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1.
Nat Genet ; 56(6): 1134-1146, 2024 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-38806714

RESUMEN

The functional impact and cellular context of mosaic structural variants (mSVs) in normal tissues is understudied. Utilizing Strand-seq, we sequenced 1,133 single-cell genomes from 19 human donors of increasing age, and discovered the heterogeneous mSV landscapes of hematopoietic stem and progenitor cells. While mSVs are continuously acquired throughout life, expanded subclones in our cohort are confined to individuals >60. Cells already harboring mSVs are more likely to acquire additional somatic structural variants, including megabase-scale segmental aneuploidies. Capitalizing on comprehensive single-cell micrococcal nuclease digestion with sequencing reference data, we conducted high-resolution cell-typing for eight hematopoietic stem and progenitor cells. Clonally expanded mSVs disrupt normal cellular function by dysregulating diverse cellular pathways, and enriching for myeloid progenitors. Our findings underscore the contribution of mSVs to the cellular and molecular phenotypes associated with the aging hematopoietic system, and establish a foundation for deciphering the molecular links between mSVs, aging and disease susceptibility in normal tissues.


Asunto(s)
Células Madre Hematopoyéticas , Mosaicismo , Humanos , Células Madre Hematopoyéticas/metabolismo , Células Madre Hematopoyéticas/citología , Persona de Mediana Edad , Adulto , Análisis de la Célula Individual/métodos , Anciano , Femenino , Masculino , Envejecimiento/genética , Anciano de 80 o más Años , Células Madre/metabolismo , Variación Genética
2.
Science ; 372(6537)2021 04 02.
Artículo en Inglés | MEDLINE | ID: mdl-33632895

RESUMEN

Long-read and strand-specific sequencing technologies together facilitate the de novo assembly of high-quality haplotype-resolved human genomes without parent-child trio data. We present 64 assembled haplotypes from 32 diverse human genomes. These highly contiguous haplotype assemblies (average minimum contig length needed to cover 50% of the genome: 26 million base pairs) integrate all forms of genetic variation, even across complex loci. We identified 107,590 structural variants (SVs), of which 68% were not discovered with short-read sequencing, and 278 SV hotspots (spanning megabases of gene-rich sequence). We characterized 130 of the most active mobile element source elements and found that 63% of all SVs arise through homology-mediated mechanisms. This resource enables reliable graph-based genotyping from short reads of up to 50,340 SVs, resulting in the identification of 1526 expression quantitative trait loci as well as SV candidates for adaptive selection within the human population.


Asunto(s)
Variación Genética , Genoma Humano , Haplotipos , Femenino , Genotipo , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Mutación INDEL , Secuencias Repetitivas Esparcidas , Masculino , Grupos de Población/genética , Sitios de Carácter Cuantitativo , Retroelementos , Análisis de Secuencia de ADN , Inversión de Secuencia , Secuenciación Completa del Genoma
3.
Cell Rep ; 31(1): 107465, 2020 04 07.
Artículo en Inglés | MEDLINE | ID: mdl-32268084

RESUMEN

TP53 deficiency is the most common alteration in cancer; however, this alone is typically insufficient to drive tumorigenesis. To identify genes promoting tumorigenesis in combination with TP53 deficiency, we perform genome-wide CRISPR-Cas9 knockout screens coupled with proliferation and transformation assays in isogenic cell lines. Loss of several known tumor suppressors enhances cellular proliferation and transformation. Loss of neddylation pathway genes promotes uncontrolled proliferation exclusively in TP53-deficient cells. Combined loss of CUL3 and TP53 activates an oncogenic transcriptional program governed by the nuclear factor κB (NF-κB), AP-1, and transforming growth factor ß (TGF-ß) pathways. This program maintains persistent cellular proliferation, induces partial epithelial to mesenchymal transition, and increases DNA damage, genomic instability, and chromosomal rearrangements. Our findings reveal CUL3 loss as a key event stimulating persistent proliferation in TP53-deficient cells. These findings may be clinically relevant, since TP53-CUL3-deficient cells are highly sensitive to ataxia telangiectasia mutated (ATM) inhibition, exposing a vulnerability that could be exploited for cancer treatment.


Asunto(s)
Proteínas Cullin/genética , Proteína p53 Supresora de Tumor/genética , Proteínas de la Ataxia Telangiectasia Mutada/antagonistas & inhibidores , Proteínas de la Ataxia Telangiectasia Mutada/genética , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Carcinogénesis/genética , Línea Celular , Línea Celular Tumoral , Proliferación Celular/fisiología , Proteínas Cullin/metabolismo , Transición Epitelial-Mesenquimal , Estudio de Asociación del Genoma Completo , Inestabilidad Genómica , Humanos , FN-kappa B/metabolismo , Epitelio Pigmentado de la Retina/citología , Factor de Crecimiento Transformador beta/metabolismo , Proteína p53 Supresora de Tumor/deficiencia , Proteína p53 Supresora de Tumor/metabolismo
4.
Nat Biotechnol ; 38(3): 343-354, 2020 03.
Artículo en Inglés | MEDLINE | ID: mdl-31873213

RESUMEN

Structural variation (SV), involving deletions, duplications, inversions and translocations of DNA segments, is a major source of genetic variability in somatic cells and can dysregulate cancer-related pathways. However, discovering somatic SVs in single cells has been challenging, with copy-number-neutral and complex variants typically escaping detection. Here we describe single-cell tri-channel processing (scTRIP), a computational framework that integrates read depth, template strand and haplotype phase to comprehensively discover SVs in individual cells. We surveyed SV landscapes of 565 single cells, including transformed epithelial cells and patient-derived leukemic samples, to discover abundant SV classes, including inversions, translocations and complex DNA rearrangements. Analysis of the leukemic samples revealed four times more somatic SVs than cytogenetic karyotyping, submicroscopic copy-number alterations, oncogenic copy-neutral rearrangements and a subclonal chromothripsis event. Advancing current methods, single-cell tri-channel processing can directly measure SV mutational processes in individual cells, such as breakage-fusion-bridge cycles, facilitating studies of clonal evolution, genetic mosaicism and SV formation mechanisms, which could improve disease classification for precision medicine.


Asunto(s)
Biología Computacional/métodos , Variación Estructural del Genoma , Leucemia/genética , Análisis de la Célula Individual/métodos , Línea Celular , Cromotripsis , Evolución Clonal , Reordenamiento Génico , Humanos , Mutación INDEL , Inversión de Secuencia , Translocación Genética
5.
Cancer Cell ; 34(6): 996-1011.e8, 2018 12 10.
Artículo en Inglés | MEDLINE | ID: mdl-30537516

RESUMEN

Identifying the earliest somatic changes in prostate cancer can give important insights into tumor evolution and aids in stratifying high- from low-risk disease. We integrated whole genome, transcriptome and methylome analysis of early-onset prostate cancers (diagnosis ≤55 years). Characterization across 292 prostate cancer genomes revealed age-related genomic alterations and a clock-like enzymatic-driven mutational process contributing to the earliest mutations in prostate cancer patients. Our integrative analysis identified four molecular subgroups, including a particularly aggressive subgroup with recurrent duplications associated with increased expression of ESRP1, which we validate in 12,000 tissue microarray tumors. Finally, we combined the patterns of molecular co-occurrence and risk-based subgroup information to deconvolve the molecular and clinical trajectories of prostate cancer from single patient samples.


Asunto(s)
Biomarcadores de Tumor/genética , Metilación de ADN , Regulación Neoplásica de la Expresión Génica , Neoplasias de la Próstata/genética , Transcriptoma , Adulto , Biomarcadores de Tumor/metabolismo , Evolución Molecular , Humanos , Masculino , Persona de Mediana Edad , Mutación , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismo , Factores de Riesgo , Secuenciación Completa del Genoma/métodos
6.
Nat Genet ; 49(1): 65-74, 2017 01.
Artículo en Inglés | MEDLINE | ID: mdl-27869826

RESUMEN

Extensive prior research focused on somatic copy-number alterations (SCNAs) affecting cancer genes, yet the extent to which recurrent SCNAs exert their influence through rearrangement of cis-regulatory elements (CREs) remains unclear. Here we present a framework for inferring cancer-related gene overexpression resulting from CRE reorganization (e.g., enhancer hijacking) by integrating SCNAs, gene expression data and information on topologically associating domains (TADs). Analysis of 7,416 cancer genomes uncovered several pan-cancer candidate genes, including IRS4, SMARCA1 and TERT. We demonstrate that IRS4 overexpression in lung cancer is associated with recurrent deletions in cis, and we present evidence supporting a tumor-promoting role. We additionally pursued cancer-type-specific analyses and uncovered IGF2 as a target for enhancer hijacking in colorectal cancer. Recurrent tandem duplications intersecting with a TAD boundary mediate de novo formation of a 3D contact domain comprising IGF2 and a lineage-specific super-enhancer, resulting in high-level gene activation. Our framework enables systematic inference of CRE rearrangements mediating dysregulation in cancer.


Asunto(s)
Variaciones en el Número de Copia de ADN/genética , Elementos de Facilitación Genéticos/genética , Regulación Neoplásica de la Expresión Génica , Proteínas Sustrato del Receptor de Insulina/genética , Factor II del Crecimiento Similar a la Insulina/genética , Neoplasias/genética , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Humanos , Regiones Promotoras Genéticas
7.
Mol Syst Biol ; 11(9): 828, 2015 Sep 28.
Artículo en Inglés | MEDLINE | ID: mdl-26415501

RESUMEN

A remarkable observation emerging from recent cancer genome analyses is the identification of chromothripsis as a one-off genomic catastrophe, resulting in massive somatic DNA structural rearrangements (SRs). Largely due to lack of suitable model systems, the mechanistic basis of chromothripsis has remained elusive. We developed an integrative method termed "complex alterations after selection and transformation (CAST)," enabling efficient in vitro generation of complex DNA rearrangements including chromothripsis, using cell perturbations coupled with a strong selection barrier followed by massively parallel sequencing. We employed this methodology to characterize catastrophic SR formation processes, their temporal sequence, and their impact on gene expression and cell division. Our in vitro system uncovered a propensity of chromothripsis to occur in cells with damaged telomeres, and in particular in hyperploid cells. Analysis of primary medulloblastoma cancer genomes verified the link between hyperploidy and chromothripsis in vivo. CAST provides the foundation for mechanistic dissection of complex DNA rearrangement processes.


Asunto(s)
Cromosomas Humanos/genética , Reordenamiento Génico , Genoma Humano/genética , Inestabilidad Genómica/genética , Neoplasias/genética , Aneuploidia , División Celular , Línea Celular , Aberraciones Cromosómicas , Variaciones en el Número de Copia de ADN/genética , Humanos , Meduloblastoma/genética , Poliploidía , Telómero/genética , Telómero/patología , Proteína 2 de Unión a Repeticiones Teloméricas/genética , Proteína 2 de Unión a Repeticiones Teloméricas/metabolismo
8.
Nat Genet ; 47(9): 1020-1029, 2015 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-26214592

RESUMEN

TCF3-HLF-positive acute lymphoblastic leukemia (ALL) is currently incurable. Using an integrated approach, we uncovered distinct mutation, gene expression and drug response profiles in TCF3-HLF-positive and treatment-responsive TCF3-PBX1-positive ALL. We identified recurrent intragenic deletions of PAX5 or VPREB1 in constellation with the fusion of TCF3 and HLF. Moreover somatic mutations in the non-translocated allele of TCF3 and a reduction of PAX5 gene dosage in TCF3-HLF ALL suggest cooperation within a restricted genetic context. The enrichment for stem cell and myeloid features in the TCF3-HLF signature may reflect reprogramming by TCF3-HLF of a lymphoid-committed cell of origin toward a hybrid, drug-resistant hematopoietic state. Drug response profiling of matched patient-derived xenografts revealed a distinct profile for TCF3-HLF ALL with resistance to conventional chemotherapeutics but sensitivity to glucocorticoids, anthracyclines and agents in clinical development. Striking on-target sensitivity was achieved with the BCL2-specific inhibitor venetoclax (ABT-199). This integrated approach thus provides alternative treatment options for this deadly disease.


Asunto(s)
Proteínas de Fusión Oncogénica/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Animales , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Técnicas de Cocultivo , Estudios de Cohortes , Análisis Mutacional de ADN , Resistencia a Antineoplásicos , Femenino , Expresión Génica , Estudios de Asociación Genética , Genómica , Humanos , Inmunoglobulina de Cadenas Ligeras Subrogadas/genética , Concentración 50 Inhibidora , Estimación de Kaplan-Meier , Masculino , Ratones Endogámicos NOD , Ratones SCID , Mutación , Proteínas de Fusión Oncogénica/metabolismo , Factor de Transcripción PAX5/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamiento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/mortalidad , Eliminación de Secuencia , Ensayos Antitumor por Modelo de Xenoinjerto
9.
Nature ; 511(7510): 428-34, 2014 Jul 24.
Artículo en Inglés | MEDLINE | ID: mdl-25043047

RESUMEN

Medulloblastoma is a highly malignant paediatric brain tumour currently treated with a combination of surgery, radiation and chemotherapy, posing a considerable burden of toxicity to the developing child. Genomics has illuminated the extensive intertumoral heterogeneity of medulloblastoma, identifying four distinct molecular subgroups. Group 3 and group 4 subgroup medulloblastomas account for most paediatric cases; yet, oncogenic drivers for these subtypes remain largely unidentified. Here we describe a series of prevalent, highly disparate genomic structural variants, restricted to groups 3 and 4, resulting in specific and mutually exclusive activation of the growth factor independent 1 family proto-oncogenes, GFI1 and GFI1B. Somatic structural variants juxtapose GFI1 or GFI1B coding sequences proximal to active enhancer elements, including super-enhancers, instigating oncogenic activity. Our results, supported by evidence from mouse models, identify GFI1 and GFI1B as prominent medulloblastoma oncogenes and implicate 'enhancer hijacking' as an efficient mechanism driving oncogene activation in a childhood cancer.


Asunto(s)
Proteínas de Unión al ADN/genética , Elementos de Facilitación Genéticos/genética , Variación Estructural del Genoma/genética , Meduloblastoma/genética , Oncogenes/genética , Proteínas Proto-Oncogénicas/genética , Proteínas Represoras/genética , Factores de Transcripción/genética , Animales , Niño , Cromosomas Humanos Par 9/genética , Proteínas de Unión al ADN/metabolismo , Humanos , Meduloblastoma/clasificación , Meduloblastoma/patología , Ratones , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Represoras/metabolismo , Factores de Transcripción/metabolismo
10.
Nat Genet ; 45(8): 927-32, 2013 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-23817572

RESUMEN

Pilocytic astrocytoma, the most common childhood brain tumor, is typically associated with mitogen-activated protein kinase (MAPK) pathway alterations. Surgically inaccessible midline tumors are therapeutically challenging, showing sustained tendency for progression and often becoming a chronic disease with substantial morbidities. Here we describe whole-genome sequencing of 96 pilocytic astrocytomas, with matched RNA sequencing (n = 73), conducted by the International Cancer Genome Consortium (ICGC) PedBrain Tumor Project. We identified recurrent activating mutations in FGFR1 and PTPN11 and new NTRK2 fusion genes in non-cerebellar tumors. New BRAF-activating changes were also observed. MAPK pathway alterations affected all tumors analyzed, with no other significant mutations identified, indicating that pilocytic astrocytoma is predominantly a single-pathway disease. Notably, we identified the same FGFR1 mutations in a subset of H3F3A-mutated pediatric glioblastoma with additional alterations in the NF1 gene. Our findings thus identify new potential therapeutic targets in distinct subsets of pilocytic astrocytoma and childhood glioblastoma.


Asunto(s)
Astrocitoma/genética , Neoplasias Encefálicas/genética , Mutación , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Receptor trkB/genética , Animales , Astrocitoma/metabolismo , Secuencia de Bases , Neoplasias Encefálicas/metabolismo , Línea Celular , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/metabolismo , Puntos de Rotura del Cromosoma , Cromosomas Humanos Par 6 , Cromosomas Humanos Par 9 , Factores de Crecimiento de Fibroblastos/metabolismo , Humanos , Sistema de Señalización de MAP Quinasas , Ratones , Modelos Moleculares , Proteínas de Fusión Oncogénica/química , Proteínas de Fusión Oncogénica/genética , Conformación Proteica , Proteínas Proto-Oncogénicas B-raf/química , Proteínas Proto-Oncogénicas B-raf/genética , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/metabolismo , Receptor trkB/metabolismo
11.
Eur J Hum Genet ; 21(10): 1177-80, 2013 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-23422942

RESUMEN

Most balanced chromosomal aberrations are not associated with a clinical phenotype, however, in some patients they may disrupt gene structure. With the development of various next-generation sequencing techniques, fast and specific analyses of the breakpoint regions of chromosomal rearrangements are possible. Here, we report on a 19-year-old woman with a de novo balanced translocation t(2;8)(p13.2;q22.1) and a severe clinical phenotype including intellectual disability, epilepsy, behavioral features resembling autism, and minor dysmorphic features. By next-generation sequencing, we defined the breakpoints and found disruption of the exocyst complex component 6B (EXOC6B) gene in intron 1 on chromosome 2p13.2 involving two Alu elements with a homology of 81%. No gene was found at the respective breakpoint on chromosome 8. Expression analysis of the EXOC6B in blood lymphocytes and buccal smear revealed reduced expression in the patient in comparison with the control. Our findings in combination with one recently published case and one other patient listed in DECIPHER v5.1 indicate EXOC6B as a gene relevant for intellectual development and electrophysiological stability.


Asunto(s)
Anomalías Múltiples/genética , Discapacidades del Desarrollo/genética , Epilepsia/genética , Proteínas de Unión al GTP/genética , Translocación Genética , Anomalías Múltiples/metabolismo , Puntos de Rotura del Cromosoma , Cromosomas Humanos Par 2/genética , Cromosomas Humanos Par 8/genética , Discapacidades del Desarrollo/metabolismo , Epilepsia/metabolismo , Femenino , Proteínas de Unión al GTP/metabolismo , Humanos , Linfocitos/metabolismo , Mucosa Bucal/metabolismo , Síndrome , Adulto Joven
12.
Cancer Cell ; 23(2): 159-70, 2013 Feb 11.
Artículo en Inglés | MEDLINE | ID: mdl-23410972

RESUMEN

Early-onset prostate cancer (EO-PCA) represents the earliest clinical manifestation of prostate cancer. To compare the genomic alteration landscapes of EO-PCA with "classical" (elderly-onset) PCA, we performed deep sequencing-based genomics analyses in 11 tumors diagnosed at young age, and pursued comparative assessments with seven elderly-onset PCA genomes. Remarkable age-related differences in structural rearrangement (SR) formation became evident, suggesting distinct disease pathomechanisms. Whereas EO-PCAs harbored a prevalence of balanced SRs, with a specific abundance of androgen-regulated ETS gene fusions including TMPRSS2:ERG, elderly-onset PCAs displayed primarily non-androgen-associated SRs. Data from a validation cohort of > 10,000 patients showed age-dependent androgen receptor levels and a prevalence of SRs affecting androgen-regulated genes, further substantiating the activity of a characteristic "androgen-type" pathomechanism in EO-PCA.


Asunto(s)
Reordenamiento Génico , Genómica , Proteínas de Fusión Oncogénica/genética , Neoplasias de la Próstata/genética , Receptores Androgénicos/genética , Serina Endopeptidasas/genética , Transactivadores/genética , Adulto , Anciano , Anciano de 80 o más Años , Biología Computacional , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Masculino , Persona de Mediana Edad , Regulador Transcripcional ERG
13.
Nature ; 488(7409): 100-5, 2012 Aug 02.
Artículo en Inglés | MEDLINE | ID: mdl-22832583

RESUMEN

Medulloblastoma is an aggressively growing tumour, arising in the cerebellum or medulla/brain stem. It is the most common malignant brain tumour in children, and shows tremendous biological and clinical heterogeneity. Despite recent treatment advances, approximately 40% of children experience tumour recurrence, and 30% will die from their disease. Those who survive often have a significantly reduced quality of life. Four tumour subgroups with distinct clinical, biological and genetic profiles are currently identified. WNT tumours, showing activated wingless pathway signalling, carry a favourable prognosis under current treatment regimens. SHH tumours show hedgehog pathway activation, and have an intermediate prognosis. Group 3 and 4 tumours are molecularly less well characterized, and also present the greatest clinical challenges. The full repertoire of genetic events driving this distinction, however, remains unclear. Here we describe an integrative deep-sequencing analysis of 125 tumour-normal pairs, conducted as part of the International Cancer Genome Consortium (ICGC) PedBrain Tumor Project. Tetraploidy was identified as a frequent early event in Group 3 and 4 tumours, and a positive correlation between patient age and mutation rate was observed. Several recurrent mutations were identified, both in known medulloblastoma-related genes (CTNNB1, PTCH1, MLL2, SMARCA4) and in genes not previously linked to this tumour (DDX3X, CTDNEP1, KDM6A, TBR1), often in subgroup-specific patterns. RNA sequencing confirmed these alterations, and revealed the expression of what are, to our knowledge, the first medulloblastoma fusion genes identified. Chromatin modifiers were frequently altered across all subgroups. These findings enhance our understanding of the genomic complexity and heterogeneity underlying medulloblastoma, and provide several potential targets for new therapeutics, especially for Group 3 and 4 patients.


Asunto(s)
Neoplasias Cerebelosas/genética , Genoma Humano/genética , Meduloblastoma/genética , Envejecimiento/genética , Secuencia de Aminoácidos , Transformación Celular Neoplásica , Neoplasias Cerebelosas/clasificación , Neoplasias Cerebelosas/diagnóstico , Neoplasias Cerebelosas/patología , Niño , Cromatina/metabolismo , Cromosomas Humanos/genética , ARN Helicasas DEAD-box/genética , ADN Helicasas/genética , Proteínas de Unión al ADN/genética , Genómica , Proteínas Hedgehog/metabolismo , Secuenciación de Nucleótidos de Alto Rendimiento , Histona Demetilasas/genética , Humanos , Meduloblastoma/clasificación , Meduloblastoma/diagnóstico , Meduloblastoma/patología , Metilación , Mutación/genética , Tasa de Mutación , Proteínas de Neoplasias/genética , Proteínas Nucleares/genética , Proteínas de Fusión Oncogénica/genética , Receptores Patched , Receptor Patched-1 , Fosfoproteínas Fosfatasas/genética , Poliploidía , Receptores de Superficie Celular/genética , Análisis de Secuencia de ARN , Transducción de Señal , Proteínas de Dominio T Box/genética , Factores de Transcripción/genética , Proteínas Wnt/metabolismo , beta Catenina/genética
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