Variation In Gene Regulation Research Articles

Sequence-based machine learning reveals 3D genome differences between bonobos and chimpanzees.

The 3D structure of the genome is an important mediator of gene expression. As phenotypic divergence is largely driven by gene regulatory variation, comparing genome 3D contacts across species can further understanding of the molecular basis of species differences. However, while experimental data on genome 3D contacts in humans is increasingly abundant, only a handful of 3D genome contact maps exist for other species. Here, we demonstrate that human experimental data can be used to close this data gap. We apply a machine learning model that predicts 3D genome contacts from DNA sequence to the genomes from 56 bonobos and chimpanzees and identify species-specific patterns of genome folding. We estimated 3D divergence between individuals from the resulting contact maps in 4,420 1 Mb genomic windows, of which ∼17% were substantially divergent in predicted genome contacts. Bonobos and chimpanzees diverged at 89 windows, overlapping genes associated with multiple traits implicated in Pan phenotypic divergence. We discovered 51 bonobo-specific variants that individually produce the observed bonobo contact pattern in bonobo-chimpanzee divergent windows. Our results demonstrate that machine learning methods can leverage human data to fill in data gaps across species, offering the first look at population-level 3D genome variation in non-human primates. We also identify loci where changes in 3D folding may contribute to phenotypic differences in our closest living relatives.

Molecular mechanisms underlying gene regulatory variation of maize metabolic traits.

Variation in gene expression levels is pervasive among individuals and races or varieties, and has substantial agronomic consequences, for example, by contributing to hybrid vigor. Gene expression level variation results from mutations in regulatory sequences (cis) and/or transcription factor (TF) activity (trans), but the mechanisms underlying cis- and/or trans-regulatory variation of complex phenotypes remain largely unknown. Here, we investigated gene expression variation mechanisms underlying the differential accumulation of the insecticidal compounds maysin and chlorogenic acid in silks of widely used maize (Zea mays) inbreds, B73 and A632. By combining transcriptomics and cistromics, we identified 1,338 silk direct targets of the maize R2R3-MYB TF Pericarp color1 (P1), consistent with it being a regulator of maysin and chlorogenic acid biosynthesis. Among these P1 targets, 464 showed allele-specific expression (ASE) between B73 and A632 silks. Allelic DNA-affinity purification sequencing identified 34 examples in which P1 allelic specific binding (ASB) correlated with cis-expression variation. From previous yeast one-hybrid studies, we identified 9 TFs potentially implicated in the control of P1 targets, with ASB to 83 out of 464 ASE genes (cis) and differential expression of 4 out of 9 TFs between B73 and A632 silks (trans). These results provide a molecular framework for understanding universal mechanisms underlying natural variation of gene expression levels, and how the regulation of metabolic diversity is established.

Genome resources for three modern cotton lines guide future breeding efforts

Cotton (Gossypium hirsutum L.) is the key renewable fibre crop worldwide, yet its yield and fibre quality show high variability due to genotype-specific traits and complex interactions among cultivars, management practices and environmental factors. Modern breeding practices may limit future yield gains due to a narrow founding gene pool. Precision breeding and biotechnological approaches offer potential solutions, contingent on accurate cultivar-specific data. Here we address this need by generating high-quality reference genomes for three modern cotton cultivars (‘UGA230’, ‘UA48’ and ‘CSX8308’) and updating the ‘TM-1’ cotton genetic standard reference. Despite hypothesized genetic uniformity, considerable sequence and structural variation was observed among the four genomes, which overlap with ancient and ongoing genomic introgressions from ‘Pima’ cotton, gene regulatory mechanisms and phenotypic trait divergence. Differentially expressed genes across fibre development correlate with fibre production, potentially contributing to the distinctive fibre quality traits observed in modern cotton cultivars. These genomes and comparative analyses provide a valuable foundation for future genetic endeavours to enhance global cotton yield and sustainability.

Diverse Gene Regulatory Mechanisms Alter Rattlesnake Venom Gene Expression at Fine Evolutionary Scales.

Understanding and predicting the relationships between genotype and phenotype is often challenging, largely due to the complex nature of eukaryotic gene regulation. A step towards this goal is to map how phenotypic diversity evolves through genomic changes that modify gene regulatory interactions. Using the Prairie Rattlesnake (Crotalus viridis) and related species, we integrate mRNA-seq, proteomic, ATAC-seq and whole-genome resequencing data to understand how specific evolutionary modifications to gene regulatory network components produce differences in venom gene expression. Through comparisons within and between species, we find a remarkably high degree of gene expression and regulatory network variation across even a shallow level of evolutionary divergence. We use these data to test hypotheses about the roles of specific trans-factors and cis-regulatory elements, how these roles may vary across venom genes and gene families, and how variation in regulatory systems drive diversity in venom phenotypes. Our results illustrate that differences in chromatin and genotype at regulatory elements play major roles in modulating expression. However, we also find that enhancer deletions, differences in transcription factor expression, and variation in activity of the insulator protein CTCF also likely impact venom phenotypes. Our findings provide insight into the diversity and gene-specificity of gene regulatory features and highlight the value of comparative studies to link gene regulatory network variation to phenotypic variation.

Tomato root specialized metabolites evolved through gene duplication and regulatory divergence within a biosynthetic gene cluster.

Tremendous plant metabolic diversity arises from phylogenetically restricted specialized metabolic pathways. Specialized metabolites are synthesized in dedicated cells or tissues, with pathway genes sometimes colocalizing in biosynthetic gene clusters (BGCs). However, the mechanisms by which spatial expression patterns arise and the role of BGCs in pathway evolution remain underappreciated. In this study, we investigated the mechanisms driving acylsugar evolution in the Solanaceae. Previously thought to be restricted to glandular trichomes, acylsugars were recently found in cultivated tomato roots. We demonstrated that acylsugars in cultivated tomato roots and trichomes have different sugar cores, identified root-enriched paralogs of trichome acylsugar pathway genes, and characterized a key paralog required for root acylsugar biosynthesis, SlASAT1-LIKE (SlASAT1-L), which is nested within a previously reported trichome acylsugar BGC. Last, we provided evidence that ASAT1-L arose through duplication of its paralog, ASAT1, and was trichome-expressed before acquiring root-specific expression in the Solanum genus. Our results illuminate the genomic context and molecular mechanisms underpinning metabolic diversity in plants.

Human gene regulatory evolution is driven by the divergence of regulatory element function in both cis and trans

Gene regulatory divergence between species can result from cis-acting local changes to regulatory element DNA sequences or global trans-acting changes to the regulatory environment. Understanding how these mechanisms drive regulatory evolution has been limited by challenges in identifying trans-acting changes. We present a comprehensive approach to directly identify cis- and trans-divergent regulatory elements between human and rhesus macaque lymphoblastoid cells using assay for transposase-accessible chromatin coupled to self-transcribing active regulatory region (ATAC-STARR) sequencing. In addition to thousands of cis changes, we discover an unexpected number (∼10,000) of trans changes and show that cis and trans elements exhibit distinct patterns of sequence divergence and function. We further identify differentially expressed transcription factors that underlie ∼37% of trans differences and trace how cis changes can produce cascades of trans changes. Overall, we find that most divergent elements (67%) experienced changes in both cis and trans, revealing a substantial role for trans divergence-alone and together with cis changes-in regulatory differences between species.

Temperature-dependent gene regulatory divergence underlies local adaptation with gene flow in the Atlantic silverside.

Gene regulatory divergence is thought to play an important role in adaptation, yet its extent and underlying mechanisms remain largely elusive for local adaptation with gene flow. Local adaptation is widespread in marine species despite generally high connectivity and is often associated with tightly linked genomic architectures, such as chromosomal inversions. To investigate gene regulatory evolution under gene flow and the role of inversions associated with local adaptation to a steep thermal gradient, we generated RNA-seq data from Atlantic silversides (Menidia menidia) from two locally adapted populations and their F1 hybrids, reared under two temperatures. We found substantial divergence in gene expression and thermal plasticity between populations, with up to 31% of genes being differentially expressed. Reduced thermal plasticity, temperature-dependent gene misexpression, and the disruption of coexpression networks in hybrids point toward a role of regulatory incompatibilities in local adaptation, particularly under colder temperatures. Chromosomal inversions show an accumulation of regulatory incompatibilities but are not consistently enriched for differentially expressed genes. Together, these results suggest that gene regulation can diverge substantially among populations despite gene flow, partly due to the accumulation of temperature-dependent regulatory incompatibilities within inversions.

Evolution and genetics of accessory gland transcriptome divergence between Drosophila melanogaster and D. simulans.

Studies of allele-specific expression in interspecific hybrids have provided important insights into gene-regulatory divergence and hybrid incompatibilities. Many such investigations in Drosophila have used transcriptome data from complex mixtures of many tissues or from gonads, however, regulatory divergence may vary widely among species, sexes, and tissues. Thus, we lack sufficiently broad sampling to be confident about the general biological principles of regulatory divergence. Here, we seek to fill some of these gaps in the literature by characterizing regulatory evolution and hybrid misexpression in a somatic male sex organ, the accessory gland, in F1 hybrids between Drosophila melanogaster and D. simulans. The accessory gland produces seminal fluid proteins, which play an important role in male and female fertility and may be subject to adaptive divergence due to male-male or male-female interactions. We find that trans differences are relatively more abundant than cis, in contrast to most of the interspecific hybrid literature, though large effect-size trans differences are rare. Seminal fluid protein genes have significantly elevated levels of expression divergence and tend to be regulated through both cis and trans divergence. We find limited misexpression (over- or underexpression relative to both parents) in this organ compared to most other Drosophila studies. As in previous studies, male-biased genes are overrepresented among misexpressed genes and are much more likely to be underexpressed. ATAC-Seq data show that chromatin accessibility is correlated with expression differences among species and hybrid allele-specific expression. This work identifies unique regulatory evolution and hybrid misexpression properties of the accessory gland and suggests the importance of tissue-specific allele-specific expression studies.

Environmentally robust cis-regulatory changes underlie rapid climatic adaptation

Changes in gene expression are thought to play a major role in adaptive evolution. While it is known that gene expression is highly sensitive to the environment, very few studies have determined the influence of genetic and environmental effects on adaptive gene expression differences in natural populations. Here, we utilize allele-specific expression to characterize cis and trans gene regulatory divergence in temperate and tropical house mice in two metabolic tissues under two thermal conditions. First, we show that gene expression divergence is pervasive between populations and across thermal conditions, with roughly 5 to 10% of genes exhibiting genotype-by-environment interactions. Second, we found that most expression divergence was due to cis-regulatory changes that were stable across temperatures. In contrast, patterns of expression plasticity were largely attributable to trans-effects, which showed greater sensitivity to temperature. Nonetheless, we found a small subset of temperature-dependent cis-regulatory changes, thereby identifying loci underlying expression plasticity. Finally, we performed scans for selection in wild house mice to identify genomic signatures of rapid adaptation. Genomic outliers were enriched in genes with evidence for cis-regulatory divergence. Notably, these genes were associated with phenotypes that affected body weight and metabolism, suggesting that cis-regulatory changes are a possible mechanism for adaptive body size evolution between populations. Our results show that gene expression plasticity, largely controlled in trans, may facilitate the colonization of new environments, but that evolved changes in gene expression are largely controlled in cis, illustrating the genetic and nongenetic mechanisms underlying the establishment of populations in new environments.

Transcriptional misexpression in hybrids between species linked by gene flow is associated with patterns of sequence divergence.

The extent to which hybridization disrupts a gene's pattern of expression likely governs its propensity for introgression, while its extent of molecular divergence can itself underlie such disruption. Together, these phenomena shape the landscape of sequence and transcriptional divergence across the genome as species diverge. To understand this process, we characterize gene expression inheritance, regulatory divergence, and molecular divergence in the reproductive transcriptomes of species linked by gene flow: the fruit flies Anastrepha fraterculus and A. obliqua, which show evidence of gene flow despite clear evolutionary divergence. We find that their transcriptional patterns are a mosaic between those typically observed within and between allopatric species. Transcripts showing transgressive expression in hybrids or cis-regulatory divergence between species are associated with greater sequence divergence. This may reflect pleiotropic constraints that make them resistant to gene flow or they may be more likely to experience divergent selection. While these more divergent gene classes are likely to be important contributors to species differences, they are relatively rare. Instead, most differentially regulated transcripts, including those linked to reproduction, show high degrees of dominance in hybrids and trans-regulated divergence between species, suggesting widespread genetic compatibility that potentially allowed for introgression. These findings provide insights into how postzygotic isolating mechanisms might evolve in the presence of gene flow: regions showing cis-regulatory divergence or transgressive expression contribute to reproductive isolation, while regions with dominant expression and trans-regulatory divergence allow for introgression. These patterns create a genomic mosaic of transcriptional regulation that is tied to sequence divergence.

Screening for functional regulatory variants in open chromatin using GenIE-ATAC.

Understanding the effects of genetic variation in gene regulatory elements is crucial to interpreting genome function. This is particularly pertinent for the hundreds of thousands of disease-associated variants identified by GWAS, which frequently sit within gene regulatory elements but whose functional effects are often unknown. Current methods are limited in their scalability and ability to assay regulatory variants in their endogenous context, independently of other tightly linked variants. Here, we present a new medium-throughput screening system: genome engineering based interrogation of enhancers assay for transposase accessible chromatin (GenIE-ATAC), that measures the effect of individual variants on chromatin accessibility in their endogenous genomic and chromatin context. We employ this assay to screen for the effects of regulatory variants in human induced pluripotent stem cells, validating a subset of causal variants, and extend our software package (rgenie) to analyse these new data. We demonstrate that this methodology can be used to understand the impact of defined deletions and point mutations within transcription factor binding sites. We thus establish GenIE-ATAC as a method to screen for the effect of gene regulatory element variation, allowing identification and prioritisation of causal variants from GWAS for functional follow-up and understanding the mechanisms of regulatory element function.

Dynamic patterns of gene expression and regulatory variation in the maize seed coat

BackgroundSeed size is an important factor contributing to maize yield, but its molecular mechanism remains unclear. The seed coat, which serves as one of the three components of the maize grain, determines seed size to a certain extent. The seed coat also shares the maternal genotype and is an ideal material for studying heterosis.ResultsIn this study, the self-pollinated seeds of the maize hybrid Yudan888 and its parental lines were continuously collected from 0 day after pollination (DAP) to 15 DAP for phenotyping, cytological observation and RNA-seq. The phenotypic data showed that 3 DAP and 8 DAP are the best time points to study maize seed coat heterosis. Cytological observations indicated that maize seed coat heterosis might be the result of the coordination between cell number and cell size. Furthermore, the RNA-seq results showed that the nonadditive genes changed significantly between 3 and 8 DAP. However, the number of genes expressed additively was not significantly different. Our findings suggest that seed coat heterosis in hybrid is the result of nonadditive expression caused by dynamic changes in genes at different time points during seed expansion and seed coat development. Gene Ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment indicated that genes related to DNA replication, cell cycle regulation, circadian rhythms and metabolite accumulation contributed significantly to hybrid seed coat heterosis.ConclusionMaize seed coat phenotyping allowed us to infer that 3 DAP and 8 DAP are important time points in the study of seed coat heterosis. Our findings provide evidence for genes involved in DNA replication, cell cycle regulation, circadian rhythms and metabolite accumulation in hybrid with high or low parental expression as major contributors to hybrid seed coat heterosis.

Gene regulatory divergence amongst echinoderms underlies appearance of pigment cells in sea urchin development

Larvae of the sea urchin, Strongylocentrotus purpuratus, have pigmented migratory cells implicated in immune defense and gut patterning. The transcription factor SpGcm activates the expression of many pigment cell-specific genes, including those involved in pigment biosynthesis (SpPks1 and SpFmo3) and immune related genes (e.g. SpMif5). Despite the importance of this cell type in sea urchins, pigmented cells are absent in larvae of the sea star, Patiria miniata. In this study, we tested the premises that sea stars lack genes to synthesize echinochrome pigment, that the genes are present but are not expressed in the larvae, or rather that the homologous gene expression does not contribute to echinochrome synthesis. Our results show that orthologs of sea urchin pigment cell-specific genes (PmPks1, PmFmo3-1 and PmMifL1-2) are present in the sea star genome and expressed in the larvae. Although no cell lineage homologous to migratory sea urchin pigment cells is present, dynamic gene activation accomplishes a similar spatial and temporal expression profile. The mechanisms regulating the expression of these genes, though, is highly divergent. In sea stars, PmGcm lacks the central role in pigment gene expression since it is not expressed in PmPks1 and PmFmo3-1-positive cells, and knockdown of Gcm does not abrogate pigment gene expression. Pigment genes are instead expressed in the coelomic mesoderm early in development before later being expressed in the ectoderm. These findings were supported by in situ RNA hybridization and comparative scRNA-seq analyses. We conclude that simply the coexpression of Pks1 and Fmo3 orthologs in cells of the sea star is not sufficient to underlie the emergence of the larval pigment cell in the sea urchin.

Reduced injury risk links sociality to survival in a group-living primate

Sociality has been linked to a longer lifespan in many mammals, including humans. Yet, how sociality results in survival benefits remains unclear. Using 10 years of data and over 1,000 recorded injuries in rhesus macaques (Macaca mulatta), we tested two injury-related mechanisms by which social status and affiliative partners might influence survival. Injuries increased individual risk of death by 3-fold in this dataset. We found that sociality can affect individuals' survival by reducing their risk of injury but had no effect on the probability of injured individuals dying. Both males and females of high social status (measured as female matrilineal rank and male group tenure) and females with more affiliative partners (estimated using the number of female relatives) experienced fewer injuries and thus were less likely to die. Collectively, our results offer rare insights into one mechanism that can mediate the well-known benefits of sociality on an individual's fitness.

The regulatory landscape of multiple brain regions in outbred heterogeneous stock rats.

Heterogeneous Stock (HS) rats are a genetically diverse outbred rat population that is widely used for studying genetics of behavioral and physiological traits. Mapping Quantitative Trait Loci (QTL) associated with transcriptional changes would help to identify mechanisms underlying these traits. We generated genotype and transcriptome data for five brain regions from 88 HS rats. We identified 21 392 cis-QTLs associated with expression and splicing changes across all five brain regions and validated their effects using allele specific expression data. We identified 80 cases where eQTLs were colocalized with genome-wide association study (GWAS) results from nine physiological traits. Comparing our dataset to human data from the Genotype-Tissue Expression (GTEx) project, we found that the HS rat data yields twice as many significant eQTLs as a similarly sized human dataset. We also identified a modest but highly significant correlation between genetic regulatory variation among orthologous genes. Surprisingly, we found less genetic variation in gene regulation in HS rats relative to humans, though we still found eQTLs for the orthologs of many human genes for which eQTLs had not been found. These data are available from the RatGTEx data portal (RatGTEx.org) and will enable new discoveries of the genetic influences of complex traits.

Selection against admixture and gene regulatory divergence in a long-term primate field study.

Genetic admixture is central to primate evolution. We combined 50 years of field observations of immigration and group demography with genomic data from ~9 generations of hybrid baboons to investigate the consequences of admixture in the wild. Despite no obvious fitness costs to hybrids, we found signatures of selection against admixture similar to those described for archaic hominins. These patterns were concentrated near genes where ancestry is strongly associated with gene expression. Our analyses also show that introgression is partially predictable across the genome. This study demonstrates the value of integrating genomic and field data for revealing how "genomic signatures of selection" (e.g., reduced introgression in low-recombination regions) manifest in nature; moreover, it underscores the importance of other primates as living models for human evolution.

Ruminant-specific retrotransposons shape regulatory evolution of bovine immunity.

Cattle are an important livestock species, and mapping the genomic architecture of agriculturally relevant traits such as disease susceptibility is a major challenge in the bovine research community. Lineage-specific transposable elements (TEs) are increasingly recognized to contribute to gene regulatory evolution and variation, but this possibility has been largely unexplored in ruminant genomes. We conducted epigenomic profiling of the type II interferon (IFN) response in bovine cells and found thousands of ruminant-specific TEs including MER41_BT and Bov-A2 elements predicted to act as IFN-inducible enhancer elements. CRISPR knockout experiments in bovine cells established that critical immune factors including IFNAR2 and IL2RB are transcriptionally regulated by TE-derived enhancers. Finally, population genomic analysis of 38 individuals revealed that a subset of polymorphic TE insertions may function as enhancers in modern cattle. Our study reveals that lineage-specific TEs have shaped the evolution of ruminant IFN responses and potentially continue to contribute to immune gene regulatory differences across modern breeds and individuals. Together with previous work in human cells, our findings demonstrate that lineage-specific TEs have been independently co-opted to regulate IFN-inducible gene expression in multiple species, supporting TE co-option as a recurrent mechanism driving the evolution of IFN-inducible transcriptional networks.

Genetic architecture of gene regulation in Indonesian populations identifies QTLs associated with global and local ancestries

SummaryLack of diversity in human genomics limits our understanding of the genetic underpinnings of complex traits, hinders precision medicine, and contributes to health disparities. To map genetic effects on gene regulation in the underrepresented Indonesian population, we have integrated genotype, gene expression, and CpG methylation data from 115 participants across three island populations that capture the major sources of genomic diversity in the region. In a comparison with European datasets, we identify eQTLs shared between Indonesia and Europe as well as population-specific eQTLs that exhibit differences in allele frequencies and/or overall expression levels between populations. By combining local ancestry and archaic introgression inference with eQTLs and methylQTLs, we identify regulatory loci driven by modern Papuan ancestry as well as introgressed Denisovan and Neanderthal variation. GWAS colocalization connects QTLs detected here to hematological traits, and further comparison with European datasets reflects the poor overall transferability of GWAS statistics across diverse populations. Our findings illustrate how population-specific genetic architecture, local ancestry, and archaic introgression drive variation in gene regulation across genetically distinct and in admixed populations and highlight the need for performing association studies on non-European populations.

Dissecting the Pathogenesis of Diabetic Retinopathy Based on the Biological ceRNA Network and Genome Variation Disturbance.

Background Diabetic retinopathy (DR) is the most important manifestation of diabetic microangiopathy. It is essential to explore the gene regulatory relationship and genomic variation disturbance of biological networks in DR progression. Methods In this study, we constructed a comprehensive lncRNA-mRNA ceRNA network of DR procession (CLMN) and explored its topological characteristics. Results Modular and functional analysis indicated that the organization of CLMN performed fundamental and specific functions in diabetes and DR pathology. The differential expression of hub ceRNA nodes and positive correlation reveals the highly connected ceRNA regulation and important roles in the regulating of DR pathology. A large proportion of SNPs in the TFBS, DHS, and enhancer regions of lncRNAs will affect lncRNA transcription and further cause expression variation. Some SNPs were found to disrupt the lncRNA functional elements such as miRNA target binding sites. These results indicate the complex nature of genotypic effects in the disturbing of CLMN and further contribute to gene expression variation and different disease phenotypes. Conclusion The identification of individual genomic variations and analysis of biological network disturbance by these genomic variations will help provide more personalized treatment plans and promote the development of precision medicine for DR.

Single-nuclei chromatin profiling of ventral midbrain reveals cell identity transcription factors and cell-type-specific gene regulatory variation

BackgroundCell types in ventral midbrain are involved in diseases with variable genetic susceptibility, such as Parkinson’s disease and schizophrenia. Many genetic variants affect regulatory regions and alter gene expression in a cell-type-specific manner depending on the chromatin structure and accessibility.ResultsWe report 20,658 single-nuclei chromatin accessibility profiles of ventral midbrain from two genetically and phenotypically distinct mouse strains. We distinguish ten cell types based on chromatin profiles and analysis of accessible regions controlling cell identity genes highlights cell-type-specific key transcription factors. Regulatory variation segregating the mouse strains manifests more on transcriptome than chromatin level. However, cell-type-level data reveals changes not captured at tissue level. To discover the scope and cell-type specificity of cis-acting variation in midbrain gene expression, we identify putative regulatory variants and show them to be enriched at differentially expressed loci. Finally, we find TCF7L2 to mediate trans-acting variation selectively in midbrain neurons.ConclusionsOur data set provides an extensive resource to study gene regulation in mesencephalon and provides insights into control of cell identity in the midbrain and identifies cell-type-specific regulatory variation possibly underlying phenotypic and behavioural differences between mouse strains.