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At least 91 records · Page 5
All sourcesNASA NTRSDOE OSTI
DOE OSTI2024

Genotypic analyses of IncHI2 plasmids from enteric bacteria

Incompatibility (Inc) HI2 plasmids are large (typically > 200 kb), transmissible plasmids that encode antimicrobial resistance (AMR), heavy metal resistance (HMR) and disinfectants/biocide resistance (DBR). To better understand the distribution and diversity of resistance-encoding genes among IncHI2 plasmids, computational approaches were used to evaluate resistance and transfer-associated genes among the plasmids. Complete IncHI2 plasmid (N - 667) sequences were extracted from GenBank and analyzed using AMRFinderPlus, IntegronFinder and Plasmid Transfer Factor database. The most common IncHI2-carrying genera included Enterobacter (N = 209), Escherichia (N = 208), and Salmonella (N = 204). Resistance genes distribution was diverse, with plasmids from Escherichia and Salmonella showing general similarity in comparison to Enterobacter and other taxa, which grouped together. Plasmids from Enterobacter and other taxa had a higher prevalence of multiple mercury resistance genes and arsenic resistance gene, arsC, compared to Escherichia and Salmonella. For sulfonamide resistance, sul1 was more common among Enterobacter and other taxa, compared to sul2 and sul3 for Escherichia and Salmonella. Similar gene diversity trends were also observed for tetracyclines, quinolones, β-lactams, and colistin. Over 99% of plasmids carried at least 25 IncHI2-associated conjugal transfer genes. These findings highlight the diversity and dissemination potential for resistance across different enteric bacteria and value of computational-based approaches for the resistance-gene assessment.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Topologically associating domain boundaries are required for normal genome function

Topologically associating domain (TAD) boundaries partition the genome into distinct regulatory territories. Anecdotal evidence suggests that their disruption may interfere with normal gene expression and cause disease phenotypes1,2,3, but the overall extent to which this occurs remains unknown. Here we demonstrate that targeted deletions of TAD boundaries cause a range of disruptions to normal in vivo genome function and organismal development. We used CRISPR genome editing in mice to individually delete eight TAD boundaries (11–80 kb in size) from the genome. All deletions examined resulted in detectable molecular or organismal phenotypes, which included altered chromatin interactions or gene expression, reduced viability, and anatomical phenotypes. We observed changes in local 3D chromatin architecture in 7 of 8 (88%) cases, including the merging of TADs and altered contact frequencies within TADs adjacent to the deleted boundary. For 5 of 8 (63%) loci examined, boundary deletions were associated with increased embryonic lethality or other developmental phenotypes. For example, a TAD boundary deletion near Smad3/Smad6 caused complete embryonic lethality, while a deletion near Tbx5/Lhx5 resulted in a severe lung malformation. Our findings demonstrate the importance of TAD boundary sequences for in vivo genome function and reinforce the critical need to carefully consider the potential pathogenicity of noncoding deletions affecting TAD boundaries in clinical genetics screening.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Atomistic origin of the entropy of melting from inelastic neutron scattering and machine learned molecular dynamics

The latent heat, L, is central to melting, but its atomic origin remains elusive. It is proportional to the entropy of fusion, ΔS fus = L/T m (T m is the melting temperature), which depends on changes of atom configurations, atom vibrations, and thermal electron excitations. Here, we combine inelastic neutron scattering and machine-learned molecular dynamics to separate ΔS fus into these components for Ge, Si, Bi, Sn, Pb, and Li. When the vibrational entropy of melting, ΔS vib , is zero, ΔS fus ≃ 1.2 kB per atom. This result provides a baseline for ΔSconfig and nearly coincides with “Richard’s Rule” of melting. The ΔSfus deviates from this value for most elements, however, and we show that this deviation originates with extra ΔS vib and extra ΔS config . These two components are correlated for positive and negative deviations from Richard’s rule – the extra ΔS config is consistently ~ 80% of ΔS vib . Our results, interpreted with potential energy landscape theory, imply a correlation between the change in the number of basins and the change in the inverse of their curvature for the melting of pure elements.

36 MATERIALS SCIENCE↗
DOE OSTI2023

Hydrophobic pockets built in polymer micelles enhance the reactivity of Cu 2+ ions

We report the hydrophobicity-enhanced reactivity of Cu2+ ions as an ester hydrolase. Using a dipicolylamine (DPA) containing reversible addition–fragmentation chain transfer agent, the synthetic sequence, either hydrophobic or hydrophilic first in amphiphilic block copolymers of polystyrene-block-poly(N'N-dimethylacrylamide) (PS-b-PDMA), was varied to control the location of the binding motif, DPA, in the hydrophobic core or on the hydrated corona of polymer micelles. The hydrophobicity of Cu 2+ sites showed a significant impact (as large as 60 times more activity) on their catalytic efficiency towards ester hydrolase. With two different kinetic modes, including Michaelis–Menten and the reverse saturation kinetics models, the binding constant Kb of the substrates to Cu 2+ sites were quantitatively analyzed and we demonstrate that hydrophobicity favors the binding of the substrates to Cu2+ sites at polymer micelles with smaller sizes, however, K b decays exponentially with micellar diameters. Despite the diffusion barrier, hydrophobicity shows a profound impact on the catalytic rate constant kc that measures the single conversion rate of bound substrates to products. There is a 16–20 times kinetic enhancement in the hydrolase activity, completely endowed by the hydrophobic microenvironment of Cu 2+ sites compared to micelles with similar sizes. Our results indicate how the hydrophobicity of Cu 2+ -containing micelles can impact the catalytic efficiency and potentially illustrate a promising way toward the design of bioinspired catalysts.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2024

Scintillation characteristics of the EJ-299-02H scintillator

Here a study of the dead layer thickness and quenching factor of a plastic scintillator for use in ultracold neutron (UCN) experiments is described. Alpha spectroscopy was used to determine the thickness of a thin surface dead layer to be 630 ± 110 nm. The relative light outputs from the decay of 241 Am and Compton scattering of electrons were used to extract Birks’ law coefficient, yielding a kB value of 0.087 ± 0.003 mm/MeV, consistent with some previous reports for other polystyrene-based scintillators. The results from these measurements are incorporated into the simulation to show that an energy threshold of (∼9 keV) can be achieved for the UCNProBe experiment. This low threshold enables high beta particle detection efficiency and the indirect measurement of UCN. The ability to make the scintillator deuterated, accompanied by its relatively thin dead layer, gives rise to unique applications in a wide range of UCN experiments, where it can be used to trap UCN and detect charged particles in situ.

73 NUCLEAR PHYSICS AND RADIATION PHYSICS↗
DOE OSTI2021

Transmission of Images on High-Temperature Nuclear-Grade Metallic Pipe with Ultrasonic Elastic Waves

Transmission of information using elastic ultrasonic waves on existing metallic pipes provides an alternative communication option for a nuclear facility. The advantages of this approach consist of transmitting information through barriers, such as the containment building wall, with minimal modification of the existing hardware. Because bit rates on the order of kilobits per second are achievable, relatively large volumes of data, such as images, can be transmitted. A viable candidate for an ultrasonic communication channel is a stainless steel pipe of the chemical volume control system (CVCS) that penetrates through the reactor containment building wall through a sealed tunnel. To study ultrasonic communication under simulated nuclear facility conditions of high temperature, a test article was developed by installing heating tapes, temperature controllers, and thermal insulation on a laboratory CVCS-like stainless steel pipe. High temperature and radiation-resilient lithium niobate ultrasonic transducers were utilized for information transmission on the heated pipe. The amplitude shift keying (ASK) digital communication protocol was developed and implemented in a GNU Radio software-defined radio environment. A root-raised-cosine filter was introduced to suppress ultrasonic transducer ringing and thus reduce inter-symbol interference. This resulted in the enhancement of the data transmission bit rate compared to information encoding with square pulses. Demonstrations of communication at high temperature included transmission of a 90-KB image at the bit rate of 10 Kbps with a bit error rate of 10 -3 across a 6-ft-long straight pipe heated up to 230 degrees C. Additional preliminary studies were conducted to evaluate ultrasonic communication system resilience to environmental degradation and damage.

42 ENGINEERING↗
DOE OSTI2023

Multiplex Editing of the Nucleoredoxin1 Tandem Array in Poplar: From Small Indels to Translocations and Complex Inversions

The CRISPR-Cas9 system has been deployed for precision mutagenesis in an ever-growing number of species, including agricultural crops and forest trees. Its application to closely linked genes with extremely high sequence similarities has been less explored. In this study, we used CRISPR-Cas9 to mutagenize a tandem array of seven Nucleoredoxin1 (NRX1) genes spanning ~100 kb in Populus tremula × Populus alba. We demonstrated efficient multiplex editing with one single guide RNA in 42 transgenic lines. The mutation profiles ranged from small insertions and deletions and local deletions in individual genes to large genomic dropouts and rearrangements spanning tandem genes. We also detected complex rearrangements including translocations and inversions resulting from multiple cleavage and repair events. Target capture sequencing was instrumental for unbiased assessments of repair outcomes to reconstruct unusual mutant alleles. The work highlights the power of CRISPR-Cas9 for multiplex editing of tandemly duplicated genes to generate diverse mutants with structural and copy number variations to aid future functional characterization.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Divergent selection and climate adaptation fuel genomic differentiation between sister species of Sphagnum (peat moss)

Abstract Background and Aims New plant species can evolve through the reinforcement of reproductive isolation via local adaptation along habitat gradients. Peat mosses (Sphagnaceae) are an emerging model system for the study of evolutionary genomics and have well-documented niche differentiation among species. Recent molecular studies have demonstrated that the globally distributed species Sphagnum magellanicum is a complex of morphologically cryptic lineages that are phylogenetically and ecologically distinct. Here, we describe the architecture of genomic differentiation between two sister species in this complex known from eastern North America: the northern S. diabolicum and the largely southern S. magniae. Methods We sampled plant populations from across a latitudinal gradient in eastern North America and performed whole genome and restriction-site associated DNA sequencing. These sequencing data were then analyzed computationally. Key Results Using sliding-window population genetic analyses we find that differentiation is concentrated within ‘islands’ of the genome spanning up to 400 kb that are characterized by elevated genetic divergence, suppressed recombination, reduced nucleotide diversity and increased rates of non-synonymous substitution. Sequence variants that are significantly associated with genetic structure and bioclimatic variables occur within genes that have functional enrichment for biological processes including abiotic stress response, photoperiodism and hormone-mediated signalling. Demographic modelling demonstrates that these two species diverged no more than 225 000 generations ago with secondary contact occurring where their ranges overlap. Conclusions We suggest that this heterogeneity of genomic differentiation is a result of linked selection and reflects the role of local adaptation to contrasting climatic zones in driving speciation. This research provides insight into the process of speciation in a group of ecologically important plants and strengthens our predictive understanding of how plant populations will respond as Earth’s climate rapidly changes.

58 GEOSCIENCES↗
DOE OSTI2021

efam: an e xpanded, metaproteome-supported HMM profile database of viral protein fam ilies

Viruses infect, reprogram and kill microbes, leading to profound ecosystem consequences, from elemental cycling in oceans and soils to microbiome-modulated diseases in plants and animals. Although metagenomic datasets are increasingly available, identifying viruses in them is challenging due to poor representation and annotation of viral sequences in databases. Here, we establish efam, an expanded collection of Hidden Markov Model (HMM) profiles that represent viral protein families conservatively identified from the Global Ocean Virome 2.0 dataset. This resulted in 240 311 HMM profiles, each with at least 2 protein sequences, making efam >7-fold larger than the next largest, pan-ecosystem viral HMM profile database. Adjusting the criteria for viral contig confidence from ‘conservative’ to ‘eXtremely Conservative’ resulted in 37 841 HMM profiles in our efam-XC database. To assess the value of this resource, we integrated efam-XC into VirSorter viral discovery software to discover viruses from less-studied, ecologically distinct oxygen minimum zone (OMZ) marine habitats. This expanded database led to an increase in viruses recovered from every tested OMZ virome by ~24% on average (up to ~42%) and especially improved the recovery of often-missed shorter contigs (<5 kb). Additionally, to help elucidate lesser-known viral protein functions, we annotated the profiles using multiple databases from the DRAM pipeline and virion-associated metaproteomic data, which doubled the number of annotations obtainable by standard, single-database annotation approaches. Together, these marine resources (efam and efam-XC) are provided as searchable, compressed HMM databases that will be updated bi-annually to help maximize viral sequence discovery and study from any ecosystem.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2021

CUT&RUN identifies centromeric DNA regions of Rhodotorula toruloides IFO0880

ABSTRACT Rhodotorula toruloides has been increasingly explored as a host for bioproduction of lipids, fatty acid derivatives and terpenoids. Various genetic tools have been developed, but neither a centromere nor an autonomously replicating sequence (ARS), both necessary elements for stable episomal plasmid maintenance, has yet been reported. In this study, cleavage under targets and release using nuclease (CUT&RUN), a method used for genome-wide mapping of DNA–protein interactions, was used to identify R. toruloides IFO0880 genomic regions associated with the centromeric histone H3 protein Cse4, a marker of centromeric DNA. Fifteen putative centromeres ranging from 8 to 19 kb in length were identified and analyzed, and four were tested for, but did not show, ARS activity. These centromeric sequences contained below average GC content, corresponded to transcriptional cold spots, were primarily nonrepetitive and shared some vestigial transposon-related sequences but otherwise did not show significant sequence conservation. Future efforts to identify an ARS in this yeast can utilize these centromeric DNA sequences to improve the stability of episomal plasmids derived from putative ARS elements.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

MINE: maximally informative next experiment—toward a new GWAS experimental design and methodology

Abstract The computational methodology of Genome Wide Association Studies (GWAS) currently has several limitations: (i) the number of observations (rows) on a quantitative trait tends to be smaller than the number of single nucleotide polymorphisms (SNPs) (columns) in the design matrix; (ii) each SNP is usually modeled separately, failing to acknowledge interaction between each other (ie epistasis); (iii) there is implicit linkage disequilibrium (LD) between neighboring SNPs due to their linkage. To overcome these issues, we developed a tool that uses ensemble methods to fit mixed linear models to GWAS data, and these ensemble methods include the development of a new experimental design approach in GWAS, which uses the resultant models and data to select the next informative experiment over time. This new adaptive and staged approach for GWAS experimental design was developed and tested in a 3 yr adaptive model-guided discovery experiment against a fixed classical design. In Sorghum bicolor a total of 79, 86, and 78 accessions were tested in years 1, 2, and 3, respectively out of 343 accessions available in the Bioenergy Association Panel (BAP) each identified for 232,303 SNPs, 1 every 2–3 kb in the genomes. We demonstrated the feasibility of MINE enacted with 8 people in the field per year over 3 yr vs in 1 large classical design enacted with 20 people in 1 yr. The MINE results for chromosomal regions identified controlling dry weight were confirmed against results from previous sorghum GWAS experiments and 1 large classical design for the BAP panel.

Genetics & Heredity↗
DOE OSTI2025

Viroid-like “obelisk” agents are widespread in the ocean and exceed the abundance of RNA viruses in the prokaryotic fraction

Abstract “Obelisks” are recently discovered ribonucleic acid (RNA) viroid-like elements present in diverse environments with no phylogenetic similarity to any known biological agent. obelisks were first identified in the human gut and in a commensal bacterium acting as a replicative host. They have a circular ∼1 kb RNA genome, rod-like secondary structures, and the encoding of a protein superfamily called “Oblins”. We performed a large-scale search of obelisks in the ocean using the Pebblescout program and the transcriptomic Sequence Archive Read databases, revealing the biogeography and abundance of these viroid-like RNA elements. We detected 55 obelisk genomes resulting in 35 marine clusters at the species level. These obelisks were detected in the prokaryotic fraction and to a lesser extent in the eukaryotic fraction, and distributed across all the oceans from surface to mesopelagic including the Arctic, and even in the coldest seawater of Earth beneath the Antarctic Ross Ice Shelf. The obelisk hallmark protein Oblin-1 confirmed by 3D models was found in various marine samples. Some of the detected marine obelisks harbor hammerhead self-cleaving ribozymes in both polarities. In the prokaryotic, but not the eukaryotic, fraction of the Tara Ocean dataset, relative abundance of obelisks calculated by transcriptomic fragment recruitment indicated that they are abundant in marine samples, reaching or even exceeding the relative abundance of the previously discovered uncultured RNA viruses. In conclusion, obelisks are abundant and widespread viroid-like elements that should be included in ocean biogeochemical models.

Environmental Sciences & Ecology↗
DOE OSTI2020

Linezolid-resistant (Tn 6246 :: fexB - poxtA ) Enterococcus faecium strains colonizing humans and bovines on different continents: similarity without epidemiological link

Abstract Objectives poxtA is the most recently described gene conferring acquired resistance to linezolid, a relevant antibiotic for treating enterococcal infections. We retrospectively screened for poxtA in diverse enterococci and aimed to characterize its genetic/genomic contexts. Methods poxtA was screened by PCR in 812 enterococci from 458 samples (hospitals/healthy humans/wastewater/animals/retail food) obtained in Portugal/Angola/Tunisia (1996–2019). Antimicrobial susceptibility testing was performed for 13 antibiotics (EUCAST/CLSI). poxtA stability (∼500 generations), transfer (filter mating), clonality (SmaI-PFGE) and location (S1-PFGE/hybridization) were tested. WGS (Illumina-HiSeq) was performed for clonal representatives. Results poxtA was detected in Enterococcus faecium from six samples (1.3%): a healthy human (rectal swab) in Porto, Portugal (ST32/2001); four farm cows (milk) in Mateur, Tunisia (ST1058/2015); and a hospitalized patient (faeces) in Matosinhos, Portugal (ST1058/2015). All expressed resistance to linezolid (MIC = 8 mg/L), chloramphenicol, tetracycline and erythromycin, with variable resistance to ciprofloxacin and streptomycin. ST1058-poxtA-carrying isolates from Tunisia and Portugal differed by two SNPs and had similar plasmid content. poxtA, located in an IS1216-flanked Tn6246-like element, co-hybridized with fexB on one or more plasmids per isolate (one to three plasmids of 30–100 kb), was stable after several generations and transferred only from ST1058. ST1058 strains carried resistance/virulence genes (Efmqnr/acm) possibly induced under selective quinolone treatment. Conclusions poxtA has been circulating in Portugal since at least 2001, corresponding to the oldest description worldwide to date. We also extend the reservoir of poxtA to bovines. The similar linezolid-resistant poxtA-carrying strains colonizing humans and livestock on different continents, and without a noticeable relationship, suggests a recent transmission event or convergent evolution of E. faecium populations in different hosts and geographic regions.

Freitas, Ana R.↗
DOE OSTI2025

The reference genome for the northeastern Pacific bull kelp, Nereocystis luetkeana

Bull kelp, Nereocystis luetkeana, is a northeastern Pacific kelp with broad distribution from Alaska to central California. Its population declines have caused severe concerns in northern California, the Salish Sea in Washington, and recently in some populations in Oregon. Despite bull kelp's accumulated ecological and physiological studies, an assembled and annotated genomic reference was still unavailable. Here, we report the complete and annotated genome of Nereocystis luetkeana, produced by the California Conservation Genomics Project (CCGP), which aims to reveal genomic diversity patterns across California by sequencing the complete genomes of approximately 150 carefully selected species. The genome was assembled into 1562 scaffolds with 449.82 Mb, 80x of coverage and 22 952 gene models. BUSCO assembly showed a completeness score of 72% for the stramenopiles gene set. The mitochondria and chloroplast genome sequences have 37 Kb and 131 Mb, respectively. The orthology analysis between 10 Phaeophycean genomes showed 1065 expanded and 286 unique orthogroups for this species. Pairwise comparisons showed 542 orthogroups present only in N. luetkeana and M. pyrifera, another large-body kelp. The enrichment analysis of these orthogroups showed important functions related to central metabolism and signaling due to ATPases enrichment in these two species. This genome assembly will provide an essential resource for the ecology, evolution, conservation, and breeding of bull kelp.

California Conservation Genomics Project—CCGP↗
DOE OSTI2021

Museomics Dissects the Genetic Basis for Adaptive Seasonal Coloration in the Least Weasel

Abstract Dissecting the link between genetic variation and adaptive phenotypes provides outstanding opportunities to understand fundamental evolutionary processes. Here, we use a museomics approach to investigate the genetic basis and evolution of winter coat coloration morphs in least weasels (Mustela nivalis), a repeated adaptation for camouflage in mammals with seasonal pelage color moults across regions with varying winter snow. Whole-genome sequence data were obtained from biological collections and mapped onto a newly assembled reference genome for the species. Sampling represented two replicate transition zones between nivalis and vulgaris coloration morphs in Europe, which typically develop white or brown winter coats, respectively. Population analyses showed that the morph distribution across transition zones is not a by-product of historical structure. Association scans linked a 200-kb genomic region to coloration morph, which was validated by genotyping museum specimens from intermorph experimental crosses. Genotyping the wild populations narrowed down the association to pigmentation gene MC1R and pinpointed a candidate amino acid change cosegregating with coloration morph. This polymorphism replaces an ancestral leucine residue by lysine at the start of the first extracellular loop of the protein in the vulgaris morph. A selective sweep signature overlapped the association region in vulgaris, suggesting that past adaptation favored winter-brown morphs and can anchor future adaptive responses to decreasing winter snow. Using biological collections as valuable resources to study natural adaptations, our study showed a new evolutionary route generating winter color variation in mammals and that seasonal camouflage can be modulated by changes at single key genes.

Miranda, Inês↗
DOE OSTI2021

Deeplasmid: deep learning accurately separates plasmids from bacterial chromosomes

Plasmids are mobile genetic elements that play a key role in microbial ecology and evolution by mediating horizontal transfer of important genes, such as antimicrobial resistance genes. Many microbial genomes have been sequenced by short read sequencers and have resulted in a mix of contigs that derive from plasmids or chromosomes. New tools that accurately identify plasmids are needed to elucidate new plasmid-borne genes of high biological importance. We have developed Deeplasmid, a deep learning tool for distinguishing plasmids from bacterial chromosomes based on the DNA sequence and its encoded biological data. It requires as input only assembled sequences generated by any sequencing platform and assembly algorithm and its runtime scales linearly with the number of assembled sequences. Deeplasmid achieves an AUC–ROC of over 89%, and it was more accurate than five other plasmid classification methods. Finally, as a proof of concept, we used Deeplasmid to predict new plasmids in the fish pathogen Yersinia ruckeri ATCC 29473 that has no annotated plasmids. Deeplasmid predicted with high reliability that a long assembled contig is part of a plasmid. Using long read sequencing we indeed validated the existence of a 102 kb long plasmid, demonstrating Deeplasmid's ability to detect novel plasmids.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

Comparative mitogenomics of kingdom Fungi – evolutionary insights and metagenomic applications

Mitochondria are essential components of eukaryotic cells, responsible for ATP production through oxidative phosphorylation. Despite their biological importance, unique challenges have hindered the adoption of automated mitochondrial genome (mitogenome) annotation methods, obstructing mitochondrial comparative genomics in a broad evolutionary context. Using Fungi as a study system and a Joint Genome Institute (JGI) annotated high-quality reference set, we observed broad patterns of mitochondrial evolution across the kingdom. We found that the median fungal mitogenome size is 58 kb and identified exceptionally large examples over 1 Mb in Pezizomycetes. All 14 expected oxidative phosphorylation protein-coding genes, plus rps3, were generally conserved. We found evidence of major evolutionary transitions within the Ascomycota, including the transfer of mitochondrially encoded atp8 and atp9 to the nuclear genomes across the Pezizomycotina and shifts in mitogenome tRNA patterns across the kingdom. We found substantial concordance between mitochondrial and nuclear evolution, enabling us to document 3131 total fungal mitogenomes from JGI-derived metagenomic datasets. We also identified 6467 total undeclared mitogenomes embedded in Genbank fungal nuclear assemblies. We provide interactive tools for mitogenome analysis through the JGI MycoCosm platform. Collectively, this work generated nearly 10 000 new fungal mitogenome annotations, providing a foundation and resources for future exploration of comparative fungal mitogenomics.

Ahrendt, Steven R. [USDOE Joint Genome Institute (↗
DOE OSTI2021

Topologically Associating Domain Boundaries are Commonly Required for Normal Genome Function

Topologically associating domain (TAD) boundaries are thought to partition the genome into distinct regulatory territories. Anecdotal evidence suggests that their disruption may interfere with normal gene expression and cause disease phenotype, but the overall extent to which this occurs remains unknown. Here we show that TAD boundary deletions commonly disrupt normal genome function in vivo . We used CRISPR genome editing in mice to individually delete eight TAD boundaries (11-80kb in size) from the genome in mice. All deletions examined resulted in at least one detectable molecular or organismal phenotype, which included altered chromatin interactions or gene expression, reduced viability, and anatomical phenotypes. For 5 of 8 (62%) loci examined, boundary deletions were associated with increased embryonic lethality or other developmental phenotypes. For example, a TAD boundary deletion near Smad3/Smad6 caused complete embryonic lethality, while a deletion near Tbx5/Lhx5 resulted in a severe lung malformation. Our findings demonstrate the importance of TAD boundary sequences for in vivo genome function and suggest that noncoding deletions affecting TAD boundaries should be carefully considered for potential pathogenicity in clinical genetics screening.

Rajderkar, Sudha↗