Summary Genome sequencing projects are discovering millions of genetic variants in humans, and interpretation of their functional effects is essential for understanding the genetic basis of variation in human traits. Here we report sequencing and deep analysis of mRNA and miRNA from lymphoblastoid cell lines of 462 individuals from the 1000 Genomes Project – the first uniformly processed RNA-seq data from multiple human populations with high-quality genome sequences. We discovered extremely widespread genetic variation affecting regulation of the majority of genes, with transcript structure and expression level variation being equally common but genetically largely independent. Our characterization of causal regulatory variation sheds light on cellular mechanisms of regulatory and loss-of-function variation, and allowed us to infer putative causal variants for dozens of disease-associated loci. Altogether, this study provides a deep understanding of the cellular mechanisms of transcriptome variation and of the landscape of functional variants in the human genome.

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The genetic architecture of common traits, including the number, frequency, and effect sizes of inherited variants that contribute to individual risk, has been long debated. Genome-wide association studies have identified scores of common variants associated with type 2 diabetes, but in aggregate, these explain only a fraction of heritability. To test the hypothesis that lower-frequency variants explain much of the remainder, the GoT2D and T2D-GENES consortia performed whole genome sequencing in 2,657 Europeans with and without diabetes, and exome sequencing in a total of 12,940 subjects from five ancestral groups. To increase statistical power, we expanded sample size via genotyping and imputation in a further 111,548 subjects. Variants associated with type 2 diabetes after sequencing were overwhelmingly common and most fell within regions previously identified by genome-wide association studies. Comprehensive enumeration of sequence variation is necessary to identify functional alleles that provide important clues to disease pathophysiology, but large-scale sequencing does not support a major role for lower-frequency variants in predisposition to type 2 diabetes.

Background Delayed afterdepolarizations (DADs) carried by Na+-Ca2+-exchange current (INCX) in response to sarcoplasmic reticulum (SR) Ca2+-leak can promote atrial fibrillation (AF). The mechanisms leading to DADs in AF-patients have not been defined. Methods and Results Protein levels (Western-blot), membrane-currents and action-potentials (patch-clamp), and [Ca2+]i (Fluo-3) were measured in right-atrial samples from 77 sinus-rhythm (Ctl) and 69 chronic-AF (cAF) patients. Diastolic [Ca2+]i and SR-Ca2+-content (integrated INCX during caffeine-induced-Ca2+-transient [cCaT]) were unchanged, whereas diastolic SR Ca2+-leak, estimated by blocking RyR2 with tetracaine, was ~50% higher in cAF vs. Ctl. Single-channel recordings from atrial RyR2 reconstituted into lipid-bilayers revealed enhanced open-probability in cAF-samples, providing a molecular basis for increased SR Ca2+-leak. Calmodulin-expression (+60%), CaMKII-autophosphorylation at Thr287 (+40%) and RyR2-phosphorylation at Ser2808 (PKA/CaMKII-site, +236%) and Ser2814 (CaMKII-site, +77%) were increased in cAF. The selective CaMKII-blocker KN-93 decreased SR Ca2+-leak, the frequency of spontaneous Ca2+-release events and RyR2 open-probability in cAF, whereas PKA-inhibition with H-89 was ineffective. Knock-in mice with constitutively-phosphorylated RyR2 at Ser2814 showed a higher incidence of Ca2+-sparks and increased susceptibility to pacing-induced AF vs. controls. The relationship between [Ca2+]i and INCX-density revealed INCX-upregulation in cAF. Spontaneous Ca2+-release events accompanied by inward INCX-currents and DADs/triggered-activity occurred more often and the sensitivity of resting membrane voltage to elevated [Ca2+]i (diastolic [Ca2+]i–voltage coupling gain) was higher in cAF vs. Ctl. Conclusions Enhanced SR Ca2+-leak through CaMKII-hyperphosphorylated RyR2, in combination with larger INCX for a given SR Ca2+-release and increased diastolic [Ca2+]i–voltage coupling gain, cause AF-promoting atrial DADs/triggered-activity in cAF patients.