Artem Mikelov, George Nefediev, Alexander Tashkeev, Oscar L Rodriguez, Diego Aguilar Ortmans, Valeriia Skatova, Mark Izraelson, Alexey N Davydov, Stanislav Poslavsky, Souad Rahmouni, Corey T Watson, Dmitriy Chudakov, Scott D Boyd, Dmitry Bolotin
Allelic variability in the adaptive immune receptor loci, which harbor the gene segments that encode B cell and T cell receptors (BCR/TCR), is of critical importance for immune responses to pathogens and vaccines. Adaptive immune receptor repertoire sequencing (AIRR-seq) has become widespread in immunology research making it the most readily available source of information about allelic diversity in immunoglobulin (IG) and T cell receptor (TR) loci. Here, we present a novel algorithm for extrasensitive and specific variable (V) and joining (J) gene allele inference, allowing the reconstruction of individual high-quality gene segment libraries. The approach can be applied for inferring allelic variants from peripheral blood lymphocyte BCR and TCR repertoire sequencing data, including hypermutated isotype-switched BCR sequences, thus allowing high-throughput novel allele discovery from a wide variety of existing data sets. The developed algorithm is a part of the MiXCR software. We demonstrate the accuracy of this approach using AIRR-seq paired with long-read genomic sequencing data, comparing it to a widely used algorithm, TIgGER. We applied the algorithm to a large set of IG heavy chain (IGH) AIRR-seq data from 450 donors of ancestrally diverse population groups, and to the largest reported full-length TCR alpha and beta chain (TRA and TRB) AIRR-seq data set, representing 134 individuals. This allowed us to assess the genetic diversity within the IGH, TRA, and TRB loci in different populations and to establish a database of alleles of V and J genes inferred from AIRR-seq data and their population frequencies with free public access through VDJ.online database.
{"title":"Ultrasensitive allele inference from immune repertoire sequencing data with MiXCR.","authors":"Artem Mikelov, George Nefediev, Alexander Tashkeev, Oscar L Rodriguez, Diego Aguilar Ortmans, Valeriia Skatova, Mark Izraelson, Alexey N Davydov, Stanislav Poslavsky, Souad Rahmouni, Corey T Watson, Dmitriy Chudakov, Scott D Boyd, Dmitry Bolotin","doi":"10.1101/gr.278775.123","DOIUrl":"10.1101/gr.278775.123","url":null,"abstract":"<p><p>Allelic variability in the adaptive immune receptor loci, which harbor the gene segments that encode B cell and T cell receptors (BCR/TCR), is of critical importance for immune responses to pathogens and vaccines. Adaptive immune receptor repertoire sequencing (AIRR-seq) has become widespread in immunology research making it the most readily available source of information about allelic diversity in immunoglobulin (IG) and T cell receptor (TR) loci. Here, we present a novel algorithm for extrasensitive and specific variable (V) and joining (J) gene allele inference, allowing the reconstruction of individual high-quality gene segment libraries. The approach can be applied for inferring allelic variants from peripheral blood lymphocyte BCR and TCR repertoire sequencing data, including hypermutated isotype-switched BCR sequences, thus allowing high-throughput novel allele discovery from a wide variety of existing data sets. The developed algorithm is a part of the MiXCR software. We demonstrate the accuracy of this approach using AIRR-seq paired with long-read genomic sequencing data, comparing it to a widely used algorithm, TIgGER. We applied the algorithm to a large set of IG heavy chain (<i>IGH</i>) AIRR-seq data from 450 donors of ancestrally diverse population groups, and to the largest reported full-length TCR alpha and beta chain (<i>TRA</i> and <i>TRB</i>) AIRR-seq data set, representing 134 individuals. This allowed us to assess the genetic diversity within the <i>IGH</i>, <i>TRA</i>, and <i>TRB</i> loci in different populations and to establish a database of alleles of V and J genes inferred from AIRR-seq data and their population frequencies with free public access through VDJ.online database.</p>","PeriodicalId":12678,"journal":{"name":"Genome research","volume":" ","pages":"2293-2303"},"PeriodicalIF":6.2,"publicationDate":"2024-12-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11694755/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142463177","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Paired-class homeodomain transcription factors (HD TFs) play essential roles in vertebrate development, and their mutations are linked to human diseases. One unique feature of paired-class HD is cooperative dimerization on specific palindrome DNA sequences. Yet, the functional significance of HD cooperative dimerization in animal development and its dysregulation in diseases remain elusive. Using the retinal TF Cone-rod Homeobox (CRX) as a model, we have studied how blindness-causing mutations in the paired HD, p.E80A and p.K88N, alter CRX’s cooperative dimerization, lead to gene misexpression and photoreceptor developmental deficits in dominant manners. CRXE80A maintains binding at monomeric WT CRX motifs but is deficient in cooperative binding at dimeric motifs. CRXE80A’s cooperativity defect impacts the exponential increase of photoreceptor gene expression in terminal differentiation and produces immature, non-functional photoreceptors in the CrxE80A retinas. CRXK88N is highly cooperative and localizes to ectopic genomic sites with strong enrichment of dimeric HD motifs. CRXK88N’s altered biochemical properties disrupt CRX’s ability to direct dynamic chromatin remodeling during development to activate photoreceptor differentiation programs and silence progenitor programs. Our study here provides in vitro and in vivo molecular evidence that paired-class HD cooperative dimerization regulates neuronal development and dysregulation of cooperative binding contributes to severe dominant blinding retinopathies.
{"title":"Aberrant homeodomain-DNA cooperative dimerization underlies distinct developmental defects in two dominant CRX retinopathy models","authors":"Yiqiao Zheng, Gary D. Stormo, Shiming Chen","doi":"10.1101/gr.279340.124","DOIUrl":"https://doi.org/10.1101/gr.279340.124","url":null,"abstract":"Paired-class homeodomain transcription factors (HD TFs) play essential roles in vertebrate development, and their mutations are linked to human diseases. One unique feature of paired-class HD is cooperative dimerization on specific palindrome DNA sequences. Yet, the functional significance of HD cooperative dimerization in animal development and its dysregulation in diseases remain elusive. Using the retinal TF Cone-rod Homeobox (CRX) as a model, we have studied how blindness-causing mutations in the paired HD, p.E80A and p.K88N, alter CRX’s cooperative dimerization, lead to gene misexpression and photoreceptor developmental deficits in dominant manners. CRX<sup>E80A</sup> maintains binding at monomeric WT CRX motifs but is deficient in cooperative binding at dimeric motifs. CRX<sup>E80A</sup>’s cooperativity defect impacts the exponential increase of photoreceptor gene expression in terminal differentiation and produces immature, non-functional photoreceptors in the <em>Crx<sup>E80A</sup></em> retinas. CRX<sup>K88N</sup> is highly cooperative and localizes to ectopic genomic sites with strong enrichment of dimeric HD motifs. CRX<sup>K88N</sup>’s altered biochemical properties disrupt CRX’s ability to direct dynamic chromatin remodeling during development to activate photoreceptor differentiation programs and silence progenitor programs. Our study here provides <em>in vitro</em> and <em>in vivo</em> molecular evidence that paired-class HD cooperative dimerization regulates neuronal development and dysregulation of cooperative binding contributes to severe dominant blinding retinopathies.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"13 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2024-12-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142879927","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Song Zhang, Chao Wang, Shenghua Qin, Choulin Chen, Yongzhou Bao, Yuanyuan Zhang, Lingna Xu, Qingyou Liu, Yunxiang Zhao, Kui Li, Zhonglin Tang, Yuwen Liu
Super-enhancers (SEs) govern the expression of genes defining cell identity. However, the dynamic landscape of SEs and their critical constituent enhancers involved in skeletal muscle development remains unclear. In this study, using pig as a model, we employed cleavage under targets and tagmentation (CUT&Tag) to profile the enhancer-associated histone modification marker H3K27ac in skeletal muscle across two prenatal and three postnatal stages, and investigated how SEs influence skeletal muscle development. We identify three SE families with distinct temporal dynamics: continuous (Con, 397), transient (TS, 434), and de novo (DN, 756). These SE families are associated with different temporal gene expression trajectories, biological functions, and DNA methylation levels. Notably, several lines of evidence suggest a potential prominent role of Con SEs in regulating porcine muscle development and meat traits. To pinpoint key cis-regulatory units in Con SEs, we developed an integrative approach that leverages information from eRNA annotation, genome-wide association study (GWAS) signals, and high-throughput capture self-transcribing active regulatory region sequencing (STARR-seq) experiments. Within Con SEs, we identify 20 candidate critical enhancers with meat and carcass-associated DNA variations that affect enhancer activity, and infer their upstream transcription factors and downstream target genes. As a proof of concept, we experimentally validate the role of one such enhancer and its potential target gene during myogenesis. Our findings reveal the dynamic regulatory features of SEs in skeletal muscle development and provide a general integrative framework for identifying critical enhancers underlying the formation of complex traits.
超级增强子（SE）控制着决定细胞特性的基因的表达。然而，参与骨骼肌发育的超级增强子及其关键组成增强子的动态图谱仍不清楚。在这项研究中，我们以猪为模型，利用 CUT&Tag 分析了骨骼肌中与增强子相关的组蛋白修饰标记 H3K27ac 在出生前两个阶段和出生后三个阶段的变化，并研究了增强子如何影响骨骼肌的发育。我们发现了三个具有不同时间动态的 SE 家族：连续 SE（Con，397 个）、瞬时 SE（TS，434 个）和新生 SE（DN，756 个）。这些 SE 家族与不同时间的基因表达轨迹、生物功能和 DNA 甲基化水平相关。值得注意的是，一些证据表明，Con SEs 在调节猪肌肉发育和肉质性状方面可能起着重要作用。为了精确定位 Con SEs 中的关键顺式调控单元，我们开发了一种综合方法，利用来自 eRNA 注释、GWAS 信号和高通量捕获 STARR-seq 实验的信息。在 Con SEs 中，我们发现了 20 个候选关键增强子，它们与肉类和胴体相关的 DNA 变异会影响增强子的活性，并推断出了它们的上游 TF 和下游靶基因。作为概念验证，我们通过实验验证了其中一个增强子及其潜在靶基因在肌形成过程中的作用。我们的研究结果揭示了骨骼肌发育过程中增强子的动态调控特征，并为确定复杂性状形成过程中的关键增强子提供了一个通用的综合框架。
{"title":"Analyzing super-enhancer temporal dynamics reveals potential critical enhancers and their gene regulatory networks underlying skeletal muscle development.","authors":"Song Zhang, Chao Wang, Shenghua Qin, Choulin Chen, Yongzhou Bao, Yuanyuan Zhang, Lingna Xu, Qingyou Liu, Yunxiang Zhao, Kui Li, Zhonglin Tang, Yuwen Liu","doi":"10.1101/gr.278344.123","DOIUrl":"10.1101/gr.278344.123","url":null,"abstract":"<p><p>Super-enhancers (SEs) govern the expression of genes defining cell identity. However, the dynamic landscape of SEs and their critical constituent enhancers involved in skeletal muscle development remains unclear. In this study, using pig as a model, we employed cleavage under targets and tagmentation (CUT&Tag) to profile the enhancer-associated histone modification marker H3K27ac in skeletal muscle across two prenatal and three postnatal stages, and investigated how SEs influence skeletal muscle development. We identify three SE families with distinct temporal dynamics: continuous (Con, 397), transient (TS, 434), and de novo (DN, 756). These SE families are associated with different temporal gene expression trajectories, biological functions, and DNA methylation levels. Notably, several lines of evidence suggest a potential prominent role of Con SEs in regulating porcine muscle development and meat traits. To pinpoint key <i>cis</i>-regulatory units in Con SEs, we developed an integrative approach that leverages information from eRNA annotation, genome-wide association study (GWAS) signals, and high-throughput capture self-transcribing active regulatory region sequencing (STARR-seq) experiments. Within Con SEs, we identify 20 candidate critical enhancers with meat and carcass-associated DNA variations that affect enhancer activity, and infer their upstream transcription factors and downstream target genes. As a proof of concept, we experimentally validate the role of one such enhancer and its potential target gene during myogenesis. Our findings reveal the dynamic regulatory features of SEs in skeletal muscle development and provide a general integrative framework for identifying critical enhancers underlying the formation of complex traits.</p>","PeriodicalId":12678,"journal":{"name":"Genome research","volume":" ","pages":"2190-2202"},"PeriodicalIF":6.2,"publicationDate":"2024-12-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11694746/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142463175","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Alejandro Paniagua, Cristina Agustin-García, Francisco J Pardo-Palacios, Thomas Brown, Maite De Maria, Nancy D Denslow, Camila Mazzoni, Ana Conesa
While the production of a draft genome has become more accessible due to long-read sequencing, the annotation of these new genomes has not been developed at the same pace. Long-read RNA sequencing (lrRNA-seq) offers a promising solution for enhancing gene annotation. In this study, we explore how sequencing platforms, Oxford Nanopore R9.4.1 chemistry or PacBio Sequel II CCS, and data processing methods influence evidence-driven genome annotation using long reads. Incorporating PacBio transcripts into our annotation pipeline significantly outperformed traditional methods, such as ab initio predictions and short-read-based annotations. We applied this strategy to a nonmodel species, the Florida manatee, and compared our results to existing short-read-based annotation. At the loci level, both annotations were highly concordant, with 90% agreement. However, at the transcript level, the agreement was only 35%. We identified 4,906 novel loci, represented by 5,707 isoforms, with 64% of these isoforms matching known sequences in other mammalian species. Overall, our findings underscore the importance of using high-quality curated transcript models in combination with ab initio methods for effective genome annotation.
虽然由于长读测序，草图基因组的制作变得更加容易，但这些新基因组的注释并没有以同样的速度发展。长读RNA测序（lrRNA-seq）为增强基因注释提供了一种很有前途的解决方案。在本研究中，我们探讨了测序平台、Oxford Nanopore R9.4.1化学或PacBio Sequel II CCS以及数据处理方法如何影响使用长读取的证据驱动基因组注释。将PacBio转录本整合到我们的注释管道中显著优于传统方法，例如从头开始预测和基于短读的注释。我们将这种策略应用于非模式物种佛罗里达海牛，并将我们的结果与现有的基于短读的注释进行比较。在位点水平上，两种注释高度一致，一致性达90%。然而，在成绩单水平上，一致性只有35%。我们鉴定了4906个新位点，由5707个同种异构体代表，其中64%的同种异构体与其他哺乳动物物种的已知序列相匹配。总的来说，我们的研究结果强调了将高质量的转录本模型与从头算方法相结合用于有效基因组注释的重要性。
{"title":"Evaluation of strategies for evidence-driven genome annotation using long-read RNA-seq","authors":"Alejandro Paniagua, Cristina Agustin-García, Francisco J Pardo-Palacios, Thomas Brown, Maite De Maria, Nancy D Denslow, Camila Mazzoni, Ana Conesa","doi":"10.1101/gr.279864.124","DOIUrl":"https://doi.org/10.1101/gr.279864.124","url":null,"abstract":"While the production of a draft genome has become more accessible due to long-read sequencing, the annotation of these new genomes has not been developed at the same pace. Long-read RNA sequencing (lrRNA-seq) offers a promising solution for enhancing gene annotation. In this study, we explore how sequencing platforms, Oxford Nanopore R9.4.1 chemistry or PacBio Sequel II CCS, and data processing methods influence evidence-driven genome annotation using long reads. Incorporating PacBio transcripts into our annotation pipeline significantly outperformed traditional methods, such as ab initio predictions and short-read-based annotations. We applied this strategy to a nonmodel species, the Florida manatee, and compared our results to existing short-read-based annotation. At the loci level, both annotations were highly concordant, with 90% agreement. However, at the transcript level, the agreement was only 35%. We identified 4,906 novel loci, represented by 5,707 isoforms, with 64% of these isoforms matching known sequences in other mammalian species. Overall, our findings underscore the importance of using high-quality curated transcript models in combination with ab initio methods for effective genome annotation.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"32 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2024-12-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142879924","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Sotiria Milia, Alexander Leonard, Xena Marie Mapel, Sandra Milena Bernal Ulloa, Cord Drögemüller, Hubert Pausch
Cattle have been selectively bred for coat color, spotting, and depigmentation patterns. The assumed autosomal dominant inherited genetic variants underlying the characteristic white head of Fleckvieh, Simmental, and Hereford cattle have not been identified yet, although the contribution of structural variation upstream the KIT gene has been proposed. Here, we construct a graph pangenome from 24 haplotype assemblies representing seven taurine cattle breeds to identify and characterize the white head-associated locus for the first time based on long-read sequencing data and pangenome analyses. We introduce a pangenome-wide association mapping approach which examines assembly path similarities within the graph to reveal an association between two most likely serial alleles of a complex structural variant 66 kb upstream KIT and facial depigmentation. The complex structural variant contains a variable number of tandemly duplicated 14.3 kb repeats, consisting of LTRs, LINEs, and other repetitive elements, leading to misleading alignments of short and long reads when using a linear reference. We align 250 short-read sequencing samples spanning 15 cattle breeds to the pangenome graph, further validating that the alleles of the structural variant segregate with head depigmentation. We estimate an increased count of repeats in Hereford relative to Simmental and other white-headed cattle breeds from the graph alignment coverage, suggesting a large under-assembly in the current Hereford-based cattle reference genome which had fewer copies. Our work shows that exploiting assembly path similarities within graph pangenomes can reveal trait-associated complex structural variants.
{"title":"Taurine pangenome uncovers a segmental duplication upstream of <i>KIT</i> associated with depigmentation in white-headed cattle.","authors":"Sotiria Milia, Alexander Leonard, Xena Marie Mapel, Sandra Milena Bernal Ulloa, Cord Drögemüller, Hubert Pausch","doi":"10.1101/gr.279064.124","DOIUrl":"10.1101/gr.279064.124","url":null,"abstract":"<p><p>Cattle have been selectively bred for coat color, spotting, and depigmentation patterns. The assumed autosomal dominant inherited genetic variants underlying the characteristic white head of Fleckvieh, Simmental, and Hereford cattle have not been identified yet, although the contribution of structural variation upstream the <i>KIT</i> gene has been proposed. Here, we construct a graph pangenome from 24 haplotype assemblies representing seven taurine cattle breeds to identify and characterize the white head-associated locus for the first time based on long-read sequencing data and pangenome analyses. We introduce a pangenome-wide association mapping approach which examines assembly path similarities within the graph to reveal an association between two most likely serial alleles of a complex structural variant 66 kb upstream <i>KIT</i> and facial depigmentation. The complex structural variant contains a variable number of tandemly duplicated 14.3 kb repeats, consisting of LTRs, LINEs, and other repetitive elements, leading to misleading alignments of short and long reads when using a linear reference. We align 250 short-read sequencing samples spanning 15 cattle breeds to the pangenome graph, further validating that the alleles of the structural variant segregate with head depigmentation. We estimate an increased count of repeats in Hereford relative to Simmental and other white-headed cattle breeds from the graph alignment coverage, suggesting a large under-assembly in the current Hereford-based cattle reference genome which had fewer copies. Our work shows that exploiting assembly path similarities within graph pangenomes can reveal trait-associated complex structural variants.</p>","PeriodicalId":12678,"journal":{"name":"Genome research","volume":" ","pages":""},"PeriodicalIF":6.2,"publicationDate":"2024-12-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142853891","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Simon Orozco-Arias, Pío Sierra, Richard Durbin, Josefa González
The number of species with high-quality genome sequences continues to increase, in part due to the scaling up of multiple large-scale biodiversity sequencing projects. While the need to annotate genic sequences in these genomes is widely acknowledged, the parallel need to annotate transposable element (TE) sequences that have been shown to alter genome architecture, rewire gene regulatory networks, and contribute to the evolution of host traits is becoming ever more evident. However, accurate genome-wide annotation of TE sequences is still technically challenging. Several de novo TE identification tools are now available, but manual curation of the libraries produced by these tools is needed to generate high-quality genome annotations. Manual curation is time-consuming, and thus impractical for large-scale genomic studies, and lacks reproducibility. In this work, we present the Manual Curator Helper tool MCHelper, which automates the TE library curation process. By leveraging MCHelper's fully automated mode with the outputs from three de novo TE identification tools, RepeatModeler2, EDTA, and REPET, in the fruit fly, rice, hooded crow, zebrafish, maize, and human, we show a substantial improvement in the quality of the TE libraries and genome annotations. MCHelper libraries are less redundant, with up to 65% reduction in the number of consensus sequences, have up to 11.4% fewer false positive sequences, and up to ∼48% fewer “unclassified/unknown” TE consensus sequences. Genome-wide TE annotations are also improved, including larger unfragmented insertions. Moreover, MCHelper is an easy-to-install and easy-to-use tool.
{"title":"MCHelper automatically curates transposable element libraries across eukaryotic species","authors":"Simon Orozco-Arias, Pío Sierra, Richard Durbin, Josefa González","doi":"10.1101/gr.278821.123","DOIUrl":"https://doi.org/10.1101/gr.278821.123","url":null,"abstract":"The number of species with high-quality genome sequences continues to increase, in part due to the scaling up of multiple large-scale biodiversity sequencing projects. While the need to annotate genic sequences in these genomes is widely acknowledged, the parallel need to annotate transposable element (TE) sequences that have been shown to alter genome architecture, rewire gene regulatory networks, and contribute to the evolution of host traits is becoming ever more evident. However, accurate genome-wide annotation of TE sequences is still technically challenging. Several de novo TE identification tools are now available, but manual curation of the libraries produced by these tools is needed to generate high-quality genome annotations. Manual curation is time-consuming, and thus impractical for large-scale genomic studies, and lacks reproducibility. In this work, we present the Manual Curator Helper tool MCHelper, which automates the TE library curation process. By leveraging MCHelper's fully automated mode with the outputs from three de novo TE identification tools, RepeatModeler2, EDTA, and REPET, in the fruit fly, rice, hooded crow, zebrafish, maize, and human, we show a substantial improvement in the quality of the TE libraries and genome annotations. MCHelper libraries are less redundant, with up to 65% reduction in the number of consensus sequences, have up to 11.4% fewer false positive sequences, and up to ∼48% fewer “unclassified/unknown” TE consensus sequences. Genome-wide TE annotations are also improved, including larger unfragmented insertions. Moreover, MCHelper is an easy-to-install and easy-to-use tool.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"20 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2024-12-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142797141","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Stephanie C. Bohaczuk, Zachary J. Amador, Chang Li, Benjamin J. Mallory, Elliott G. Swanson, Jane Ranchalis, Mitchell R. Vollger, Katherine M. Munson, Tom Walsh, Morgan O. Hamm, Yizi Mao, Andre Lieber, Andrew B. Stergachis
Accurately quantifying the functional consequences of noncoding mosaic variants requires the pairing of DNA sequences with both accessible and closed chromatin architectures along individual DNA molecules—a pairing that cannot be achieved using traditional fragmentation-based chromatin assays. We demonstrate that targeted single-molecule chromatin fiber sequencing (Fiber-seq) achieves this, permitting single-molecule, long-read genomic, and epigenomic profiling across targeted >100 kb loci with ∼10-fold enrichment over untargeted sequencing. Targeted Fiber-seq reveals that pathogenic expansions of the DMPK CTG repeat that underlie Myotonic Dystrophy 1 are characterized by somatic instability and disruption of multiple nearby regulatory elements, both of which are repeat length-dependent. Furthermore, we reveal that therapeutic adenine base editing of the segmentally duplicated γ-globin (HBG1/HBG2) promoters in primary human hematopoietic cells induced toward an erythroblast lineage increases the accessibility of the HBG1 promoter as well as neighboring regulatory elements. Overall, we find that these non–protein coding mosaic variants can have complex impacts on chromatin architectures, including extending beyond the regulatory element harboring the variant.
{"title":"Resolving the chromatin impact of mosaic variants with targeted Fiber-seq","authors":"Stephanie C. Bohaczuk, Zachary J. Amador, Chang Li, Benjamin J. Mallory, Elliott G. Swanson, Jane Ranchalis, Mitchell R. Vollger, Katherine M. Munson, Tom Walsh, Morgan O. Hamm, Yizi Mao, Andre Lieber, Andrew B. Stergachis","doi":"10.1101/gr.279747.124","DOIUrl":"https://doi.org/10.1101/gr.279747.124","url":null,"abstract":"Accurately quantifying the functional consequences of noncoding mosaic variants requires the pairing of DNA sequences with both accessible and closed chromatin architectures along individual DNA molecules—a pairing that cannot be achieved using traditional fragmentation-based chromatin assays. We demonstrate that targeted single-molecule chromatin fiber sequencing (Fiber-seq) achieves this, permitting single-molecule, long-read genomic, and epigenomic profiling across targeted >100 kb loci with ∼10-fold enrichment over untargeted sequencing. Targeted Fiber-seq reveals that pathogenic expansions of the <em>DMPK</em> CTG repeat that underlie Myotonic Dystrophy 1 are characterized by somatic instability and disruption of multiple nearby regulatory elements, both of which are repeat length-dependent. Furthermore, we reveal that therapeutic adenine base editing of the segmentally duplicated γ-globin (<em>HBG1</em>/<em>HBG2</em>) promoters in primary human hematopoietic cells induced toward an erythroblast lineage increases the accessibility of the <em>HBG1</em> promoter as well as neighboring regulatory elements. Overall, we find that these non–protein coding mosaic variants can have complex impacts on chromatin architectures, including extending beyond the regulatory element harboring the variant.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"1 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2024-12-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142797142","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Vanessa L. Porter, Michelle Ng, Kieran O'Neill, Signe MacLennan, Richard D. Corbett, Luka Culibrk, Zeid Hamadeh, Marissa Iden, Rachel Schmidt, Shirng-Wern Tsaih, Carolyn Nakisige, Martin Origa, Jackson Orem, Glenn Chang, Jeremy Fan, Ka Ming Nip, Vahid Akbari, Simon K. Chan, James Hopkins, Richard A. Moore, Eric Chuah, Karen L. Mungall, Andrew J. Mungall, Inanc Birol, Steven J.M. Jones, Janet S. Rader, Marco A. Marra
Human papillomavirus (HPV) integration has been implicated in transforming HPV infection into cancer. To resolve genome dysregulation associated with HPV integration, we performed Oxford Nanopore long-read sequencing on 72 cervical cancer genomes from an Ugandan dataset that was previously characterized using short-read sequencing. We found recurrent structural rearrangement patterns at HPV integration events, which we categorized as: del(etion)-like, dup(lication)-like, translocation, multibreakpoint, or repeat region integrations. Integrations involving amplified HPV-human concatemers, particularly multibreakpoint events, frequently harbored heterogeneous forms and copy numbers of the viral genome. Transcriptionally active integrants were characterized by unmethylated regions in both the viral and human genomes downstream from the viral transcription start site, resulting in HPV-human fusion transcripts. In contrast, integrants without evidence of expression lacked consistent methylation patterns. Furthermore, whereas transcriptional dysregulation was limited to genes within 200 kilobases of an HPV integrant, dysregulation of the human epigenome in the form of allelic differentially methylated regions affected megabase expanses of the genome, irrespective of the integrant's transcriptional status. By elucidating the structural, epigenetic, and allele-specific impacts of HPV integration, we provide insight into the role of integrated HPV in cervical cancer.
{"title":"Rearrangements of viral and human genomes at human papillomavirus integration events and their allele-specific impacts on cancer genome regulation","authors":"Vanessa L. Porter, Michelle Ng, Kieran O'Neill, Signe MacLennan, Richard D. Corbett, Luka Culibrk, Zeid Hamadeh, Marissa Iden, Rachel Schmidt, Shirng-Wern Tsaih, Carolyn Nakisige, Martin Origa, Jackson Orem, Glenn Chang, Jeremy Fan, Ka Ming Nip, Vahid Akbari, Simon K. Chan, James Hopkins, Richard A. Moore, Eric Chuah, Karen L. Mungall, Andrew J. Mungall, Inanc Birol, Steven J.M. Jones, Janet S. Rader, Marco A. Marra","doi":"10.1101/gr.279041.124","DOIUrl":"https://doi.org/10.1101/gr.279041.124","url":null,"abstract":"Human papillomavirus (HPV) integration has been implicated in transforming HPV infection into cancer. To resolve genome dysregulation associated with HPV integration, we performed Oxford Nanopore long-read sequencing on 72 cervical cancer genomes from an Ugandan dataset that was previously characterized using short-read sequencing. We found recurrent structural rearrangement patterns at HPV integration events, which we categorized as: del(etion)-like, dup(lication)-like, translocation, multibreakpoint, or repeat region integrations. Integrations involving amplified HPV-human concatemers, particularly multibreakpoint events, frequently harbored heterogeneous forms and copy numbers of the viral genome. Transcriptionally active integrants were characterized by unmethylated regions in both the viral and human genomes downstream from the viral transcription start site, resulting in HPV-human fusion transcripts. In contrast, integrants without evidence of expression lacked consistent methylation patterns. Furthermore, whereas transcriptional dysregulation was limited to genes within 200 kilobases of an HPV integrant, dysregulation of the human epigenome in the form of allelic differentially methylated regions affected megabase expanses of the genome, irrespective of the integrant's transcriptional status. By elucidating the structural, epigenetic, and allele-specific impacts of HPV integration, we provide insight into the role of integrated HPV in cervical cancer.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"68 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2024-12-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142783300","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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