Abstract
Genome-wide association studies (GWAS) have linked single nucleotide polymorphisms (SNPs) at >250 loci in the human genome to type 2 diabetes (T2D) risk. For each locus, identifying the functional variant(s) among multiple SNPs in high linkage disequilibrium is critical to understand molecular mechanisms underlying T2D genetic risk. Using massively parallel reporter assays (MPRA), we test the cis-regulatory effects of SNPs associated with T2D and altered in vivo islet chromatin accessibility in MIN6 β cells under steady state and pathophysiologic endoplasmic reticulum (ER) stress conditions. We identify 1,982/6,621 (29.9%) SNP-containing elements that activate transcription in MIN6 and 879 SNP alleles that modulate MPRA activity. Multiple T2D-associated SNPs alter the activity of short interspersed nuclear element (SINE)-containing elements that are strongly induced by ER stress. We identify 220 functional variants at 104 T2D association signals, narrowing 54 signals to a single candidate SNP. Together, this study identifies elements driving β cell steady state and ER stress-responsive transcriptional activation, nominates causal T2D SNPs, and uncovers potential roles for repetitive elements in β cell transcriptional stress response and T2D genetics.
Highlights
Genome-wide association studies (GWAS) have linked single nucleotide polymorphisms (SNPs) at >250 loci in the human genome to type 2 diabetes (T2D) risk
The massively parallel reporter assays (MPRA) library consisted of twohundred base pair sequences from the human genome containing each allele for SNPs including: (i) 2512 index or linked (EUR r2 ≥ 0.8) SNPs/indels from 259 T2D and related quantitative trait association signals in the NHGRI/EBI GWAS Catalogue[28] (“T2D SNPs”); (ii) 1910 SNPs significantly associated with changes in human islet chromatin accessibility (“caQTL SNPs”)[7]; and (iii) 2214 SNPs that overlapped human islet ATAC-seq peaks, but were not significantly associated with changes in human islet chromatin accessibility (“non-caQTL SNPs”)[7] (Methods; Fig. 1a)
We found that T2D functional SNPs nominated by MPRA were more enriched for islet eQTLs than T2D SNPs without MPRA activity (Fig. 4a)
Summary
Genome-wide association studies (GWAS) have linked single nucleotide polymorphisms (SNPs) at >250 loci in the human genome to type 2 diabetes (T2D) risk. Studies over the past several years clearly implicate altered islet cis-regulatory element (CRE) activity in T2D genetic risk and progression, they have co-localized only ~1/4 of T2D-associated loci to altered chromatin accessibility and/or gene expression levels in islets[5,7,8,9,10]. This may be partially because previous studies measured the effect of genetic variants on chromatin accessibility (caQTLs) and gene expression levels (eQTLs) in islets under steadystate conditions[7,11], missing the role of genetic variants whose functions emerge only under certain cellular conditions. Non-coding T2D risk alleles may modulate the transcription of genes and pathways that alter ER stress responses and the UPR
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