Altered Transcription Factor Binding Research Articles

Aberrant DNA methylation distorts developmental trajectories in atypical teratoid/rhabdoid tumors.

Atypical teratoid/rhabdoid tumors (AT/RTs) are pediatric brain tumors known for their aggressiveness and aberrant but still unresolved epigenetic regulation. To better understand their malignancy, we investigated how AT/RT-specific DNA hypermethylation was associated with gene expression and altered transcription factor binding and how it is linked to upstream regulation. Medulloblastomas, choroid plexus tumors, pluripotent stem cells, and fetal brain were used as references. A part of the genomic regions, which were hypermethylated in AT/RTs similarly as in pluripotent stem cells and demethylated in the fetal brain, were targeted by neural transcriptional regulators. AT/RT-unique DNA hypermethylation was associated with polycomb repressive complex 2 and linked to suppressed genes with a role in neural development and tumorigenesis. Activity of the several NEUROG/NEUROD pioneer factors, which are unable to bind to methylated DNA, was compromised via the suppressed expression or DNA hypermethylation of their target sites, which was also experimentally validated for NEUROD1 in medulloblastomas and AT/RT samples. These results highlight and characterize the role of DNA hypermethylation in AT/RT malignancy and halted neural cell differentiation.

Modeling methyl-sensitive transcription factor motifs with an expanded epigenetic alphabet

BackgroundTranscription factors bind DNA in specific sequence contexts. In addition to distinguishing one nucleobase from another, some transcription factors can distinguish between unmodified and modified bases. Current models of transcription factor binding tend not to take DNA modifications into account, while the recent few that do often have limitations. This makes a comprehensive and accurate profiling of transcription factor affinities difficult.ResultsHere, we develop methods to identify transcription factor binding sites in modified DNA. Our models expand the standard A/C/G/T DNA alphabet to include cytosine modifications. We develop Cytomod to create modified genomic sequences and we also enhance the MEME Suite, adding the capacity to handle custom alphabets. We adapt the well-established position weight matrix (PWM) model of transcription factor binding affinity to this expanded DNA alphabet. Using these methods, we identify modification-sensitive transcription factor binding motifs. We confirm established binding preferences, such as the preference of ZFP57 and C/EBPβ for methylated motifs and the preference of c-Myc for unmethylated E-box motifs.ConclusionsUsing known binding preferences to tune model parameters, we discover novel modified motifs for a wide array of transcription factors. Finally, we validate our binding preference predictions for OCT4 using cleavage under targets and release using nuclease (CUT&RUN) experiments across conventional, methylation-, and hydroxymethylation-enriched sequences. Our approach readily extends to other DNA modifications. As more genome-wide single-base resolution modification data becomes available, we expect that our method will yield insights into altered transcription factor binding affinities across many different modifications.

Type 1 diabetes risk genes mediate pancreatic beta cell survival in response to proinflammatory cytokines

We combined functional genomics and human genetics to investigate processes that affect type 1 diabetes (T1D) risk by mediating beta cell survival in response to proinflammatory cytokines. We mapped 38,931 cytokine-responsive candidate cis-regulatory elements (cCREs) in beta cells using ATAC-seq and snATAC-seq and linked them to target genes using co-accessibility and HiChIP. Using a genome-wide CRISPR screen in EndoC-βH1 cells, we identified 867 genes affecting cytokine-induced survival, and genes promoting survival and up-regulated in cytokines were enriched at T1D risk loci. Using SNP-SELEX, we identified 2,229 variants in cytokine-responsive cCREs altering transcription factor (TF) binding, and variants altering binding of TFs regulating stress, inflammation, and apoptosis were enriched for T1D risk. At the 16p13 locus, a fine-mapped T1D variant altering TF binding in a cytokine-induced cCRE interacted with SOCS1, which promoted survival in cytokine exposure. Our findings reveal processes and genes acting in beta cells during inflammation that modulate T1D risk.

PRKCE non-coding variants influence on transcription as well as translation of its gene

ABSTRACT Untranslated regions of the gene play a crucial role in gene expression regulation at mRNA and protein levels. Mutations at UTRs impact expression by altering transcription factor binding, transcriptional/translational efficacy, miRNA-mediated gene regulation, mRNA secondary structure, ribosomal translocation, and stability. PKCε, a serine/threonine kinase, is aberrantly expressed in numerous diseases such as cardiovascular disorders, neurological disorders, and cancers; its probable cause is unknown. Therefore, in the current study, the influence of PRKCE 5’-and 3'UTR variants was explored for their potential impact on its transcription and translation through several bioinformatics approaches. UTR variants data was obtained through different databases and initially evaluated for their regulatory function. Variants with regulatory function were then studied for their effect on PRKCE binding with transcription factors (TF) and miRNAs, as well as their impact on mRNA secondary structure. Study outcomes indicated the regulatory function of 73 5'UTR and 17 3'UTR variants out of 376. 5'UTR variants introduced AP1 binding sites and promoted the PRKCE transcription. Four 3'UTR variants introduced a circular secondary structure, increasing PRKCE translational efficacy. A region in 5'UTR position 45,651,564 to 45,651,644 was found where variants readily influenced the miRNA-PRKCE mRNA binding. The study further highlighted a PKCε-regulated feedback loop mechanism that induces the activity of TFs, promoting its gene transcription. The study provides foundations for experimentation to understand these variants’ role in diseases. These variants can also serve as the genetic markers for different diseases’ diagnoses after validation at the cell and population levels.

Characterization of sequence determinants of enhancer function using natural genetic variation.

Sequence variation in enhancers that control cell-type-specific gene transcription contributes significantly to phenotypic variation within human populations. However, it remains difficult to predict precisely the effect of any given sequence variant on enhancer function due to the complexity of DNA sequence motifs that determine transcription factor (TF) binding to enhancers in their native genomic context. Using F1-hybrid cells derived from crosses between distantly related inbred strains of mice, we identified thousands of enhancers with allele-specific TF binding and/or activity. We find that genetic variants located within the central region of enhancers are most likely to alter TF binding and enhancer activity. We observe that the AP-1 family of TFs (Fos/Jun) are frequently required for binding of TEAD TFs and for enhancer function. However, many sequence variants outside of core motifs for AP-1 and TEAD also impact enhancer function, including sequences flanking core TF motifs and AP-1 half sites. Taken together, these data represent one of the most comprehensive assessments of allele-specific TF binding and enhancer function to date and reveal how sequence changes at enhancers alter their function across evolutionary timescales.

Thymocyte regulatory variant alters transcription factor binding and protects from type 1 diabetes in infants

We recently mapped a genetic susceptibility locus on chromosome 6q22.33 for type 1 diabetes (T1D) diagnosed below the age of 7 years between the PTPRK and thymocyte-selection-associated (THEMIS) genes. As the thymus plays a central role in shaping the T cell repertoire, we aimed to identify the most likely causal genetic factors behind this association using thymocyte genomic data. In four thymocyte populations, we identified 253 DNA sequence motifs underlying histone modifications. The G insertion allele of rs138300818, associated with protection from diabetes, created thymocyte motifs for multiple histone modifications and thymocyte types. In a parallel approach to identifying variants that alter transcription factor binding motifs, the same variant disrupted a predicted motif for Rfx7, which is abundantly expressed in the thymus. Chromatin state and RNA sequencing data suggested strong transcription overlapping rs138300818 in fetal thymus, while expression quantitative trait locus and chromatin conformation data associate the insertion with lower THEMIS expression. Extending the analysis to other T1D loci further highlighted rs66733041 affecting the GATA3 transcription factor binding in the AFF3 locus. Taken together, our results support a role for thymic THEMIS gene expression and the rs138300818 variant in promoting the development of early-onset T1D.

Interferon-α-mediated therapeutic resistance in early rheumatoid arthritis implicates epigenetic reprogramming

ObjectivesAn interferon (IFN) gene signature (IGS) is present in approximately 50% of early, treatment naive rheumatoid arthritis (eRA) patients where it has been shown to negatively impact initial response to...

H1.0 C Terminal Domain Is Integral for Altering Transcription Factor Binding within Nucleosomes.

The linker histone H1 is a highly prevalent protein that compacts chromatin and regulates DNA accessibility and transcription. However, the mechanisms behind H1 regulation of transcription factor (TF) binding within nucleosomes are not well understood. Using in vitro fluorescence assays, we positioned fluorophores throughout human H1 and the nucleosome, then monitored the distance changes between H1 and the histone octamer, H1 and nucleosomal DNA, or nucleosomal DNA and the histone octamer to monitor the H1 movement during TF binding. We found that H1 remains bound to the nucleosome dyad, while the C terminal domain (CTD) releases the linker DNA during nucleosome partial unwrapping and TF binding. In addition, mutational studies revealed that a small 16 amino acid region at the beginning of the H1 CTD is largely responsible for altering nucleosome wrapping and regulating TF binding within nucleosomes. We then investigated physiologically relevant post-translational modifications (PTMs) in human H1 by preparing fully synthetic H1 using convergent hybrid phase native chemical ligation. Both individual PTMs and combinations of phosphorylation and citrullination of H1 had no detectable influence on nucleosome binding and nucleosome wrapping, and had only a minor impact on H1 regulation of TF occupancy within nucleosomes. This suggests that these H1 PTMs function by other mechanisms. Our results highlight the importance of the H1 CTD, in particular, the first 16 amino acids, in regulating nucleosome linker DNA dynamics and TF binding within the nucleosome.

Mutations and variants of ONECUT1 in diabetes

Genes involved in distinct diabetes types suggest shared disease mechanisms. Here we show that One Cut Homeobox 1 (ONECUT1) mutations cause monogenic recessive syndromic diabetes in two unrelated patients, characterized by intrauterine growth retardation, pancreas hypoplasia and gallbladder agenesis/hypoplasia, and early-onset diabetes in heterozygous relatives. Heterozygous carriers of rare coding variants of ONECUT1 define a distinctive subgroup of diabetic patients with early-onset, nonautoimmune diabetes, who respond well to diabetes treatment. In addition, common regulatory ONECUT1 variants are associated with multifactorial type 2 diabetes. Directed differentiation of human pluripotent stem cells revealed that loss of ONECUT1 impairs pancreatic progenitor formation and a subsequent endocrine program. Loss of ONECUT1 altered transcription factor binding and enhancer activity and NKX2.2/NKX6.1 expression in pancreatic progenitor cells. Collectively, we demonstrate that ONECUT1 controls a transcriptional and epigenetic machinery regulating endocrine development, involved in a spectrum of diabetes, encompassing monogenic (recessive and dominant) as well as multifactorial inheritance. Our findings highlight the broad contribution of ONECUT1 in diabetes pathogenesis, marking an important step toward precision diabetes medicine.

Genetic and Epigenetic Fine-Tuning of TGFB1 Expression Within the Human Osteoarthritic Joint.

Osteoarthritis (OA) is an age-related disease characterized by articular cartilage degeneration. It is largely heritable, and genetic screening has identified single-nucleotide polymorphisms (SNPs) marking genomic risk loci. One such locus is marked by the G>A SNP rs75621460, downstream of TGFB1. This gene encodes transforming growth factor β1, the correct expression of which is essential for cartilage maintenance. This study investigated the regulatory activity of rs75621460 to characterize its impact on TGFB1 expression in disease-relevant patient samples (n = 319) and in Tc28a2 immortalized chondrocytes. Articular cartilage samples from human patients were genotyped, and DNA methylation levels were quantified using pyrosequencing. Gene reporter and electrophoretic mobility shift assays were used to determine differential nuclear protein binding to the region. The functional impact of DNA methylation on TGFB1 expression was tested using targeted epigenome editing. The analyses showed that SNP rs75621460 was located within a TGFB1 enhancer region, and the OA risk allele A altered transcription factor binding, with decreased enhancer activity. Protein complexes binding to A (but not G) induced DNA methylation at flanking CG dinucleotides. Strong correlations between patient DNA methylation levels and TGFB1 expression were observed, with directly opposing effects in the cartilage and the synovium at this locus. This demonstrated biologic pleiotropy in the impact of the SNP within different tissues of the articulating joint. The OA risk SNP rs75621460 impacts TGFB1 expression by modulating the function of a gene enhancer. We propose a mechanism by which the SNP impacts enhancer function, providing novel biologic insight into one mechanism of OA genetic risk, which may facilitate the development of future pharmacologic therapies.

Regulatory SNPs: Altered Transcription Factor Binding Sites Implicated in Complex Traits and Diseases.

The vast majority of the genetic variants (mainly SNPs) associated with various human traits and diseases map to a noncoding part of the genome and are enriched in its regulatory compartment, suggesting that many causal variants may affect gene expression. The leading mechanism of action of these SNPs consists in the alterations in the transcription factor binding via creation or disruption of transcription factor binding sites (TFBSs) or some change in the affinity of these regulatory proteins to their cognate sites. In this review, we first focus on the history of the discovery of regulatory SNPs (rSNPs) and systematized description of the existing methodical approaches to their study. Then, we brief the recent comprehensive examples of rSNPs studied from the discovery of the changes in the TFBS sequence as a result of a nucleotide substitution to identification of its effect on the target gene expression and, eventually, to phenotype. We also describe state-of-the-art genome-wide approaches to identification of regulatory variants, including both making molecular sense of genome-wide association studies (GWAS) and the alternative approaches the primary goal of which is to determine the functionality of genetic variants. Among these approaches, special attention is paid to expression quantitative trait loci (eQTLs) analysis and the search for allele-specific events in RNA-seq (ASE events) as well as in ChIP-seq, DNase-seq, and ATAC-seq (ASB events) data.

Altering transcription factor binding reveals comprehensive transcriptional kinetics of a basic gene.

Transcription is a vital process activated by transcription factor (TF) binding. The active gene releases a burst of transcripts before turning inactive again. While the basic course of transcription is well understood, it is unclear how binding of a TF affects the frequency, duration and size of a transcriptional burst. We systematically varied the residence time and concentration of a synthetic TF and characterized the transcription of a synthetic reporter gene by combining single molecule imaging, single molecule RNA-FISH, live transcript visualisation and analysis with a novel algorithm, Burst Inference from mRNA Distributions (BIRD). For this well-defined system, we found that TF binding solely affected burst frequency and variations in TF residence time had a stronger influence than variations in concentration. This enabled us to device a model of gene transcription, in which TF binding triggers multiple successive steps before the gene transits to the active state and actual mRNA synthesis is decoupled from TF presence. We quantified all transition times of the TF and the gene, including the TF search time and the delay between TF binding and the onset of transcription. Our quantitative measurements and analysis revealed detailed kinetic insight, which may serve as basis for a bottom-up understanding of gene regulation.

The Mediator complex kinase module is necessary for fructose regulation of liver glycogen levels through induction of glucose-6-phosphatase catalytic subunit (G6pc)

ObjectiveLiver glycogen levels are dynamic and highly regulated by nutrient availability as the levels decrease during fasting and are restored during the feeding cycle. However, feeding in the presence of fructose in water suppresses glycogen accumulation in the liver by upregulating the expression of the glucose-6-phosphatase catalytic subunit (G6pc) gene, although the exact mechanism is unknown. We generated liver-specific knockout MED13 mice that lacked the transcriptional Mediator complex kinase module to examine its effect on the transcriptional activation of inducible target gene expression, such as the ChREBP- and FOXO1-dependent control of the G6pc gene promoter. MethodsThe relative changes in liver expression of lipogenic and gluconeogenic genes as well as glycogen levels were examined in response to feeding standard low-fat laboratory chow supplemented with water or water containing sucrose or fructose in control (Med13fl/fl) and liver-specific MED13 knockout (MED13-LKO) mice. ResultsAlthough MED13 deficiency had no significant effect on constitutive gene expression, all the dietary inducible gene transcripts were significantly reduced despite the unchanged insulin sensitivity in the MED13-LKO mice compared to that in the control mice. G6pc gene transcription displayed the most significant difference between the Med13 fl/fl and MED13-LKO mice, particularly when fed fructose. Following fasting that depleted liver glycogen, feeding induced the restoration of glycogen levels except in the presence of fructose. MED13 deficiency rescued the glycogen accumulation defect in the presence of fructose. This resulted from the suppression of G6pc expression and thus G6PC enzymatic activity.Among two transcriptional factors that regulate G6pc gene expression, FOXO1 binding to the G6pc promoter was not affected, whereas ChREBP binding was dramatically reduced in MED13-LKO hepatocytes. In addition, there was a marked suppression of FOXO1 and ChREBP-β transcriptional activities in MED13-LKO hepatocytes. ConclusionsTaken together, our data suggest that the kinase module of the Mediator complex is necessary for the transcriptional activation of metabolic genes such as G6pc and has an important role in regulating glycogen levels in the liver through altering transcription factor binding and activity at the G6pc promoter.

Alpha synuclein (SNCA) rs7684318 variant contributes to Parkinson’s disease risk by altering transcription factor binding related with Notch and Wnt signaling

In view of inconsistencies in the association studies of alpha synuclein (SNCA) rs7684318 (chr4: 90655003 T > C) with Parkinson’s disease (PD), we conducted a meta-analysis to establish the association of this variant with PD and examined changes in transcription factor binding. SNCA rs7684318 C-allele was identified as genetic risk factor for PD in fixed (OR: 1.53, 95 % CI: 1.40–1.68, p < 0.0001) and random effect (OR: 1.65, 95 % CI: 1.30–2.09, p = 0.0003) models. Heterogeneity was observed in association (Tau2: 0.0576, H: 2.32, I2: 0.815, Q: 21.64, p = 0.0002). Egger’s test showed no evidence of publication bias (p = 0.37). Subgroup analysis showed that rs7684318 is contributing to PD risk in Japanese (OR: 1.46, 95 % CI: 1.30–1.64) and Indian (OR: 2.63, 95 % CI: 1.79–3.86) populations while showing no significant association in Chinese population (OR: 1.68, 95 % CI: 0.93–3.02). Sensitivity analysis showed that exclusion of any one of the studies has no significant impact on the association, which justifies the robustness of the analysis. Tissue-specific DNase foot print analysis revealed that this variant contributes to increased transcription factor binding in midbrain, putamen and caudate nucleus. The substitution of T > C increased binding of RBPJ and GATA-family transcription factors; and decreased binding of NKX2 family, SNAI2, SNAI3, DMRT1, HOXA13, HOXB13, HOXC13, HOXD13, WT1, POU4F1, POU4F2, POU4F3 transcriptional factors. TRANSFAC and DNA curvature analyses substantiate the association of this variant with increased binding of GATA1 that contribute to intensity of DNA curvature peaks and splitting pattern. These studies along with the meta-analysis strongly suggest that the rs7684318 variant contributes to the pathophysiology of PD by modulating binding of transcription factors related to Notch and Wnt signalling pathways that are likely to impair dopmanergic transmission.

Integrative analysis identifies bHLH transcription factors as contributors to Parkinson\u2019s disease risk mechanisms

Genome-wide association studies (GWAS) have identified multiple genetic risk signals for Parkinson’s disease (PD), however translation into underlying biological mechanisms remains scarce. Genomic functional annotations of neurons provide new resources that may be integrated into analyses of GWAS findings. Altered transcription factor binding plays an important role in human diseases. Insight into transcriptional networks involved in PD risk mechanisms may thus improve our understanding of pathogenesis. We analysed overlap between genome-wide association signals in PD and open chromatin in neurons across multiple brain regions, finding a significant enrichment in the superior temporal cortex. The involvement of transcriptional networks was explored in neurons of the superior temporal cortex based on the location of candidate transcription factor motifs identified by two de novo motif discovery methods. Analyses were performed in parallel, both finding that PD risk variants significantly overlap with open chromatin regions harboring motifs of basic Helix-Loop-Helix (bHLH) transcription factors. Our findings show that cortical neurons are likely mediators of genetic risk for PD. The concentration of PD risk variants at sites of open chromatin targeted by members of the bHLH transcription factor family points to an involvement of these transcriptional networks in PD risk mechanisms.

Allele-specific open chromatin in human iPSC neurons elucidates functional disease variants.

Most neuropsychiatric disease risk variants are in noncoding sequences and lack functional interpretation. Because regulatory sequences often reside in open chromatin, we reasoned that neuropsychiatric disease risk variants may affect chromatin accessibility during neurodevelopment. Using human induced pluripotent stem cell (iPSC)-derived neurons that model developing brains, we identified thousands of genetic variants exhibiting allele-specific open chromatin (ASoC). These neuronal ASoCs were partially driven by altered transcription factor binding, overrepresented in brain gene enhancers and expression quantitative trait loci, and frequently associated with distal genes through chromatin contacts. ASoCs were enriched for genetic variants associated with brain disorders, enabling identification of functional schizophrenia risk variants and their cis-target genes. This study highlights ASoC as a functional mechanism of noncoding neuropsychiatric risk variants, providing a powerful framework for identifying disease causal variants and genes.

Altered Transcription Factor Binding and Gene Bivalency in Islets of Intrauterine Growth Retarded Rats.

Intrauterine growth retardation (IUGR), which induces epigenetic modifications and permanent changes in gene expression, has been associated with the development of type 2 diabetes. Using a rat model of IUGR, we performed ChIP-Seq to identify and map genome-wide histone modifications and gene dysregulation in islets from 2- and 10-week rats. IUGR induced significant changes in the enrichment of H3K4me3, H3K27me3, and H3K27Ac marks in both 2-wk and 10-wk islets, which were correlated with expression changes of multiple genes critical for islet function in IUGR islets. ChIP-Seq analysis showed that IUGR-induced histone mark changes were enriched at critical transcription factor binding motifs, such as C/EBPs, Ets1, Bcl6, Thrb, Ebf1, Sox9, and Mitf. These transcription factors were also identified as top upstream regulators in our previously published transcriptome study. In addition, our ChIP-seq data revealed more than 1000 potential bivalent genes as identified by enrichment of both H3K4me3 and H3K27me3. The poised state of many potential bivalent genes was altered by IUGR, particularly Acod1, Fgf21, Serpina11, Cdh16, Lrrc27, and Lrrc66, key islet genes. Collectively, our findings suggest alterations of histone modification in key transcription factors and genes that may contribute to long-term gene dysregulation and an abnormal islet phenotype in IUGR rats.

79: Genetic variants at AGTR2 locus increase risk of preterm birth

Preterm birth (PTB) complicates approximately 10% of pregnancies in the U.S. While genetic factors contribute to PTB, there has been limited replication of specific genetic risk loci associated with PTB. Recently, we conducted the first large-scale, maternal genome-wide association study (GWAS), utilizing 43,568 women from 23andMe, that identified a locus of variants near the angiotensin II receptor type 2 receptor (AGTR2) gene associated with both PTB and gestational length. We hypothesized that a causative variant in this locus influences expression of AGTR2. We assessed genetic variants previously identified in our GWAS located near AGTR2 (combined analysis p-value of 7.2x10-16 in association with gestational length) to identify genotype-dependent regulation of AGTR2 expression. We computationally prioritized disease variants using available functional genomics data. Implicated variants were then evaluated for possible effects on transcription factor (TF) binding using protein-binding microarray data. Selected variants predicted to alter TF binding were analyzed using electrophoretic mobility shift assays (EMSAs). Luciferase reporter assays were performed to identify rs7889204 genotype-dependent regulation of the AGTR2 promoter. Our computational analyses identified rs7889204 as the top functional candidate variant. EMSA analysis confirmed predicted allele-dependent binding for the transcription factors CEBPB and HOXA10. Luciferase reporter assays revealed allele-dependent reporter activity for rs7889204, with decreased expression for the non-reference “C” allele. We identified candidate mechanisms in which rs7889204 at the AGTR2 locus differentially binds CEBPB and HOXA10. Reporter assays demonstrate decreased enhancer activity for the non-reference allele at rs7889204. These results are consistent with a previously established eQTL for AGTR2 in the uterus, suggesting decreased binding of CEBPB and HOXA10 to the preterm birth rs7889204 risk allele results in decreased AGTR2 expression and possible downstream decidual implantation abnormalities leading to PTB.View Large Image Figure ViewerDownload Hi-res image Download (PPT)

Multiple selective sweeps of ancient polymorphisms in and around LT\u03b1 located in the MHC class III region on chromosome 6

BackgroundLymphotoxin-α (LTα), located in the Major Histocompatibility Complex (MHC) class III region on chromosome 6, encodes a cytotoxic protein that mediates a variety of antiviral responses among other biological functions. Furthermore, several genotypes at this gene have been implicated in the onset of a number of complex diseases, including myocardial infarction, autoimmunity, and various types of cancer. However, little is known about levels of nucleotide variation and linkage disequilibrium (LD) in and near LTα, which could also influence phenotypic variance. To address this gap in knowledge, we examined sequence variation across ~ 10 kilobases (kbs), encompassing LTα and the upstream region, in 2039 individuals from the 1000 Genomes Project originating from 21 global populations.ResultsHere, we observed striking patterns of diversity, including an excess of intermediate-frequency alleles, the maintenance of multiple common haplotypes and a deep coalescence time for variation (dating > 1.0 million years ago), in global populations. While these results are generally consistent with a model of balancing selection, we also uncovered a signature of positive selection in the form of long-range LD on chromosomes with derived alleles primarily in Eurasian populations. To reconcile these findings, which appear to support different models of selection, we argue that selective sweeps (particularly, soft sweeps) of multiple derived alleles in and/or near LTα occurred in non-Africans after their ancestors left Africa. Furthermore, these targets of selection were predicted to alter transcription factor binding site affinity and protein stability, suggesting they play a role in gene function. Additionally, our data also showed that a subset of these functional adaptive variants are present in archaic hominin genomes.ConclusionsOverall, this study identified candidate functional alleles in a biologically-relevant genomic region, and offers new insights into the evolutionary origins of these loci in modern human populations.

Functional Annotation of Genetic Loci Associated With Sepsis Prioritizes Immune and Endothelial Cell Pathways.

Due to limited sepsis patient cohort size and extreme heterogeneity, only one significant locus and suggestive associations at several independent loci were implicated by three genome-wide association studies. However, genes from such suggestive loci may also provide crucial information to unravel genetic mechanisms that determine sepsis heterogeneity. Therefore, in this study, we made use of integrative approaches to prioritize genes and pathways affected by sepsis associated genetic variants. By integrating expression quantitative trait loci (eQTL) results from the largest whole-blood eQTL database, cytokine QTLs from pathogen-stimulated peripheral blood mononuclear cells (PBMCs), publicly available blood transcriptome data from pneumoniae-derived sepsis patients, and transcriptome data from pathogen-stimulated PBMCs, we identified 55 potential genes affected by 39 independent loci. By performing pathway enrichment analysis at these loci we found enrichment of genes for adherences-junction pathway. Finally, we investigated the functional role of the only one GWAS significant SNP rs4957796 on sepsis survival in altering transcription factor binding affinity in monocytes and endothelial cells. We also found that transient deficiency of FER and MAN2A1 affect endothelial response to stimulation, indicating that both FER and MAN2A1 could be the causal genes at this locus. Taken together, our study suggests that in addition to immune pathways, genetic variants may also affect non-immune related pathways.