Genome-wide association studies (GWAS) have long relied on proposed statistical significance thresholds to be able to differentiate true positives from false positives. Although the genome-wide ...significance P-value threshold of 5 × 10(-8) has become a standard for common-variant GWAS, it has not been updated to cope with the lower allele frequency spectrum used in many recent array-based GWAS studies and sequencing studies. Using a whole-genome- and -exome-sequencing data set of 2875 individuals of European ancestry from the Genetics of Type 2 Diabetes (GoT2D) project and a whole-exome-sequencing data set of 13 000 individuals from five ancestries from the GoT2D and T2D-GENES (Type 2 Diabetes Genetic Exploration by Next-generation sequencing in multi-Ethnic Samples) projects, we describe guidelines for genome- and exome-wide association P-value thresholds needed to correct for multiple testing, explaining the impact of linkage disequilibrium thresholds for distinguishing independent variants, minor allele frequency and ancestry characteristics. We emphasize the advantage of studying recent genetic isolate populations when performing rare and low-frequency genetic association analyses, as the multiple testing burden is diminished due to higher genetic homogeneity.
Depletion of loss-of-function (LoF) mutations may provide a rank of genic functional intolerance and consequently susceptibility to disease.
Here we have studied LoF mutations in 60 706 unrelated ...individuals and show that the most intolerant quartile of ranked genes is enriched in rare and early onset diseases and explains 87% of de novo haploinsufficient OMIM mutations, 17% more than any other gene scoring tool. We detected particular enrichment in expression of the depleted LoF genes in brain (odds ratio = 1.5; P -value = 4.2e-07). By searching for de novo haploinsufficient mutations putatively associated with neurodevelopmental disorders in four recent studies, we were able to explain 81% of them. Taken together, this study provides a novel gene intolerance ranking system, called LoFtool, which may help in ranking genes of interest based on their LoF intolerance and tissue expression.
The LoFtool gene scores are available in the Supplementary data .
joaofadista@gmail.com.
Supplementary data are available at Bioinformatics online.
Significance We provide a comprehensive catalog of novel genetic variants influencing gene expression and metabolic phenotypes in human pancreatic islets. The data also show that the path from ...genetic variation (SNP) to gene expression is more complex than hitherto often assumed, and that we need to consider that genetic variation can also influence function of a gene by influencing exon usage or splice isoforms (sQTL), allelic imbalance, RNA editing, and expression of noncoding RNAs, which in turn can influence expression of target genes.
Genetic variation can modulate gene expression, and thereby phenotypic variation and susceptibility to complex diseases such as type 2 diabetes (T2D). Here we harnessed the potential of DNA and RNA sequencing in human pancreatic islets from 89 deceased donors to identify genes of potential importance in the pathogenesis of T2D. We present a catalog of genetic variants regulating gene expression (eQTL) and exon use (sQTL), including many long noncoding RNAs, which are enriched in known T2D-associated loci. Of 35 eQTL genes, whose expression differed between normoglycemic and hyperglycemic individuals, siRNA of tetraspanin 33 (TSPAN33), 5′-nucleotidase, ecto (NT5E), transmembrane emp24 protein transport domain containing 6 (TMED6), and p21 protein activated kinase 7 (PAK7) in INS1 cells resulted in reduced glucose-stimulated insulin secretion. In addition, we provide a genome-wide catalog of allelic expression imbalance, which is also enriched in known T2D-associated loci. Notably, allelic imbalance in paternally expressed gene 3 (PEG3) was associated with its promoter methylation and T2D status. Finally, RNA editing events were less common in islets than previously suggested in other tissues. Taken together, this study provides new insights into the complexity of gene regulation in human pancreatic islets and better understanding of how genetic variation can influence glucose metabolism.
Copy number variations (CNVs), which represent a significant source of genetic diversity in mammals, have been shown to be associated with phenotypes of clinical relevance and to be causative of ...disease. Notwithstanding, little is known about the extent to which CNV contributes to genetic variation in cattle.
We designed and used a set of NimbleGen CGH arrays that tile across the assayable portion of the cattle genome with approximately 6.3 million probes, at a median probe spacing of 301 bp. This study reports the highest resolution map of copy number variation in the cattle genome, with 304 CNV regions (CNVRs) being identified among the genomes of 20 bovine samples from 4 dairy and beef breeds. The CNVRs identified covered 0.68% (22 Mb) of the genome, and ranged in size from 1.7 to 2,031 kb (median size 16.7 kb). About 20% of the CNVs co-localized with segmental duplications, while 30% encompass genes, of which the majority is involved in environmental response. About 10% of the human orthologous of these genes are associated with human disease susceptibility and, hence, may have important phenotypic consequences.
Together, this analysis provides a useful resource for assessment of the impact of CNVs regarding variation in bovine health and production traits.
Type 2 diabetes (T2D) is a global pandemic. Genome-wide association studies (GWASs) have identified >100 genetic variants associated with the disease, including a common variant in the melatonin ...receptor 1 b gene (MTNR1B). Here, we demonstrate increased MTNR1B expression in human islets from risk G-allele carriers, which likely leads to a reduction in insulin release, increasing T2D risk. Accordingly, in insulin-secreting cells, melatonin reduced cAMP levels, and MTNR1B overexpression exaggerated the inhibition of insulin release exerted by melatonin. Conversely, mice with a disruption of the receptor secreted more insulin. Melatonin treatment in a human recall-by-genotype study reduced insulin secretion and raised glucose levels more extensively in risk G-allele carriers. Thus, our data support a model where enhanced melatonin signaling in islets reduces insulin secretion, leading to hyperglycemia and greater future risk of T2D. The findings also imply that melatonin physiologically serves to inhibit nocturnal insulin release.
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•rs10830963 is an eQTL in human islets conferring increased MTNR1B mRNA expression•Melatonin inhibits cAMP rises in mouse islets and clonal insulin-secreting cells•Melatonin blocks insulin release in mouse islets and clonal insulin-secreting cells•Melatonin’s inhibition of insulin release is stronger in risk allele carriers
Tuomi et al. show that a common (about 30%) human type 2 diabetes risk variant of the melatonin receptor 1B gene affects insulin release. A recall-by-genotype study demonstrated that melatonin treatment inhibits insulin secretion, with at-risk carriers exhibiting higher glucose levels. Melatonin might have a protective role in preventing nocturnal hypoglycemia.
Genome-wide association studies have revealed >60 loci associated with type 2 diabetes (T2D), but the underlying causal variants and functional mechanisms remain largely elusive. Although variants in ...TCF7L2 confer the strongest risk of T2D among common variants by presumed effects on islet function, the molecular mechanisms are not yet well understood. Using RNA-sequencing, we have identified a TCF7L2-regulated transcriptional network responsible for its effect on insulin secretion in rodent and human pancreatic islets. ISL1 is a primary target of TCF7L2 and regulates proinsulin production and processing via MAFA, PDX1, NKX6.1, PCSK1, PCSK2 and SLC30A8, thereby providing evidence for a coordinated regulation of insulin production and processing. The risk T-allele of rs7903146 was associated with increased TCF7L2 expression, and decreased insulin content and secretion. Using gene expression profiles of 66 human pancreatic islets donors', we also show that the identified TCF7L2-ISL1 transcriptional network is regulated in a genotype-dependent manner. Taken together, these results demonstrate that not only synthesis of proinsulin is regulated by TCF7L2 but also processing and possibly clearance of proinsulin and insulin. These multiple targets in key pathways may explain why TCF7L2 has emerged as the gene showing one of the strongest associations with T2D.
Integration of genomic variation with phenotypic information is an effective approach for uncovering genotype-phenotype associations. This requires an accurate identification of the different types ...of variation in individual genomes.
We report the integration of the whole genome sequence of a single Holstein Friesian bull with data from single nucleotide polymorphism (SNP) and comparative genomic hybridization (CGH) array technologies to determine a comprehensive spectrum of genomic variation. The performance of resequencing SNP detection was assessed by combining SNPs that were identified to be either in identity by descent (IBD) or in copy number variation (CNV) with results from SNP array genotyping. Coding insertions and deletions (indels) were found to be enriched for size in multiples of 3 and were located near the N- and C-termini of proteins. For larger indels, a combination of split-read and read-pair approaches proved to be complementary in finding different signatures. CNVs were identified on the basis of the depth of sequenced reads, and by using SNP and CGH arrays.
Our results provide high resolution mapping of diverse classes of genomic variation in an individual bovine genome and demonstrate that structural variation surpasses sequence variation as the main component of genomic variability. Better accuracy of SNP detection was achieved with little loss of sensitivity when algorithms that implemented mapping quality were used. IBD regions were found to be instrumental for calculating resequencing SNP accuracy, while SNP detection within CNVs tended to be less reliable. CNV discovery was affected dramatically by platform resolution and coverage biases. The combined data for this study showed that at a moderate level of sequencing coverage, an ensemble of platforms and tools can be applied together to maximize the accurate detection of sequence and structural variants.
Idiopathic pulmonary fibrosis (IPF) is a complex lung disease, characterized by progressive lung scarring. Severe COVID-19 is associated with substantial pneumonitis and has a number of shared major ...risk factors with IPF. This study aimed to determine the genetic correlation between IPF and severe COVID-19 and assess a potential causal role of genetically increased risk of IPF on COVID-19 severity.
The genetic correlation between IPF and COVID-19 severity was estimated with linkage disequilibrium (LD) score regression. We performed a Mendelian randomization (MR) study for IPF causality in COVID-19. Genetic variants associated with IPF susceptibility (P<5 × 10−8) in previous genome-wide association studies (GWAS) were used as instrumental variables (IVs). Effect estimates of those IVs on COVID-19 severity were gathered from the GWAS meta-analysis by the COVID-19 Host Genetics Initiative (4,336 cases & 623,902 controls).
We detected a positive genetic correlation of IPF with COVID-19 severity (rg=0·31 95% CI 0·04–0·57, P = 0·023). The MR estimates for severe COVID-19 did not reveal any genetic association (OR 1·05, 95% CI 0·92–1·20, P = 0·43). However, outlier analysis revealed that the IPF risk allele rs35705950 at MUC5B had a different effect compared with the other variants. When rs35705950 was excluded, MR results provided evidence that genetically increased risk of IPF has a causal effect on COVID-19 severity (OR 1·21, 95% CI 1·06–1·38, P = 4·24 × 10−3). Furthermore, the IPF risk-allele at MUC5B showed an apparent protective effect against COVID-19 hospitalization only in older adults (OR 0·86, 95% CI 0·73–1·00, P = 2·99 × 10−2) .
The strongest genetic determinant of IPF, rs35705950 at MUC5B, seems to confer protection against COVID-19, whereas the combined effect of all other IPF risk loci seem to confer risk of COVID-19 severity. The observed effect of rs35705950 could either be due to protective effects of mucin over-production on the airways or a consequence of selection bias due to (1) a patient group that is heavily enriched for the rs35705950 T undertaking strict self-isolation and/or (2) due to survival bias of the rs35705950 non-IPF risk allele carriers. Due to the diverse impact of IPF causal variants on SARS-CoV-2 infection, with a possible selection bias as an explanation, further investigation is needed to address this apparent paradox between variance at MUC5B and other IPF genetic risk factors.
Novo Nordisk Foundation and Oak Foundation.
Genetics, epigenetics, and environment may together affect the susceptibility for type 2 diabetes (T2D). Our aim was to dissect molecular mechanisms underlying T2D using genome-wide expression and ...DNA methylation data in adipose tissue from monozygotic twin pairs discordant for T2D and independent case-control cohorts. In adipose tissue from diabetic twins, we found decreased expression of genes involved in oxidative phosphorylation; carbohydrate, amino acid, and lipid metabolism; and increased expression of genes involved in inflammation and glycan degradation. The most differentially expressed genes included ELOVL6, GYS2, FADS1, SPP1 (OPN), CCL18, and IL1RN. We replicated these results in adipose tissue from an independent case-control cohort. Several candidate genes for obesity and T2D (e.g., IRS1 and VEGFA) were differentially expressed in discordant twins. We found a heritable contribution to the genome-wide DNA methylation variability in twins. Differences in methylation between monozygotic twin pairs discordant for T2D were subsequently modest. However, 15,627 sites, representing 7,046 genes including PPARG, KCNQ1, TCF7L2, and IRS1, showed differential DNA methylation in adipose tissue from unrelated subjects with T2D compared with control subjects. A total of 1,410 of these sites also showed differential DNA methylation in the twins discordant for T2D. For the differentially methylated sites, the heritability estimate was 0.28. We also identified copy number variants (CNVs) in monozygotic twin pairs discordant for T2D. Taken together, subjects with T2D exhibit multiple transcriptional and epigenetic changes in adipose tissue relevant to the development of the disease.
Most signals detected by genome-wide association studies map to non-coding sequence and their tissue-specific effects influence transcriptional regulation. However, key tissues and cell-types ...required for functional inference are absent from large-scale resources. Here we explore the relationship between genetic variants influencing predisposition to type 2 diabetes (T2D) and related glycemic traits, and human pancreatic islet transcription using data from 420 donors. We find: (a) 7741 cis-eQTLs in islets with a replication rate across 44 GTEx tissues between 40% and 73%; (b) marked overlap between islet cis-eQTL signals and active regulatory sequences in islets, with reduced eQTL effect size observed in the stretch enhancers most strongly implicated in GWAS signal location; (c) enrichment of islet cis-eQTL signals with T2D risk variants identified in genome-wide association studies; and (d) colocalization between 47 islet cis-eQTLs and variants influencing T2D or glycemic traits, including DGKB and TCF7L2. Our findings illustrate the advantages of performing functional and regulatory studies in disease relevant tissues.
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