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At least 829 records · Page 46
All sourcesNASA NTRSDOE OSTI
DOE OSTI2024

Discovery of genes that positively affect biomass and stress associated traits in poplar

Woody biomass serves as a renewable resource for various industries, including pulp and paper production, construction, biofuels, and electricity generation. However, the molecular mechanisms behind biomass traits are poorly understood, which significantly curtails the speed and efficiency of their improvement. We used activation tagging to discover genes that can positively affect tree biomass-associated traits. We generated and screened under greenhouse conditions a population of 2,700 independent activation tagging lines. A total of 761 lines, which had significantly and positively affected at least one biomass-associated trait, were discovered. The tag was positioned in the genome for forty lines which were affected in multiple traits and activation of proximal genes validated for a subset. For two lines we fully recapitulated the phenotype of the original lines through overexpression. Moreover, the overexpression led to more pronounced and additional improvements, not observed in the original lines. Importantly, the overexpression of a Fasciclin-like gene (PtaFLA10) and a Patatin-like gene (PtaPAT) was found to substantially improve biomass, with a 40% increase in dry-stem weight, and enhance drought tolerance, respectively. Additionally, PtaPAT overexpression increased cellulose content, which is crucial for biofuel production. Our work shows that the activation tagging approach applied even on a non-genome saturation scale in a poplar tree can be successfully used for the discovery of genes positively modify biomass productivity. Such dominant forward genetics approaches can aid in biotechnological manipulation of woody biomass traits and help unravel the functions and mechanisms of individual genes, gene families, and regulatory modules.

Plant Sciences↗
DOE OSTI2019

CRISPR interference to interrogate genes that control biofilm formation in Pseudomonas fluorescens

Bacterial biofilm formation involves signaling and regulatory pathways that control the transition from motile to sessile lifestyle, production of extracellular polymeric matrix, and maturation of the biofilm 3D structure. Biofilms are extensively studied because of their importance in biomedical, ecological and industrial settings. Gene inactivation is a powerful approach for functional studies but it is often labor intensive, limiting systematic gene surveys to the most tractable bacterial hosts. Here, we adapted the CRISPR interference (CRISPRi) system for use in diverse strain isolates of P. fluorescens, SBW25, WH6 and Pf0-1. We found that CRISPRi is applicable to study complex phenotypes such as cell morphology, motility and biofilm formation over extended periods of time. In SBW25, CRISPRi-mediated silencing of genes encoding the GacA/S two-component system and regulatory proteins associated with the cylic di-GMP signaling messenger produced swarming and biofilm phenotypes similar to those obtained after gene inactivation. Combined with detailed confocal microscopy of biofilms, our study also revealed novel phenotypes associated with extracellular matrix biosynthesis as well as the potent inhibition of SBW25 biofilm formation mediated by the PFLU1114 operon. We conclude that CRISPRi is a reliable and scalable approach to investigate gene networks in the diverse P. fluorescens group.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2019

Evolutionary characteristics of intergenic transcribed regions indicate rare novel genes and widespread noisy transcription in the Poaceae

Abstract Extensive transcriptional activity occurring in intergenic regions of genomes has raised the question whether intergenic transcription represents the activity of novel genes or noisy expression. To address this, we evaluated cross-species and post-duplication sequence and expression conservation of intergenic transcribed regions (ITRs) in four Poaceae species. Among 43,301 ITRs across the four species, 34,460 (80%) are species-specific. ITRs found across species tend to be more divergent in expression and have more recent duplicates compared to annotated genes. To assess if ITRs are functional (under selection), machine learning models were established in Oryza sativa (rice) that could accurately distinguish between phenotype genes and pseudogenes (area under curve-receiver operating characteristic = 0.94). Based on the models, 584 (8%) and 4391 (61%) rice ITRs are classified as likely functional and nonfunctional with high confidence, respectively. ITRs with conserved expression and ancient retained duplicates, features that were not part of the model, are frequently classified as likely-functional, suggesting these characteristics could serve as pragmatic rules of thumb for identifying candidate sequences likely to be under selection. This study also provides a framework to identify novel genes using comparative transcriptomic data to improve genome annotation that is fundamental for connecting genotype to phenotype in crop and model systems.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2018

Pleiotropic and Epistatic Network-Based Discovery: Integrated Networks for Target Gene Discovery

Biological organisms are complex systems that are composed of functional networks of interacting molecules and macro-molecules. Complex phenotypes are the result of orchestrated, hierarchical, heterogeneous collections of expressed genomic variants. However, the effects of these variants are the result of historic selective pressure and current environmental and epigenetic signals, and, as such, their co-occurrence can be seen as genome-wide correlations in a number of different manners. Biomass recalcitrance (i.e., the resistance of plants to degradation or deconstruction, which ultimately enables access to a plant's sugars) is a complex polygenic phenotype of high importance to biofuels initiatives. This study makes use of data derived from the re-sequenced genomes from over 800 different Populus trichocarpa genotypes in combination with metabolomic and pyMBMS data across this population, as well as co-expression and co-methylation networks in order to better understand the molecular interactions involved in recalcitrance, and identify target genes involved in lignin biosynthesis/degradation. A Lines Of Evidence (LOE) scoring system is developed to integrate the information in the different layers and quantify the number of lines of evidence linking genes to target functions. This new scoring system was applied to quantify the lines of evidence linking genes to lignin-related genes and phenotypes across the network layers, and allowed for the generation of new hypotheses surrounding potential new candidate genes involved in lignin biosynthesis in P. trichocarpa, including various AGAMOUS-LIKE genes. Lastly, the resulting Genome Wide Association Study networks, integrated with Single Nucleotide Polymorphism (SNP) correlation, co-methylation, and co-expression networks through the LOE scores are proving to be a powerful approach to determine the pleiotropic and epistatic relationships underlying cellular functions and, as such, the molecular basis for complex phenotypes, such as recalcitrance.

09 BIOMASS FUELS↗
DOE OSTI2017

Function of Nup98 subtypes and their fusion proteins, Nup98-TopIIβ and Nup98-SETBP1 in nuclear-cytoplasmic transport

Nup98 is a component of the nuclear pore complex. The nup98-fusion genes derived by chromosome translocations are involved in hematopoietic malignancies. Here, we investigated the functions of Nup98 isoforms and two unexamined Nup98-fusion proteins, Nup98-TopIIβ and Nup98-SETBP1. We first demonstrated that two Nup98 isoforms are expressed in various mouse tissues and similarly localized in the nucleus and the nuclear envelope. We also showed that Nup98-TopIIβ and Nup98-SETBP1 are localized in the nucleus and partially co-localized with full-length Nup98 and a nuclear export receptor XPO1. We demonstrated that Nup98-TopIIβ and Nup98-SETBP1 negatively regulate the XPO1-mediated protein export. Our results will contribute to the understanding of the molecular mechanism by which the Nup98-fusion proteins induce tumorigenesis. - Highlights: • Two nup98 variants are expressed among various mouse tissues and human cell lines. • Both variants show similar localization patterns. • Nup98-TopIIβ and Nup98-SETBP1 change the localization of full-length Nup98. • Nup98-TopIIβ and Nup98-SETBP1 reduce the nuclear export activity of XPO1.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

A large sequenced mutant library – valuable reverse genetic resource that covers 98% of sorghum genes

SUMMARY Mutant populations are crucial for functional genomics and discovering novel traits for crop breeding. Sorghum , a drought and heat‐tolerant C4 species, requires a vast, large‐scale, annotated, and sequenced mutant resource to enhance crop improvement through functional genomics research. Here, we report a sorghum large‐scale sequenced mutant population with 9.5 million ethyl methane sulfonate (EMS)‐induced mutations that covered 98% of sorghum's annotated genes using inbred line BTx623. Remarkably, a total of 610 320 mutations within the promoter and enhancer regions of 18 000 and 11 790 genes, respectively, can be leveraged for novel research of cis ‐regulatory elements. A comparison of the distribution of mutations in the large‐scale mutant library and sorghum association panel (SAP) provides insights into the influence of selection. EMS‐induced mutations appeared to be random across different regions of the genome without significant enrichment in different sections of a gene, including the 5′ UTR, gene body, and 3′‐UTR. In contrast, there were low variation density in the coding and UTR regions in the SAP. Based on the K a / K s value, the mutant library (~1) experienced little selection, unlike the SAP (0.40), which has been strongly selected through breeding. All mutation data are publicly searchable through SorbMutDB ( https://www.depts.ttu.edu/igcast/sorbmutdb.php ) and SorghumBase ( https://sorghumbase.org/ ). This current large‐scale sequence‐indexed sorghum mutant population is a crucial resource that enriched the sorghum gene pool with novel diversity and a highly valuable tool for the Poaceae family, that will advance plant biology research and crop breeding.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Single-cell RNA sequencing reveals plasmid constrains bacterial population heterogeneity and identifies a non-conjugating subpopulation

Transcriptional heterogeneity in isogenic bacterial populations can play various roles in bacterial evolution, but its detection remains technically challenging. Here, we use microbial split-pool ligation transcriptomics to study the relationship between bacterial subpopulation formation and plasmid-host interactions at the single-cell level. We find that single-cell transcript abundances are influenced by bacterial growth state and plasmid carriage. Moreover, plasmid carriage constrains the formation of bacterial subpopulations. Plasmid genes, including those with core functions such as replication and maintenance, exhibit transcriptional heterogeneity associated with cell activity. Notably, we identify a cell subpopulation that does not transcribe conjugal plasmid transfer genes, which may help reduce plasmid burden on a subset of cells. Our study advances the understanding of plasmid-mediated subpopulation dynamics and provides insights into the plasmid-bacteria interplay.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Emerging Role of Exosomal Long Non-coding RNAs in Spaceflight-Associated Risks in Astronauts

During spaceflight, astronauts are exposed to multiple unique environmental factors, particularly microgravity and ionizing radiation, that can cause a range of harmful health consequences. Over the past decades, increasing evidence demonstrates that the space environment can induce changes in gene expression and RNA processing. Long non-coding RNA (lncRNA) represent an emerging area of focus in molecular biology as they modulate chromatin structure and function, the transcription of neighboring genes, and affect RNA splicing, stability, and translation. They have been implicated in cancer development and associated with diverse cardiovascular conditions and associated risk factors. However, their role on astronauts’ health after spaceflight remains poorly understood. In this perspective article, we provide new insights into the potential role of exosomal lncRNA after spaceflight. We analyzed the transcriptional profile of exosomes isolated from peripheral blood plasma of three astronauts who flew on various Shuttle missions between 1998–2001 by RNA-sequencing. Computational analysis of the transcriptome of these exosomes identified 27 differentially expressed lncRNAs with a Log 2 fold change, with molecular, cellular, and clinical implications.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2017

Early Transcriptomic Changes in the Ileal Pouch Provide Insight into the Molecular Pathogenesis of Pouchitis and Ulcerative Colitis

Background: Ulcerative colitis (UC) only involves the colonic mucosa. Yet, nearly 50% of patients with UC who undergo total proctocolectomy with ileal pouch anal anastomosis develop UC-like inflammation of the ileal pouch (pouchitis). By contrast, patients with familial adenomatous polyposis (FAP) with ileal pouch anal anastomosis develop pouchitis far less frequently. We hypothesized that pathogenic events associated with the development of UC are recapitulated by colonic-metaplastic transcriptomic reprogramming of the UC pouch. Methods: We prospectively sampled pouch and prepouch ileum mucosal biopsies in patients with UC with ileal pouch anal anastomosis 4, 8, and 12 months after their pouch was in continuity. Mucosal samples were also obtained from patients with FAP. Transcriptional profiles of the UC and FAP pouch and prepouch ileum were investigated via RNA sequencing and compared with data from a previously published microarray study. Results: Unlike patients with FAP, subjects with UC exhibited a large set of differentially expressed genes between the pouch and prepouch ileum as early as 4 months after pouch functionalization. Functional pathway analysis of differentially expressed genes in the UC pouch revealed an enhanced state of immune/inflammatory response and extracellular matrix remodeling. Moreover, >70% of differentially expressed genes mapped to published inflammatory bowel diseases microarray data sets displayed directional changes consistent with active UC but not with Crohn's disease. Conclusions: The UC pouch, well before histologic inflammation, already displays a systems-level gain of colon-associated genes and loss of ileum-associated genes. Patients with UC exhibit a unique transcriptomic response to ileal pouch creation that can be observed well before disease and may in part explain their susceptibility to the development of pouchitis.

familial adenomatous polyposis↗
DOE OSTI2021

MultiPhATE2: code for functional annotation and comparison of phage genomes

To address a need for improved tools for annotation and comparative genomics of bacteriophage genomes, we developed multiPhATE2. As an extension of multiPhATE, a functional annotation code released previously, multiPhATE2 performs gene finding using multiple algorithms, compares the results of the algorithms, performs functional annotation of coding sequences, and incorporates additional search algorithms and databases to extend the search space of the original code. MultiPhATE2 performs gene matching among sets of closely related bacteriophage genomes, and uses multiprocessing to speed computations. MultiPhATE2 can be re-started at multiple points within the workflow to allow the user to examine intermediate results and adjust the subsequent computations accordingly. In addition, multiPhATE2 accommodates custom gene calls and sequence databases, again adding flexibility. MultiPhATE2 was implemented in Python 3.7 and runs as a command-line code under Linux or MAC operating systems. Full documentation is provided as a README file and a Wiki website.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2019

Development of a CRISPR/Cas9 System for Methylococcus capsulatus In Vivo Gene Editing

Methanotrophic bacteria play a crucial role in the Earth’s biogeochemical cycle and have the potential to be employed in industrial biomanufacturing processes due to their capacity to use natural gas- and biogas-derived methane as a sole carbon and energy source. Advanced gene-editing systems have the potential to enable rapid, high-throughput methanotrophic genetics and biocatalyst development. To this end, we employed a series of broad-host-range expression plasmids to construct a conjugatable clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 gene-editing system in Methylococcus capsulatus (Bath). Heterologous coexpression of the Streptococcus pyogenes Cas9 endonuclease and a synthetic single guide RNA (gRNA) showed efficient Cas9 DNA targeting and double-stranded DNA (dsDNA) cleavage that resulted in cell death. We demonstrated effective in vivo editing of plasmid DNA using both Cas9 and Cas9 D10A nickase to convert green fluorescent protein (GFP)- to blue fluorescent protein (BFP)-expressing cells with 71% efficiency. Further, we successfully introduced a premature stop codon into the soluble methane monooxygenase (sMMO) hydroxylase component-encoding mmoX gene with the Cas9 D10A nickase, disrupting sMMO function. These data provide proof of concept for CRISPR/Cas9-mediated gene editing in M. capsulatus . Given the broad-host-range replicons and conjugation capability of these CRISPR/Cas9 tools, they have potential utility in other methanotrophs and a wide array of Gram-negative microorganisms.

09 BIOMASS FUELS↗
DOE OSTI2022

Virioplankton assemblages from challenger deep, the deepest place in the oceans

Hadal ocean biosphere, that is, the deepest part of the world’s oceans, harbors a unique microbial community, suggesting a potential uncovered co-occurring virioplankton assemblage. Herein, we reveal the unique virioplankton assemblages of the Challenger Deep, comprising 95,813 non-redundant viral contigs from the surface to the hadal zone. Almost all of the dominant viral contigs in the hadal zone were unclassified, potentially related to Alteromonadales and Oceanospirillales. 2,586 viral auxiliary metabolic genes from 132 different KEGG orthologous groups were mainly related to the carbon, nitrogen, sulfur, and arsenic metabolism. Lysogenic viral production and integrase genes were augmented in the hadal zone, suggesting the prevalence of viral lysogenic life strategy. Abundant rve genes in the hadal zone, which function as transposase in the caudoviruses, further suggest the prevalence of viral-mediated horizontal gene transfer. This study provides fundamental insights into the virioplankton assemblages of the hadal zone, reinforcing the necessity of incorporating virioplankton into the hadal biogeochemical cycles.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2019

Gene Regulation Shifts Shed Light on Fungal Adaption in Plant Biomass Decomposers

ABSTRACT Fungi dominate the recycling of carbon sequestered in woody biomass. This process of organic turnover was first evolved among “white rot” fungi that degrade lignin to access carbohydrates and later evolved multiple times toward more efficient strategies to selectively target carbohydrates—“brown rot.” The brown rot adaption was often explained by mechanisms to deploy reactive oxygen species (ROS) to oxidatively attack wood structures. However, its genetic basis remains unclear, especially in the context of gene contractions of conventional carbohydrate-active enzymes (CAZYs) relative to white rot ancestors. Here, we hypothesized that these apparent gains in brown rot efficiency despite gene losses were due, in part, to upregulation of the retained genes. We applied comparative transcriptomics to multiple species of both rot types grown across a wood wafer to create a gradient of progressive decay and to enable tracking temporal gene expression. Dozens of “decay-stage-dependent” ortho-genes were isolated, narrowing a pool of candidate genes with time-dependent regulation unique to brown rot fungi. A broad comparison of the expression timing of CAZY families indicated a temporal regulatory shift of lignocellulose-oxidizing genes toward early stages in brown rot compared to white rot, enabling the segregation of oxidative treatment ahead of hydrolysis. These key brown rot ROS-generating genes with iron ion binding functions were isolated. Moreover, transcription energy was shifted to be invested on the retained GHs in brown rot fungi to strengthen carbohydrate conversion. Collectively, these results support the hypothesis that gene regulation shifts played a pivotal role in brown rot adaptation. IMPORTANCE Fungi dominate the turnover of wood, Earth’s largest pool of aboveground terrestrial carbon. Fungi first evolved this capacity by degrading lignin to access and hydrolyze embedded carbohydrates (white rot). Multiple lineages, however, adapted faster reactive oxygen species (ROS) pretreatments to loosen lignocellulose and selectively extract sugars (brown rot). This brown rot “shortcut” often coincided with losses (>60%) of conventional lignocellulolytic genes, implying that ROS adaptations supplanted conventional pathways. We used comparative transcriptomics to further pursue brown rot adaptations, which illuminated the clear temporal expression shift of ROS genes, as well as the shift toward synthesizing more GHs in brown rot relative to white rot. These imply that gene regulatory shifts, not simply ROS innovations, were key to brown rot fungal evolution. These results not only reveal an important biological shift among these unique fungi, but they may also illuminate a trait that restricts brown rot fungi to certain ecological niches.

plant biomas↗
DOE OSTI2026

Data for A Fluorescence-Based Transient Expression Assay for the Analysis of Upstream Open Reading Frames in Plants

Scripts for the manuscript "A fluorescence-based transient expression assay for the analysis of upstream open reading frames in plant" by Haas et al. Upstream open reading frames (uORFs) are regulatory elements present in the 5′ leaders of mRNA that can significantly impact downstream gene expression in eukaryotes. In crop engineering, editing of uORFs can provide an avenue to upregulate expression of native genes without the need to add persistent transgenic copies. Even with genome- wide methods to identify translated uORFs such as ribosome profiling, their functional characterization depends on validation through reporter gene assays and mutagenesis studies. Current screening methods for plants use luciferases or protoplasts to measure differential gene expression between wild- type and mutated transcript leaders, which requires tissue processing and/or substrate addition. Here, we present a time- and cost- efficient alternative to investigate transcript leaders by co- expression of two fluorescent proteins in Nicotiana benthamiana leaf tissue and test our assay on genes involved in photoprotection, editing of which could provide a pathway to increase CO2 assimilation during sun–shade transitions.

Gene Editing↗
DOE OSTI2015

Identification and molecular characterization of the switchgrass AP2/ERF transcription factor superfamily, and overexpression of PvERF001 for improvement of biomass characteristics for biofuel

The APETALA2/ethylene response factor (AP2/ERF) superfamily of transcription factors (TFs) plays essential roles in the regulation of various growth and developmental programs including stress responses. Members of these TFs in other plant species have been implicated to play a role in the regulation of cell wall biosynthesis. Here, we identified a total of 207 AP2/ERF TF genes in the switchgrass genome and grouped into four gene families comprised of 25 AP2-, 121 ERF-, 55 DREB (dehydration responsive element binding)-, and 5 RAV (related to API3/VP) genes, as well as a singleton gene not fitting any of the above families. The ERF and DREB subfamilies comprised seven and four distinct groups, respectively. Analysis of exon/intron structures of switchgrass AP2/ERF genes showed high diversity in the distribution of introns in AP2 genes versus a single or no intron in most genes in the ERF and RAV families. The majority of the subfamilies or groups within it were characterized by the presence of one or more specific conserved protein motifs. In silico functional analysis revealed that many genes in these families might be associated with the regulation of responses to environmental stimuli via transcriptional regulation of the response genes. Moreover, these genes had diverse endogenous expression patterns in switchgrass during seed germination, vegetative growth, flower development, and seed formation. Interestingly, several members of the ERF and DREB families were found to be highly expressed in plant tissues where active lignification occurs. These results provide vital resources to select candidate genes to potentially impart tolerance to environmental stress as well as reduced recalcitrance. Furthermore, overexpression of one of the ERF genes ( PvERF001 ) in switchgrass was associated with increased biomass yield and sugar release efficiency in transgenic lines, exemplifying the potential of these TFs in the development of lignocellulosic feedstocks with improved biomass characteristics for biofuels.

09 BIOMASS FUELS↗
DOE OSTI2023

Prolonging genetic circuit stability through adaptive evolution of overlapping genes

Abstract The development of synthetic biological circuits that maintain functionality over application-relevant time scales remains a significant challenge. Here, we employed synthetic overlapping sequences in which one gene is encoded or ‘entangled’ entirely within an alternative reading frame of another gene. In this design, the toxin-encoding relE was entangled within ilvA, which encodes threonine deaminase, an enzyme essential for isoleucine biosynthesis. A functional entanglement construct was obtained upon modification of the ribosome-binding site of the internal relE gene. Using this optimized design, we found that the selection pressure to maintain functional IlvA stabilized the production of burdensome RelE for >130 generations, which compares favorably with the most stable kill-switch circuits developed to date. This stabilizing effect was achieved through a complete alteration of the allowable landscape of mutations such that mutations inactivating the entangled genes were disfavored. Instead, the majority of lineages accumulated mutations within the regulatory region of ilvA. By reducing baseline relE expression, these more ‘benign’ mutations lowered circuit burden, which suppressed the accumulation of relE-inactivating mutations, thereby prolonging kill-switch function. Overall, this work demonstrates the utility of sequence entanglement paired with an adaptive laboratory evolution campaign to increase the evolutionary stability of burdensome synthetic circuits.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Potential for functional divergence in ectomycorrhizal fungal communities across a precipitation gradient

Abstract Functional traits influence the assembly of microbial communities, but identifying these traits in the environment has remained challenging. We studied ectomycorrhizal fungal (EMF) communities inhabiting Populus trichocarpa roots distributed across a precipitation gradient in the Pacific Northwest, USA. We profiled these communities using taxonomic (meta-barcoding) and functional (metagenomic) approaches. We hypothesized that genes involved in fungal drought-stress tolerance and fungal mediated plant water uptake would be most abundant in drier soils. We were unable to detect support for this hypothesis; instead, the abundance of genes involved in melanin synthesis, hydrophobins, aquaporins, trehalose-synthases, and other gene families exhibited no significant shifts across the gradient. Finally, we studied variation in sequence homology for certain genes, finding that fungal communities in dry soils are composed of distinct aquaporin and hydrophobin gene sequences. Altogether, our results suggest that while EMF communities exhibit significant compositional shifts across this gradient, coupled functional turnover, at least as inferred using community metagenomics is limited. Accordingly, the consequences of these distinct EMF communities on plant water uptake remain critically unknown, and future studies targeting the expression of genes involved in drought stress tolerance are required.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Structural basis of differential gene expression at eQTLs loci from high-resolution ensemble models of 3D single-cell chromatin conformations

Abstract Motivation Techniques such as high-throughput chromosome conformation capture (Hi-C) have provided a wealth of information on nucleus organization and genome important for understanding gene expression regulation. Genome-Wide Association Studies have identified numerous loci associated with complex traits. Expression quantitative trait loci (eQTL) studies have further linked the genetic variants to alteration in expression levels of associated target genes across individuals. However, the functional roles of many eQTLs in noncoding regions remain unclear. Current joint analyses of Hi-C and eQTLs data lack advanced computational tools, limiting what can be learned from these data. Results We developed a computational method for simultaneous analysis of Hi-C and eQTL data, capable of identifying a small set of nonrandom interactions from all Hi-C interactions. Using these nonrandom interactions, we reconstructed large ensembles (×105) of high-resolution single-cell 3D chromatin conformations with thorough sampling, accurately replicating Hi-C measurements. Our results revealed many-body interactions in chromatin conformation at the single-cell level within eQTL loci, providing a detailed view of how 3D chromatin structures form the physical foundation for gene regulation, including how genetic variants of eQTLs affect the expression of associated eGenes. Furthermore, our method can deconvolve chromatin heterogeneity and investigate the spatial associations of eQTLs and eGenes at subpopulation level, revealing their regulatory impacts on gene expression. Together, ensemble modeling of thoroughly sampled single-cell chromatin conformations combined with eQTL data, helps decipher how 3D chromatin structures provide the physical basis for gene regulation, expression control, and aid in understanding the overall structure-function relationships of genome organization. Availability and implementation It is available at https://github.com/uic-liang-lab/3DChromFolding-eQTL-Loci.

Du, Lin (ORCID:0009000289869812)↗