Genome research最新文献

{"title":"Optical genome mapping enables accurate testing of large repeat expansions","authors":"Bart van der Sanden, Kornelia Neveling, Syukri Shukor, Michael D. Gallagher, Joyce Lee, Stephanie L. Burke, Maartje Pennings, Ronald van Beek, Michiel Oorsprong, Ellen Kater-Baats, Eveline Kamping, Alide A. Tieleman, Nicol C. Voermans, Ingrid E. Scheffer, Jozef Gecz, Mark A. Corbett, Lisenka E.L.M. Vissers, Andy Wing Chun Pang, Alex Hastie, Erik-Jan Kamsteeg, Alexander Hoischen","doi":"10.1101/gr.279491.124","DOIUrl":"https://doi.org/10.1101/gr.279491.124","url":null,"abstract":"Short tandem repeats (STRs) are common variations in human genomes that frequently expand or contract, causing genetic disorders, mainly when expanded. Traditional diagnostic methods for identifying these expansions, such as repeat-primed PCR and Southern blotting, are often labor-intensive, locus-specific, and are unable to precisely determine long repeat expansions. Sequencing-based methods, although capable of genome-wide detection, are limited by inaccuracy (short-read technologies) and high associated costs (long-read technologies). This study evaluated optical genome mapping (OGM) as an efficient, accurate approach for measuring STR lengths and assessing somatic stability in 85 samples with known pathogenic repeat expansions in <em>DMPK</em>, <em>CNBP</em>, and <em>RFC1</em>, causing myotonic dystrophy types 1 and 2 and cerebellar ataxia, neuropathy, and vestibular areflexia syndrome (CANVAS), respectively. Three workflows—manual de novo assembly, local guided assembly (local-GA), and a molecule distance script—were applied, of which the latter two were developed as part of this study to assess the repeat sizes and somatic repeat stability. OGM successfully identified 84/85 (98.8%) of the pathogenic expansions, distinguishing between wild-type and expanded alleles or between two expanded alleles in recessive cases, with greater accuracy than standard of care (SOC) for long repeats and no apparent upper size limit. Notably, OGM detected somatic instability in a subset of <em>DMPK</em>, <em>CNBP</em>, and <em>RFC1</em> samples. These findings suggest OGM could advance diagnostic accuracy for large repeat expansions, providing a more comprehensive genome-wide assay for repeat expansion disorders by measuring exact repeat lengths and somatic instability across multiple loci simultaneously.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"27 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2025-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143666279","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}

{"title":"De novo detection of somatic variants in high-quality long-read single-cell RNA sequencing data","authors":"Arthur Dondi, Nico Borgsmüller, Pedro F. Ferreira, Brian J. Haas, Francis Jacob, Viola Heinzelmann-Schwarz, Tumor Profiler Consortium, Niko Beerenwinkel","doi":"10.1101/gr.279281.124","DOIUrl":"https://doi.org/10.1101/gr.279281.124","url":null,"abstract":"In cancer, genetic and transcriptomic variations generate clonal heterogeneity, leading to treatment resistance. Long-read single-cell RNA sequencing (LR scRNA-seq) has the potential to detect genetic and transcriptomic variations simultaneously. Here, we present LongSom, a computational workflow leveraging high-quality LR scRNA-seq data to call de novo somatic single-nucleotide variants (SNVs), including in mitochondria (mtSNVs), copy number alterations (CNAs), and gene fusions, to reconstruct the tumor clonal heterogeneity. Before somatic variant calling, LongSom reannotates marker gene-based cell types using cell mutational profiles. LongSom distinguishes somatic SNVs from noise and germline polymorphisms by applying an extensive set of hard filters and statistical tests. Applying LongSom to human ovarian cancer samples, we detected clinically relevant somatic SNVs that were validated against matched DNA samples. Leveraging somatic SNVs and fusions, LongSom found subclones with different predicted treatment outcomes. In summary, LongSom enables de novo variant detection without the need for normal samples, facilitating the study of cancer evolution, clonal heterogeneity, and treatment resistance.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"15 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2025-03-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143661416","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}

{"title":"Heterogeneous and novel transcript expression in single cells of patient-derived clear cell renal cell carcinoma organoids","authors":"Tülay Karakulak, Natalia Zajac, Hella Anna Bolck, Anna Bratus-Neuenschwander, Qin Zhang, Weihong Qi, Debleena Basu, Tamara Carrasco Oltra, Hubert Rehrauer, Christian von Mering, Holger Moch, Abdullah Kahraman","doi":"10.1101/gr.279345.124","DOIUrl":"https://doi.org/10.1101/gr.279345.124","url":null,"abstract":"Splicing is often dysregulated in cancer, leading to alterations in the expression of canonical and alternatively spliced isoforms. We used the multiplexed arrays sequencing (MAS-seq) protocol of PacBio to sequence full-length transcripts in patient-derived organoid (PDO) cells of clear cell renal cell carcinoma (ccRCC). The sequencing revealed a heterogeneous dysregulation of splicing across 2599 single ccRCC cells. The majority of novel transcripts could be removed with stringent filtering criteria. The remaining 31,531 transcripts (36.6% of the 86,182 detected transcripts on average) were previously uncharacterized. In contrast to known transcripts, many of the novel transcripts have cell-specific expression. Novel transcripts common to ccRCC cells belong to genes involved in ccRCC-related pathways, such as hypoxia and oxidative phosphorylation. A novel transcript of the ccRCC-related gene nicotinamide N-methyltransferase is validated using PCR. Moreover, &gt;50% of novel transcripts possess a predicted complete protein-coding open reading frame. An analysis of the most dominant transcript-switching events between ccRCC and non-ccRCC cells shows many switching events that are cell- and sample-specific, underscoring the heterogeneity of alternative splicing events in ccRCC. Overall, our study elucidates the intricate transcriptomic architecture of ccRCC, underlying its aggressive phenotype and providing insights into its molecular complexity.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"61 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2025-03-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143661415","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}

{"title":"Optical genome mapping identifies rare structural variants in neural tube defects","authors":"Nikhil S. Sahajpal, Jane Dean, Benjamin Hilton, Timothy Fee, Cindy Skinner, Alex Hastie, Barbara R. DuPont, Alka Chaubey, Michael J. Friez, Roger E. Stevenson","doi":"10.1101/gr.279318.124","DOIUrl":"https://doi.org/10.1101/gr.279318.124","url":null,"abstract":"Neural tube defects (NTDs) are the most common birth defects of the central nervous system and occur as either isolated malformations or accompanied by anomalies of other systems. The genetic basis of NTDs remains poorly understood using karyotyping, chromosomal microarray, and short-read sequencing, with only a limited number of pathogenic variants identified. Collectively, these technologies may fail to detect rare structural variants (SVs) in the genome, which may cause these birth defects. Therefore, optical genome mapping (OGM) was applied to investigate 104 NTD cases, of which 74 were isolated NTDs and 30 were NTDs with other malformations. A stepwise approach was undertaken to ascertain candidate variants using population and internal databases and performing parental studies when possible. This analysis identifies diagnostic findings in 8% of cases (8/104) and candidate findings in an additional 22% of cases (23/104). Of the candidate findings, 9% of cases (9/104) have SVs impacting genes associated with NTDs in mouse, and 13% of cases (14/104) have SVs impacting genes implicated in the neural tube development pathways. This study identifies <em>RMND5A, HNRNPC</em>, <em>FOXD4</em>, and <em>RBBP4</em> as strong candidate genes associated with NTDs, and expands the phenotypic spectrum of <em>AMER1</em> and <em>TGIF1</em> to include NTDs. This study constitutes the first systematic investigation of SVs using OGM to elucidate the genetic determinants of NTDs. The data provide key insights into the pathogenesis of NTDs and demonstrate the contribution of SVs in the genome to these birth defects.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"19 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2025-03-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143661539","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}

{"title":"Closing the gaps, and improving somatic structural variant analysis and benchmarking using CHM13-T2T","authors":"Luis F. Paulin, Jeremy Fan, Kieran O'Neill, Erin Pleasance, Vanessa L. Porter, Steven J.M. Jones, Fritz J. Sedlazeck","doi":"10.1101/gr.279352.124","DOIUrl":"https://doi.org/10.1101/gr.279352.124","url":null,"abstract":"The complexities of cancer genomes are becoming more easily interpreted due to advancements in sequencing technologies and improved bioinformatic analysis. Structural variants (SVs) represent an important subset of somatic events in tumors. While the detection of SVs has been markedly improved by the development of long-read sequencing, somatic variant identification and annotation remain challenging. We hypothesized that the use of a completed human reference genome (CHM13-T2T) would improve somatic SV calling. Our findings in a tumor–normal matched benchmark sample and three patient samples show that the CHM13-T2T improves SV detection accuracy compared to GRCh38 with a notable reduction in false-positive calls, and thus supports improved prioritization. We also overcame the lack of annotation resources for CHM13-T2T by lifting over CHM13-T2T-aligned reads to the GRCh38 genome, therefore combining both improved alignment and advanced annotations. In this process, we assessed the current SV benchmark set for COLO829/COLO829BL across four replicates sequenced at different centers with different long-read technologies. We discovered instability of this cell line across these replicates; 346 SVs (1.13%) were only discoverable in a single replicate. We identify 54 somatic SVs, which appear to be stable as they are consistently present across the four replicates. As such, we propose this consensus set as an updated benchmark for somatic SV calling and include both GRCh38 and CHM13-T2T coordinates in our benchmark. Our work demonstrates new approaches to optimize somatic SV detection in cancer with potential improvements in other genetic diseases.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"33 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2025-03-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143641043","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}

{"title":"BINDER achieves accurate identification of hierarchical TADs by comprehensively characterizing consensus TAD boundaries","authors":"Yangyang Liu, Sr., Bingqiang Liu, Sr., Juntao Liu, Sr.","doi":"10.1101/gr.279647.124","DOIUrl":"https://doi.org/10.1101/gr.279647.124","url":null,"abstract":"As a crucial chromatin structure, hierarchical TADs play important roles in epigenetic organization, transcriptional activity, gene regulation, and cell differentiation. Currently, it remains a highly challenging task to accurately identify hierarchical TADs in a computational manner. The key bottleneck for existing TAD callers lies in the difficulty on prediction of precise TAD boundaries. We solve this problem by introducing a novel algorithm, called BINDER, which conducts a boundary consensus approach, and then precisely locate hierarchical TAD boundaries by developing a multifaceted boundary characterization strategy. After comparison with other leading TAD callers, BINDER shows great improvement in identifying hierarchical TADs and exhibits the strongest robustness with ultrasparse data, which fully indicates the importance of boundary identification in calling hierarchical TADs. Applying BINDER to experimental data and mouse hematopoietic cases, we find that the hierarchical TADs identified by BINDER show strong biological relevance with epigenetic organization, transcriptional activity, DNA motifs, and coregulation during cellular differentiation. BINDER discovers differences in the enrichment of two specific transcription factors, CHD1 and CHD2 at TAD boundaries with different hierarchies. It also observes variations in gene expression of TADs with different hierarchies during cellular differentiation.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"60 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2025-03-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143641044","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}

{"title":"Biosurfer for systematic tracking of regulatory mechanisms leading to protein isoform diversity","authors":"Mayank Murali, Jamie Saquing, Senbao Lu, Ziyang Gao, Emily F. Watts, Ben Jordan, Zachary Peters Wakefield, Ana Fiszbein, David R. Cooper, Peter J. Castaldi, Dmitry Korkin, Gloria M. Sheynkman","doi":"10.1101/gr.279317.124","DOIUrl":"https://doi.org/10.1101/gr.279317.124","url":null,"abstract":"Long-read RNA-seq has shed light on transcriptomic complexity, but questions remain about the functionality of downstream protein products. We introduce Biosurfer, a computational approach for comparing protein isoforms, while systematically tracking the transcriptional, splicing, and translational variations that underlie differences in the sequences of the protein products. Using Biosurfer, we analyzed the differences in 35,082 pairs of GENCODE annotated protein isoforms, finding a majority (70%) of variable N-termini are due to the alternative transcription start sites, while only 9% arise from 5′ UTR alternative splicing (AS). Biosurfer's detailed tracking of nucleotide-to-residue relationships helps reveal an uncommonly tracked source of single amino acid residue changes arising from the codon splits at junctions. For 17% of internal sequence changes, such split codon patterns lead to single residue differences, termed “ragged codons.” Of variable C-termini, 72% involve splice- or intron retention-induced reading frameshifts. We systematically characterize an unusual pattern of reading frame changes, in which the first frameshift is closely followed by a distinct second frameshift that restores the original frame, which we term a “snapback” frameshift. We analyze the long-read RNA-seq-predicted proteome of a human cell line and find similar trends as compared to our GENCODE analysis, with the exception of a higher proportion of transcripts predicted to undergo nonsense-mediated decay. Biosurfer's comprehensive characterization of long-read RNA-seq data sets should accelerate insights of the functional role of protein isoforms, providing mechanistic explanation of the origins of the proteomic diversity driven by the AS. Biosurfer is available as a Python package.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"32 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2025-03-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143627426","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}

{"title":"Taurine pangenome uncovers a segmental duplication upstream of <i>KIT</i> associated with depigmentation in white-headed cattle.","authors":"Sotiria Milia, Alexander S 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 of 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 that examines assembly path similarities within the graph to reveal an association between two most likely serial alleles of a complex structural variant (SV) 66 kb upstream of <i>KIT</i> and facial depigmentation. The complex SV 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 SV 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 SVs.</p>","PeriodicalId":12678,"journal":{"name":"Genome research","volume":" ","pages":""},"PeriodicalIF":6.2,"publicationDate":"2025-03-14","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}

{"title":"Accurate fusion transcript identification from long- and short-read isoform sequencing at bulk or single-cell resolution","authors":"Qian Qin, Victoria Popic, Kirsty Wienand, Houlin Yu, Emily White, Akanksha Khorgade, Asa Shin, Christophe Georgescu, Catarina D. Campbell, Arthur Dondi, Niko Beerenwinkel, Francisca Vazquez, Aziz M. Al'Khafaji, Brian J. Haas","doi":"10.1101/gr.279200.124","DOIUrl":"https://doi.org/10.1101/gr.279200.124","url":null,"abstract":"Gene fusions are found as cancer drivers in diverse adult and pediatric cancers. Accurate detection of fusion transcripts is essential in cancer clinical diagnostics and prognostics and for guiding therapeutic development. Most currently available methods for fusion transcript detection are compatible with Illumina RNA-seq involving highly accurate short-read sequences. Recent advances in long-read isoform sequencing enable the detection of fusion transcripts at unprecedented resolution in bulk and single-cell samples. Here, we developed a new computational tool, CTAT-LR-Fusion, to detect fusion transcripts from long-read RNA-seq with or without companion short reads, with applications to bulk or single-cell transcriptomes. We demonstrate that CTAT-LR-Fusion exceeds the fusion detection accuracy of alternative methods as benchmarked with simulated and genuine long-read RNA-seq. Using short- and long-read RNA-seq, we further apply CTAT-LR-Fusion to bulk transcriptomes of nine tumor cell lines and to tumor single cells derived from a melanoma sample and three metastatic high-grade serous ovarian carcinoma samples. In both bulk and single-cell RNA-seq, long isoform reads yield higher sensitivity for fusion detection than short reads with notable exceptions. By combining short and long reads in CTAT-LR-Fusion, we are able to further maximize the detection of fusion splicing isoforms and fusion-expressing tumor cells.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"6 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2025-03-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143627427","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}

{"title":"KRAB zinc-finger proteins regulate endogenous retroviruses to sculpt germline transcriptomes and genome evolution","authors":"Kai Otsuka, Akihiko Sakashita, So Maezawa, Richard M. Schultz, Satoshi H. Namekawa","doi":"10.1101/gr.279924.124","DOIUrl":"https://doi.org/10.1101/gr.279924.124","url":null,"abstract":"As transposable elements (TEs) coevolved with the host genome, the host genome exploited TEs as functional regulatory elements of gene expression. Here we show that a subset of KRAB domain–containing zinc-finger proteins (KZFPs), which are highly expressed in mitotically dividing spermatogonia, repress the enhancer function of endogenous retroviruses (ERVs) and that the release from KZFP-mediated repression allows activation of ERV enhancers upon entry into meiosis. This regulatory feature is observed for independently evolved KZFPs and ERVs in mice and humans, suggesting evolutionary conservation in mammals. Further, we show that KZFP-targeted ERVs are underrepresented on the sex chromosomes in meiosis, suggesting that meiotic sex chromosome inactivation (MSCI) may antagonize the coevolution of KZFPs and ERVs in mammals. Our study uncovers a mechanism by which a subset of KZFPs regulate ERVs to sculpt germline transcriptomes. We propose that epigenetic programming during the transition from mitotic spermatogonia to meiotic spermatocytes facilitates the coevolution of KZFPs and TEs on autosomes and is antagonized by MSCI.","PeriodicalId":12678,"journal":{"name":"Genome research","volume":"208 1","pages":""},"PeriodicalIF":7.0,"publicationDate":"2025-03-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143608348","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}