Biology subjects

Wray, G. A.

Publications and source records attributed to Wray, G. A..

18 recordsLinked to original sources

bioRxiv2026

HiFi-Helper: A reproducible workflow for genome assembly from HiFi reads alone

The PacBio Revio simplifies genome assembly by generating very long reads with very few errors at an affordable price point. Comparative ease of assembly is democratizing access, leading to a larger niche for assembly workflows. HiFi-Helper is a user-friendly snakemake workflow designed to facilitate genome assembly from HiFi data alone. This tool produces a visual summary that provides intuitive feedback on the quality of the resulting assembly and guides assembly parameter optimization. The case studies presented here confirm that HiFi-only assemblies produced by HiFi-helper can meet or exceed the quality of genomes assembled in prior decades with a much lower investment of time and resources.

genomics↗

bioRxiv2026

The gene ivory:mir-193 controls scale type differentiation in Heliconius butterflies

The ivory:mir-193 locus is a genetic hotspot underlying melanic wing pattern variation across Lepidoptera, acting as a master regulator of melanic scale fate. This includes the genus Heliconius, where aposematic mimicry is driven by three scale cell types: light Type I, dark-melanic Type II, and red Type III scales. We tested functions of ivory:mir-193 in Heliconius using CRISPR-induced somatic mosaic knockouts of ivory and mir-193 across multiple pattern morphs. Knockouts converted Type II scales to Type I scales. Effects on Type III scales varied among and within individuals, indicating that ivory:mir-193 is permissive for Type III development. Using single-nucleus RNA-seq, we profiled the transcriptional landscape of H. melpomene-{Delta}78k mutants lacking mir-193. Loss of mir-193 produced readthrough transcription, consistent with a model where miR-193 acts as a co-transcriptional terminator. These data show that ivory drives wing pattern variation, while mir-193 mediates downstream diversity in scale fate across Lepidoptera.

evolutionary biology↗

bioRxiv2024

Single-nucleus transcriptomics of wing sexual dimorphism and scale cell specialization in sulphur butterflies

The evolution of sexual secondary characteristics necessitates regulatory factors that confer sexual identity to differentiating tissues and cells. In Colias eurytheme butterflies, males exhibit two specialized wing scale types -- UV-iridescent (UVI) and lanceolate scales -- absent in females and likely integral to male courtship behavior. This study investigates the regulatory mechanisms and single-nucleus transcriptomics underlying these two sexually dimorphic cell types during wing development. We show that Doublesex (Dsx) expression is itself dimorphic and required to repress the UVI cell state in females, while unexpectedly, UVI activation in males is independent from Dsx. In the melanic marginal band, Dsx is required in each sex to enforce the presence of lanceolate scales in males, and their absence in females. Single-nucleus RNAseq reveals that UV-iridescent and lanceolate scale cell precursors each show distinctive gene expression profiles at 40% of pupal development, with marker genes that include regulators of transcription, cell signaling, cytoskeletal patterning, and chitin secretion. Both male-specific cell types share a low expression of the Bric-a-brac (Bab) transcription factor, a key repressor of the UVI fate. Bab ChIP-seq profiling suggests that Bab binds the cis-regulatory regions of gene markers associated to UVI fate, including potential effector genes involved in the regulation of cytoskeletal processes and chitin secretion, and loci showing signatures of recent selective sweeps in an UVI-polymorphic population. These findings open new avenues for exploring wing patterning and scale development, shedding light on the mechanisms driving the specification of sex-specific cell states and the differentiation of specialized cell ultrastructures.

developmental biology↗

bioRxiv2024

An RNA interference approach for functional studies in the sea urchin ands its use in analysis of Nodal signaling gradients

Dicer substrate interfering RNAs (DsiRNAs) destroy targeted transcripts using the RNA-Induced Silencing Complex (RISC) through a process called RNA interference (RNAi). This process is ubiquitous among eukaryotes. Here we report the utility of DsiRNA in embryos of the sea urchin Lytechinus variagatus (Lv). Specific knockdowns phenocopy known morpholino and inhibitor knockdowns, and DsiRNA offers a useful alternative to morpholinos. Methods for designing and obtaining specific DsiRNAs that lead to destruction of targeted mRNA are described. DsiRNAs directed against pks1, an enzyme necessary for pigment production, show how successful DsiRNA perturbations are monitored by RNA in situ analysis and by qPCR to determine relative destruction of targeted mRNA. DsiRNA-based knockdowns phenocopy morpholino- and drug-based inhibition of nodal and lefty. Other knockdowns demonstrate that the RISC operates early in development as well as on genes that are first transcribed hours after gastrulation is completed. Thus, DsiRNAs effectively mediate destruction of targeted mRNA in the sea urchin embryo. The approach offers significant advantages over other widely used methods in the urchin in terms of cost, and ease of procurement, and offers sizeable experimental advantages in terms of ease of handling, injection, and knockdown validation. HighlightsDsiRNA provides an RNAi approach for perturbation of sea urchin embryos. A dilution series of DsiRNA oligos reveals properties of the Nodal gradient in establishing the Dorsal-Ventral axis.

developmental biology↗

bioRxiv2024

Lepidopteran scale cells derive from sensory organ precursors through a canonical lineage

The success of butterflies and moths is tightly linked to the origin of scales within the group. A long-standing hypothesis postulates that scales are homologous to the well-described mechanosensory bristles found in the fruit fly Drosophila melanogaster, where both derive from an epithelial precursor specified by lateral inhibition that then undergoes multiple rounds of division. Previous histological examination and candidate gene approaches identified parallels in genes involved in scale and bristle development. Here, we provide definitive developmental and transcriptomic evidence that the differentiation of lepidopteran scales derives from the canonical cell lineage, known as the Sensory Organ Precursor (SOP). Live imaging in moth and butterfly pupae shows that SOP cells undergo two rounds of asymmetric divisions that first abrogate the neurogenic lineage, and then lead to a differentiated scale precursor and its associated socket cell. Single-nucleus RNA sequencing across a time-series of early pupal development revealed differential gene expression patterns that mirror canonical lineage development, including Notch-Delta signalling components, cell adhesion molecules, cell cycling factors, and terminal cell differentiation markers, suggesting a shared origin of the SOP developmental program. Additionally, we recovered a novel gene, the POU-domain transcription factor pdm3, involved in the proper differentiation of butterfly wing scales. Altogether, these data open up avenues for understanding scale type specification and development, and illustrate how single-cell transcriptomics provide a powerful platform for understanding the evolution of cell types.

developmental biology↗

bioRxiv2024

Single-cell transcriptomics reveals evolutionary reconfiguration of embryonic cell fate specification in the sea urchin Heliocidaris erythrogramma

Altered regulatory interactions during development likely underlie a large fraction of phenotypic diversity within and between species, yet identifying specific evolutionary changes remains challenging. Analysis of single-cell developmental transcriptomes from multiple species provides a powerful framework for unbiased identification of evolutionary changes in developmental mechanisms. Here, we leverage a "natural experiment" in developmental evolution in sea urchins, where a major life history switch recently evolved in the lineage leading to Heliocidaris erythrogramma, precipitating extensive changes in early development. Comparative analyses of scRNA-seq developmental time courses from H. erythrogramma and Lytechinus variegatus (representing the derived and ancestral states respectively) reveals numerous evolutionary changes in embryonic patterning. The earliest cell fate specification events, and the primary signaling center are co-localized in the ancestral dGRN but remarkably, in H. erythrogramma they are spatially and temporally separate. Fate specification and differentiation are delayed in most embryonic cell lineages, although in some cases, these processes are conserved or even accelerated. Comparative analysis of regulator-target gene co-expression is consistent with many specific interactions being preserved but delayed in H. erythrogramma, while some otherwise widely conserved interactions have likely been lost. Finally, specific patterning events are directly correlated with evolutionary changes in larval morphology, suggesting that they are directly tied to the life history shift. Together, these findings demonstrate that comparative scRNA-seq developmental time courses can reveal a diverse set of evolutionary changes in embryonic patterning and provide an efficient way to identify likely candidate regulatory interactions for subsequent experimental validation.

evolutionary biology↗

bioRxiv2024

Contrasting the development of larval and adult body plans during the evolution of biphasic lifecycles in sea urchins

Biphasic lifecycles are widespread among animals, but little is known about how the developmental transition between larvae and adults is regulated. Sea urchins are a unique system for studying this phenomenon because of the stark differences between their bilateral larval and pentaradial adult body plans. Here, we use single cell RNA-sequencing to analyze the development of Heliocidaris erythrogramma (He), a sea urchin species with an accelerated, non-feeding mode of larval development. The sequencing time course extends from early embryogenesis to roughly a day before the onset of metamorphosis in He larvae, which is a period that has not been covered by previous datasets. We find that the non-feeding developmental strategy of He is associated with several changes in the specification of larval cell types compared to sea urchins with feeding larvae, such as the loss of a larva-specific skeletal cell population. Furthermore, the development of the larval and adult body plans in sea urchins may utilize largely different sets of regulatory genes. These findings lay the groundwork for extending existing developmental gene regulatory networks to cover additional stages of biphasic lifecycles. Summary statementAnalysis of a new single cell transcriptomic atlas of sea urchin development reveals the rapid evolution of larval cell type trajectories and provides a candidate list of regulators for adult rudiment development.

developmental biology↗

bioRxiv2024

A human-specific enhancer fine-tunes radial glia potency and corticogenesis

Humans evolved an extraordinarily expanded and complex cerebral cortex, associated with developmental and gene regulatory modifications1-3. Human accelerated regions (HARs) are highly conserved genomic sequences with human-specific nucleotide substitutions. Although there are thousands of annotated HARs, their functional contribution to human-specific cortical development is largely unknown4,5. HARE5 is a HAR transcriptional enhancer of the WNT signaling receptor Frizzled8 (FZD8) active during brain development6. Here, using genome-edited mouse and primate models, we demonstrate that human (Hs) HARE5 fine-tunes cortical development and connectivity by controlling the proliferative and neurogenic capacity of neural progenitor cells (NPCs). Hs-HARE5 knock-in mice have significantly enlarged neocortices containing more neurons. By measuring neural dynamics in vivo we show these anatomical features correlate with increased functional independence between cortical regions. To understand the underlying developmental mechanisms, we assess progenitor fate using live imaging, lineage analysis, and single-cell RNA sequencing. This reveals Hs-HARE5 modifies radial glial progenitor behavior, with increased self-renewal at early developmental stages followed by expanded neurogenic potential. We use genome-edited human and chimpanzee (Pt) NPCs and cortical organoids to assess the relative enhancer activity and function of Hs-HARE5 and Pt-HARE5. Using these orthogonal strategies we show four human-specific variants in HARE5 drive increased enhancer activity which promotes progenitor proliferation. These findings illustrate how small changes in regulatory DNA can directly impact critical signaling pathways and brain development. Our study uncovers new functions for HARs as key regulatory elements crucial for the expansion and complexity of the human cerebral cortex.

neuroscience↗

bioRxiv2023

Hybrid epigenomes reveal extensive local genetic changes to chromatin accessibility contribute to divergence in embryonic gene expression between species

Chromatin accessibility plays an important role in shaping gene expression patterns across development and evolution; however, little is known about the genetic and molecular mechanisms that influence chromatin configuration itself. Because cis and trans influences can both theoretically influence the accessibility of the epigenome, we sought to better characterize the role that both of these mechanisms play in altering chromatin accessibility in two closely related sea urchin species. Using hybrids of the two species, and adapting a statistical framework previously developed for the analysis of cis and trans influences on the transcriptome, we examined how these mechanisms shape the regulatory landscape at three important developmental stages, and compared our results to similar patterns in the transcriptome. We found extensive cis- and trans-based influences on evolutionary changes in chromatin, with cis effects slightly more numerous and larger in effect. Genetic mechanisms influencing gene expression and chromatin configuration are correlated, but differ in several important ways. Maternal influences also appear to have more of an effect on chromatin accessibility than on gene expression, persisting well past the maternal-to-zygotic transition. Furthermore, chromatin accessibility near GRN genes appears to be regulated differently than the rest of the epigenome, and indicates that trans factors may play an outsized role in the configuration of chromatin near these genes. Together, our results represent the first attempt to quantify cis and trans influences on evolutionary divergence in chromatin configuration in an outbred natural study system, and suggest that the regulation of chromatin is more genetically complex than was previously appreciated.

evolutionary biology↗

bioRxiv2022

Genomic insights into Plasmodium vivax population structure and diversity in central Africa

Plasmodium vivax malaria has not traditionally been a major concern in central Africa given the high prevalence of the human Duffy-negative phenotype that is believed to prevent infection. Increasing reports of asymptomatic and symptomatic infections in Duffy-negative individuals throughout Africa raise the possibility that P. vivax is evolving to evade host resistance, but there are few parasite samples with genomic data available from this part of the world. In this study, we perform whole genome sequencing of a new P. vivax isolate from the Democratic Republic of the Congo (DRC) and assess how this central African isolate fits into the global context of this species. We use population genomics methods to show that P. vivax from DRC is similar to other African parasite populations and is not closely related to the non-human primate parasite P. vivax-like. SignificanceThe second most common malaria species to infect humans, Plasmodium vivax, is not considered a major threat to human health in central Africa because people in this region frequently have a genetic variant that prevents the P. vivax species from being able to cause illness. Recent research shows that P. vivax can be found in individuals who should be immune, but there is insufficient data to understand why. Our study investigates the genome of one P. vivax sample collected from central Africa to show that the DRC population is closely related to other P.vivax populations in Africa.

evolutionary biology↗

bioRxiv2022

Recent Reconfiguration Of An Ancient Developmental Gene Regulatory Network In Heliocidaris Sea Urchins

Changes in developmental gene regulatory networks (dGRNs) underlie much of the diversity of life1, but the evolutionary mechanisms that operate on interactions with these networks remain poorly understood. Closely related species with extreme phenotypic divergence provide a valuable window into the genetic and molecular basis for changes in dGRNs and their relationship to adaptive changes in organismal traits. Here we analyze genomes, epigenomes, and transcriptomes during early development in two sea urchin species in the genus Heliocidaris that exhibit highly divergent life histories and in an outgroup species. Signatures of positive selection and changes in chromatin status within putative gene regulatory elements are both enriched on the branch leading to the derived life history, and particularly so near dGRN genes; in contrast, positive selection within protein-coding regions have at most a modest enrichment in branch and function. Single-cell transcriptomes reveal a dramatic delay in cell fate specification in the derived state, which also has far fewer open chromatin regions, especially near dGRN genes with conserved roles in cell fate specification. Experimentally perturbing the function of three key transcription factors reveals profound evolutionary changes in the earliest events that pattern the embryo, disrupting regulatory interactions previously conserved for [~]225 million years. Together, these results demonstrate that natural selection can rapidly reshape developmental gene expression on a broad scale when selective regimes abruptly change and that even highly conserved dGRNs and patterning mechanisms in the early embryo remain evolvable under appropriate ecological circumstances.

evolutionary biology↗

bioRxiv2022

Evolutionary changes in the chromatin landscape reshape a developmental gene regulatory network during rapid life history divergence in sea urchins

Chromatin configuration is highly dynamic during embryonic development in animals, exerting an important point of control in transcriptional regulation. Yet there exists remarkably little information about the role of evolutionary changes in chromatin configuration to the evolution of gene expression and organismal traits. Genome-wide assays of chromatin configuration, coupled with whole-genome alignments, can help address this gap in knowledge in several ways. In this study we present a comparative analysis of regulatory element sequences and accessibility throughout embryogenesis in three sea urchin species with divergent life histories: a lecithotroph Heliocidaris erythrogramma, a closely related planktotroph H. tuberculata, and a distantly related planktotroph Lytechinus variegatus. We identified distinct epigenetic and mutational signatures of evolutionary modifications to the function of putative cis-regulatory elements in H. erythrogramma that have accumulated non-uniformly throughout the genome, suggesting selection, rather than drift, underlies many modifications associated with the derived life history. Specifically, regulatory elements composing the sea urchin developmental gene regulatory network are enriched for signatures of positive selection and accessibility changes which may function to alter binding affinity and access of developmental transcription factors to these sites. Furthermore, regulatory element changes often correlate with divergent expression patterns of genes involved in cell type specification, morphogenesis, and development of other derived traits, suggesting these evolutionary modifications have been consequential for phenotypic evolution in H. erythrogramma. Collectively, our results demonstrate that selective pressures imposed by changes in developmental life history rapidly reshape the cis-regulatory landscape of core developmental genes to generate novel traits and embryonic programs.

evolutionary biology↗

bioRxiv2020

Positive selection within the genomes of SARS-CoV-2 and other Coronaviruses independent of impact on protein function

BackgroundThe emergence of a novel coronavirus (SARS-CoV-2) associated with severe acute respiratory disease (COVID-19) has prompted efforts to understand the genetic basis for its unique characteristics and its jump from non-primate hosts to humans. Tests for positive selection can identify apparently nonrandom patterns of mutation accumulation within genomes, highlighting regions where molecular function may have changed during the origin of a species. Several recent studies of the SARS-CoV-2 genome have identified signals of conservation and positive selection within the gene encoding Spike protein based on the ratio of synonymous to nonsynonymous substitution. Such tests cannot, however, detect changes in the function of RNA molecules. MethodsHere we apply a test for branch-specific oversubstitution of mutations within narrow windows of the genome without reference to the genetic code. ResultsWe recapitulate the finding that the gene encoding Spike protein has been a target of both purifying and positive selection. In addition, we find other likely targets of positive selection within the genome of SARS-CoV-2, specifically within the genes encoding Nsp4 and Nsp16. Homology-directed modeling indicates no change in either Nsp4 or Nsp16 protein structure relative to the most recent common ancestor. Thermodynamic modeling of RNA stability and structure, however, indicates that RNA secondary structure within both genes in the SARS-CoV-2 genome differs from those of RaTG13, the reconstructed common ancestor, and Pan-CoV-GD (Guangdong). These SARS-CoV-2-specific mutations may affect molecular processes mediated by the positive or negative RNA molecules, including transcription, translation, RNA stability, and evasion of the host innate immune system. Our results highlight the importance of considering mutations in viral genomes not only from the perspective of their impact on protein structure, but also how they may impact other molecular processes critical to the viral life cycle.

evolutionary biology↗

bioRxiv2020

An early cell shape transition drives evolutionary expansion of the human forebrain

The human brain has undergone rapid expansion since humans diverged from other great apes, but the mechanism of this human-specific enlargement is still unknown. Here, we use cerebral organoids derived from human, gorilla and chimpanzee cells to study developmental mechanisms driving evolutionary brain expansion. We find that the differentiation of neuroepithelial cells to neurogenic radial glia is a protracted process in apes, involving a previously unrecognized transition state characterized by a change in cell shape. Furthermore, we show that human organoids are larger due to a delay in this transition. Temporally resolved RNA-seq from human and gorilla organoids reveals differences in gene expression patterns associated with cell morphogenesis, and in particular highlights ZEB2, a known regulator of epithelial-mesenchymal transition and cell shape. We show, through loss- and gain-of-function experiments, that ZEB2 promotes the progression of neuroepithelial differentiation, and its ectopic overexpression in human is sufficient to trigger a premature transition. Thus, by mimicking the nonhuman ape expression in human organoids, we are able to force the acquisition of nonhuman ape architecture, establishing for the first time, an instructive role of neuroepithelial cell shape in human brain expansion.

evolutionary biology↗

bioRxiv2019

Identifying branch-specific positive selection throughout the regulatory genome using an appropriate neutral proxy

Adaptive changes in cis-regulatory elements are an essential component of evolution by natural selection. Identifying adaptive and functional noncoding DNA elements throughout the genome is therefore crucial for understanding the relationship between phenotype and genotype. Here, we introduce a method we called adaptyPhy, which adds significant improvements to our earlier method that tests for branch-specific directional selection in noncoding sequences. The motivation for these improvements is to provide a more sensitive and better targeted characterization of directional selection and neutral evolution across the genome. We use ENCODE annotations to identify appropriate proxy neutral sequences and demonstrate that the conservativeness of the test can be modulated during the filtration of reference alignments. We apply the method to noncoding Human Accelerated Elements as well as open chromatin elements previously identified in 125 human tissues and cell lines to demonstrate its utility. We also simulate sequence alignments under different classes of evolution in order to validate the ability of adaptiPhy to distinguish positive selection from relaxation of constraint and neutral evolution. Finally, we evaluate the impact of query region length, proxy neutral sequence length, and branch count on test sensitivity.

evolutionary biology↗

bioRxiv2019

Evolution of abbreviated development in Heliocidaris erythrogramma dramatically re-wired the highly conserved sea urchin developmental gene regulatory network to decouple signaling center function from ultimate fate

Developmental gene regulatory networks (GRNs) describe the interactions among gene products that drive the differential transcriptional and cell regulatory states that pattern the embryo and specify distinct cell fates. GRNs are often deeply conserved, but whether this is the product of constraint inherent to the network structure or stabilizing selection remains unclear. We have constructed the first formal GRN for early development in Heliocidaris erythrogramma, a species with dramatically accelerated, direct development. This life history switch has important ecological consequences, arose rapidly, and has evolved independently many times in echinoderms, suggesting it is a product of selection. We find that H. erythrogramma exhibits dramatic differences in GRN topology compared with ancestral, indirect-developing sea urchins. In particular, the GRN sub-circuit that directs the early and autonomous commitment of skeletogenic cell precursors in indirect developers appears to be absent in H. erythrogramma, a particularly striking change in relation to both the prior conservation of this sub-circuit and the key role that these cells play ancestrally in early development as the embryonic signaling center. These results show that even highly conserved molecular mechanisms of early development can be substantially reconfigured in a relatively short evolutionary time span, suggesting that selection rather than constraint is responsible for the striking conservation of the GRN among other sea urchins.

developmental biology↗

bioRxiv2019

Scalable method for groupwise comparisons and its application to chromatin accessibility among primates

Changes in transcriptional regulation are thought to be a major contributor to the evolution of phenotypic traits, but the contribution of changes in chromatin accessibility to the evolution of gene expression remains almost entirely unknown. To address this important gap in knowledge, we developed a new method to identify DNase I Hypersensitive (DHS) sites with differential chromatin accessibility between species using a joint modeling approach. Our method overcomes several limitations inherent to conventional threshold-based pairwise comparisons that become increasingly apparent as the number of species analyzed rises. Our approach employs a single quantitative test which is more sensitive than existing pairwise methods. To illustrate, we applied our joint approach to DHS sites in fibroblast cells from five primates (human, chimpanzee, gorilla, orangutan, and rhesus macaque). We identified 89,744 DHS sites, of which 41% are identified as differential between species using the joint model compared with 33% using the conventional pairwise approach. The joint model provides a principled approach to distinguishing single from multiple chromatin accessibility changes among species. We found that non differential DHS sites are enriched for nucleotide conservation. Differential DHS sites with decreased chromatin accessibility relative to rhesus macaque occur more commonly near transcription start sites (TSS), while those with increased chromatin accessibility occur more commonly distal to TSS. Further, differential DHS sites near TSS are less cell type-specific than more distal regulatory elements. Taken together, these results point to distinct classes of DHS sites, each with distinct characteristics of selection, genomic location, and cell type specificity.

evolutionary biology↗

bioRxiv2017

Evaluation Of Chromatin Accessibility In Prefrontal Cortex Of Schizophrenia Cases And Controls

Schizophrenia genome-wide association (GWA) studies have identified over 150 regions of the genome that are associated with disease risk, yet there is little evidence that coding mutations contribute to this disorder. To explore the mechanism of non-coding regulatory elements in schizophrenia, we performed ATAC-seq on adult prefrontal cortex brain samples from 135 individuals with schizophrenia and 137 controls, and identified 118,152 ATAC-seq peaks. These accessible chromatin regions in brain are highly enriched for SNP-heritability for schizophrenia (10.6 fold enrichment, P=2.4x10-4, second only to genomic regions conserved in Eutherian mammals) and replicated in an independent dataset (9.0 fold enrichment, P=2.7x10-4). This degree of enrichment of schizophrenia heritability was higher than in open chromatin found in 138 different cell and tissue types. Brain open chromatin regions that overlapped highly conserved regions exhibited an even higher degree of heritability enrichment, indicating that conservation can identify functional subsets within regulatory elements active in brain. However, we did not identify chromatin accessibility differences between schizophrenia cases and controls, nor did we find an interaction of chromatin QTLs with case-control status. This indicates that although causal variants map within regulatory elements, mechanisms other than differential chromatin may govern the contribution of regulatory element variation to schizophrenia risk. Our results strongly implicate gene regulatory processes involving open chromatin in the pathogenesis of schizophrenia, and suggest a strategy to understand the hundreds of common variants emerging from large genomic studies of complex brain diseases.

genomics↗