adipocytes carrying FTO CC had lower thermogenic gene (e.g., UCP1, PM20D1, CIDEA) expression and thermogenesis measured by proton leak respiration as compared to TT carriers. In addition, active beige adipocytes with CC alleles exerted lower expression of ASC-1 neutral amino acid transporter (encoded by SLC7A10) and less consumption of Ala, Ser, Cys, and Gly as compared to risk-free carriers. We did not observe any influence of the FTO rs1421085 SNP on white and inactive beige adipocytes highlighting its exclusive and critical effect when adipocytes were activated for thermogenesis.

Dataset Information

Linking the FTO obesity rs1421085 variant circuitry to cellular, metabolic, and organismal phenotypes in vivo.

Publications

Linking the <i>FTO</i> obesity rs1421085 variant circuitry to cellular, metabolic, and organismal phenotypes in vivo.

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