ABSTRACT A recent U.S. survey indicated that 40% of US adults are obese, and a worldwide survey of 195 countries showed that 2.2 billion people globally are obese or overweight. It would be vital to understand the genetic mechanisms and gene-environment interactions that underlie obesity-related phenotypes to improve treatment options and drug development to combat this global obesity epidemic. This emphasizes the need to discover additional DNA variants contributing to obesity. The goal of this project is to improve the understanding of genetic regulatory mechanisms of gene expression in obesity-related pathways in human adipocytes and adipose tissue. In Aim 1, we will integrate promoter Capture Hi-C (pCHi-C) data produced in primary Human White Adipocytes (HWAs), which elucidates the physical interaction between enhancers and promoters, with genotype, obesogenic phenotypes, and adipose tissue transcriptomic data from the Finnish METabolic Syndrome in Men (METSIM) cohort to directly follow up the looping cis expression quantitative trait loci (eQTLs) and BMI-correlated genes identified in my first paper (Pan et al. Nature Communications, 2018 in press). Our preliminary data show that looping cis-eQTLs in DNAse I Hypersensitivity sites (DHSs) significantly contribute to the variation in local gene expression. In Aim 1, we will refine this analysis of cis-eQTLs in DHSs using Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) in primary HWAs to identify the open chromatin regions in HWAs, a currently publically unavailable dataset. We will incorporate ATAC-seq data to identify the variants in regions of open chromatin within the chromosomal interactions where proteins, such as transcription factors (TFs), may bind. We will also use our previously identified list of 38 non-GWAS BMI-correlated genes as our obesity candidates, potentially reacting to obesogenic cellular environment via differential TF binding. Using Systematic high-resolution activation and repression profiling with the reporter- tiling Massively Parallel Reporter Assay (SHARPR-MPRA), we will confirm the enhancer potential of the looping cis-eQTL for the 38 BMI-correlated genes and search for their networks related to obesogenic phenotypes in METSIM using Weighted Gene Coexpression Network Analysis (WGCNA). In Aim 2, we propose to identify longer range trans-eQTLs mediated by looping cis-eQTLs and cis-genes. Using the TFs identified in obesogenic networks from WGCNA as potential cis-genes, we aim to discover their cis-eQTLs and then examine if those cis-eQTLs are also trans-eQTLs, confirming thus the direction of effect in a mediation analysis. The gene regulatory architecture that we identify and their obesogenic networks should help further the understanding of the genetic mechanisms that underlie adipose and adipocyte biology and provide potential clinically actionable targets for obesity. Aims 1 and 2 align with the mission of the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) to decrease the prevalence of obesity and cardiometabolic disorders through the understanding of the underlying molecular mechanisms.