Integration of human adipocyte chromosomal interactions with adipose gene expression prioritizes obesity-related genes from GWAS

Abstract

Increased adiposity is a hallmark of obesity and overweight, which affect 2.2 billion people world-wide. Understanding the genetic and molecular mechanisms that underlie obesity-related phenotypes can help to improve treatment options and drug development. Here we perform promoter Capture Hi-C in human adipocytes to investigate interactions between gene promoters and distal elements as a transcription-regulating mechanism contributing to these phenotypes. We find that promoter-interacting elements in human adipocytes are enriched for adipose-related transcription factor motifs, such as PPARG and CEBPB, and contribute to heritability of cis-regulated gene expression. We further intersect these data with published genome-wide association studies for BMI and BMI-related metabolic traits to identify the genes that are under genetic cis regulation in human adipocytes via chromosomal interactions. This integrative genomics approach identifies four cis-eQTL-eGene relationships associated with BMI or obesity-related traits, including rs4776984 and MAP2K5, which we further confirm by EMSA, and highlights 38 additional candidate genes.

Fig. 1

Open chromatin sites (DHSs) within adipocyte promoter CHi-C chromosomal interactions show significant enrichment in cis expression. Enrichments in cis expression with error bars for different categories using LD score regression analysis (see Methods). For the horizontal axis labels, the value in parentheses shows the percentage of SNPs contained within the respective annotation category that contributed to the enrichment calculation. For the significance threshold after Bonferroni correction above each bar, * indicates a p-value < 0.05; **, a p-value < 0.001; and ***, a p-value < 0.0001, respectively. The p-values were estimated based on Z scores calculated from the normal distribution. Error bars represent jackknife standard errors around the estimates of enrichment

Fig. 2

Overview of the study design targeted to identify new genes for obesity and related metabolic traits. A schematic illustrating the integration of multi-omics data utilized in this study to elucidate genetics of obesity-related traits.

Fig. 3

Promoter Capture Hi–C enables refinement of the BMI GWAS locus that colocalizes with cis-eQTLs interacting with the target gene promoter of MAP2K5. Genomic landscape of the BMI locus, MAP2K5 (panels a, b), modified from the WashU Genome Browser to show the histone mark calls from ChIP-seq data; gene transcripts; promoter and eQTL HindIII fragments that interact in primary human white adipocytes (HWA); and GWAS SNPs (A, the rs number indicated in the magnified box) or their LD proxies (B, r² > 0.8) located in the interacting HindIII fragment. The vertical yellow band highlights the cis-eQTL variant (the rs number is indicated in the magnified box). a Genomic landscape containing MAP2K5 and the interacting cis-eQTL variant and corresponding BMI GWAS SNP. b Magnification of the boxed region in (a)

Fig. 4

Predicted TF motifs and electrophoretic mobility shift assay (EMSA) at the rs4776984 site indicate allele-specific binding. a Predicted TF motifs for CTCF and p300, as well as the hg19 reference genome sequence. b Biotinylated (labeled probe) 31-bp oligonucleotide complexes with ±15 bp flanking the reference or alternate allele for variant rs4776984 were incubated with nuclear protein extracted from primary HWA and resolved on a 6% polyacrylamide gel. Competitor assays were performed by incubating the reaction with ×100 excess of unlabeled (no biotin) oligonucleotide complexes with identical sequence. Arrow denotes specific binding of HWA nuclear protein to reference (left) and alternate (right) allele