How somatic mutations accumulate in normal cells is poorly understood. A comprehensive analysis of RNA sequencing data from ~6700 samples across 29 normal tissues revealed multiple somatic variants, ...demonstrating that macroscopic clones can be found in many normal tissues. We found that sun-exposed skin, esophagus, and lung have a higher mutation burden than other tested tissues, which suggests that environmental factors can promote somatic mosaicism. Mutation burden was associated with both age and tissue-specific cell proliferation rate, highlighting that mutations accumulate over both time and number of cell divisions. Finally, normal tissues were found to harbor mutations in known cancer genes and hotspots. This study provides a broad view of macroscopic clonal expansion in human tissues, thus serving as a foundation for associating clonal expansion with environmental factors, aging, and risk of disease.
Post-sequencing quality control is a crucial component of RNA sequencing (RNA-seq) data generation and analysis, as sample quality can be affected by sample storage, extraction and sequencing ...protocols. RNA-seq is increasingly applied to cohorts ranging from hundreds to tens of thousands of samples in size, but existing tools do not readily scale to these sizes, and were not designed for a wide range of sample types and qualities. Here, we describe RNA-SeQC 2, an efficient reimplementation of RNA-SeQC (DeLuca et al., 2012) that adds multiple metrics designed to characterize sample quality across a wide range of RNA-seq protocols.
The command-line tool, documentation and C++ source code are available at the GitHub repository https://github.com/getzlab/rnaseqc. Code and data for reproducing the figures in this paper are available at https://github.com/getzlab/rnaseqc2-paper.
Supplementary data are available at Bioinformatics online.
Tens of thousands of genetic variants associated with gene expression (cis-eQTLs) have been discovered in the human population. These eQTLs are active in various tissues and contexts, but the ...molecular mechanisms of eQTL variability are poorly understood, hindering our understanding of genetic regulation across biological contexts. Since many eQTLs are believed to act by altering transcription factor (TF) binding affinity, we hypothesized that analyzing eQTL effect size as a function of TF level may allow discovery of mechanisms of eQTL variability. Using GTEx Consortium eQTL data from 49 tissues, we analyzed the interaction between eQTL effect size and TF level across tissues and across individuals within specific tissues and generated a list of 10,098 TF-eQTL interactions across 2,136 genes that are supported by at least two lines of evidence. These TF-eQTLs were enriched for various TF binding measures, supporting with orthogonal evidence that these eQTLs are regulated by the implicated TFs. We also found that our TF-eQTLs tend to overlap genes with gene-by-environment regulatory effects and to colocalize with GWAS loci, implying that our approach can help to elucidate mechanisms of context-specificity and trait associations. Finally, we highlight an interesting example of IKZF1 TF regulation of an APBB1IP gene eQTL that colocalizes with a GWAS signal for blood cell traits. Together, our findings provide candidate TF mechanisms for a large number of eQTLs and offer a generalizable approach for researchers to discover TF regulators of genetic variant effects in additional QTL datasets.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
BACKGROUND:The human heart requires a complex ensemble of specialized cell types to perform its essential function. A greater knowledge of the intricate cellular milieu of the heart is critical to ...increase our understanding of cardiac homeostasis and pathology. As recent advances in low-input RNA sequencing have allowed definitions of cellular transcriptomes at single-cell resolution at scale, we have applied these approaches to assess the cellular and transcriptional diversity of the nonfailing human heart.
METHODS:Microfluidic encapsulation and barcoding was used to perform single nuclear RNA sequencing with samples from 7 human donors, selected for their absence of overt cardiac disease. Individual nuclear transcriptomes were then clustered based on transcriptional profiles of highly variable genes. These clusters were used as the basis for between-chamber and between-sex differential gene expression analyses and intersection with genetic and pharmacologic data.
RESULTS:We sequenced the transcriptomes of 287 269 single cardiac nuclei, revealing 9 major cell types and 20 subclusters of cell types within the human heart. Cellular subclasses include 2 distinct groups of resident macrophages, 4 endothelial subtypes, and 2 fibroblast subsets. Comparisons of cellular transcriptomes by cardiac chamber or sex reveal diversity not only in cardiomyocyte transcriptional programs but also in subtypes involved in extracellular matrix remodeling and vascularization. Using genetic association data, we identified strong enrichment for the role of cell subtypes in cardiac traits and diseases. Intersection of our data set with genes on cardiac clinical testing panels and the druggable genome reveals striking patterns of cellular specificity.
CONCLUSIONS:Using large-scale single nuclei RNA sequencing, we defined the transcriptional and cellular diversity in the normal human heart. Our identification of discrete cell subtypes and differentially expressed genes within the heart will ultimately facilitate the development of new therapeutics for cardiovascular diseases.
Understanding gene function and regulation in homeostasis and disease requires knowledge of the cellular and tissue contexts in which genes are expressed. Here, we applied four single-nucleus RNA ...sequencing methods to eight diverse, archived, frozen tissue types from 16 donors and 25 samples, generating a cross-tissue atlas of 209,126 nuclei profiles, which we integrated across tissues, donors, and laboratory methods with a conditional variational autoencoder. Using the resulting cross-tissue atlas, we highlight shared and tissue-specific features of tissue-resident cell populations; identify cell types that might contribute to neuromuscular, metabolic, and immune components of monogenic diseases and the biological processes involved in their pathology; and determine cell types and gene modules that might underlie disease mechanisms for complex traits analyzed by genome-wide association studies.
We apply integrative approaches to expression quantitative loci (eQTLs) from 44 tissues from the Genotype-Tissue Expression project and genome-wide association study data. About 60% of known ...trait-associated loci are in linkage disequilibrium with a cis-eQTL, over half of which were not found in previous large-scale whole blood studies. Applying polygenic analyses to metabolic, cardiovascular, anthropometric, autoimmune, and neurodegenerative traits, we find that eQTLs are significantly enriched for trait associations in relevant pathogenic tissues and explain a substantial proportion of the heritability (40-80%). For most traits, tissue-shared eQTLs underlie a greater proportion of trait associations, although tissue-specific eQTLs have a greater contribution to some traits, such as blood pressure. By integrating information from biological pathways with eQTL target genes and applying a gene-based approach, we validate previously implicated causal genes and pathways, and propose new variant and gene associations for several complex traits, which we replicate in the UK BioBank and BioVU.
Transcriptional regulation and posttranscriptional processing underlie many cellular and organismal phenotypes. We used RNA sequence data generated by Genotype-Tissue Expression (GTEx) project to ...investigate the patterns of transcriptome variation across individuals and tissues. Tissues exhibit characteristic transcriptional signatures that show stability in postmortem samples. These signatures are dominated by a relatively small number of genes–which is most clearly seen in blood–though few are exclusive to a particular tissue and vary more across tissues than individuals. Genes exhibiting high interindividual expression variation include disease candidates associated with sex, ethnicity, and age. Primary transcription is the major driver of cellular specificity, with splicing playing mostly a complementary role; except for the brain, which exhibits a more divergent splicing program. Variation in splicing, despite its stochasticity, may play in contrast a comparatively greater role in defining individual phenotypes.
Genetic studies of the transcriptome help bridge the gap between genetic variation and phenotypes. To maximize the potential of such studies, efficient methods to identify expression quantitative ...trait loci (eQTLs) and perform fine-mapping and genetic prediction of gene expression traits are needed. Current methods that leverage both total read counts and allele-specific expression to identify eQTLs are generally computationally intractable for large transcriptomic studies. Here, we describe a unified framework that addresses these needs and is scalable to thousands of samples. Using simulations and data from GTEx, we demonstrate its calibration and performance. For example, mixQTL shows a power gain equivalent to a 29% increase in sample size for genes with sufficient allele-specific read coverage. To showcase the potential of mixQTL, we apply it to 49 GTEx tissues and find 20% additional eQTLs (FDR < 0.05, per tissue) that are significantly more enriched among trait associated variants and candidate cis-regulatory elements comparing to the standard approach.
Post-mortem tissues samples are a key resource for investigating patterns of gene expression. However, the processes triggered by death and the post-mortem interval (PMI) can significantly alter ...physiologically normal RNA levels. We investigate the impact of PMI on gene expression using data from multiple tissues of post-mortem donors obtained from the GTEx project. We find that many genes change expression over relatively short PMIs in a tissue-specific manner, but this potentially confounding effect in a biological analysis can be minimized by taking into account appropriate covariates. By comparing ante- and post-mortem blood samples, we identify the cascade of transcriptional events triggered by death of the organism. These events do not appear to simply reflect stochastic variation resulting from mRNA degradation, but active and ongoing regulation of transcription. Finally, we develop a model to predict the time since death from the analysis of the transcriptome of a few readily accessible tissues.
PDF
