RESUMO
With the increasing number of sequencing projects involving families, quality control tools optimized for family genome sequencing are needed. However, accurately quantifying contamination in a DNA mixture is particularly difficult when genetically related family members are the sources. We developed TrioMix, a maximum likelihood estimation (MLE) framework based on Mendel's law of inheritance, to quantify DNA mixture between family members in genome sequencing data of parent-offspring trios. TrioMix can accurately deconvolute any intrafamilial DNA contamination, including parent-offspring, sibling-sibling, parent-parent, and even multiple familial sources. In addition, TrioMix can be applied to detect genomic abnormalities that deviate from Mendelian inheritance patterns, such as uniparental disomy (UPD) and chimerism. A genome-wide depth and variant allele frequency plot generated by TrioMix facilitates tracing the origin of Mendelian inheritance deviations. We showed that TrioMix could accurately deconvolute genomes in both simulated and real data sets.
Assuntos
Contaminação por DNA , Genoma , Humanos , Mapeamento Cromossômico , Dissomia Uniparental , Bases de Dados GenéticasRESUMO
Identifying genomic variants is a fundamental first step toward the understanding of the role of inherited and acquired variation in disease. The accelerating growth in the corpus of sequencing data that underpins such analysis is making the data-download bottleneck more evident, placing substantial burdens on the research community to keep pace. As a result, the search for alternative approaches to the traditional "download and analyze" paradigm on local computing resources has led to a rapidly growing demand for cloud-computing solutions for genomics analysis. Here, we introduce the Genome Variant Investigation Platform (GenomeVIP), an open-source framework for performing genomics variant discovery and annotation using cloud- or local high-performance computing infrastructure. GenomeVIP orchestrates the analysis of whole-genome and exome sequence data using a set of robust and popular task-specific tools, including VarScan, GATK, Pindel, BreakDancer, Strelka, and Genome STRiP, through a web interface. GenomeVIP has been used for genomic analysis in large-data projects such as the TCGA PanCanAtlas and in other projects, such as the ICGC Pilots, CPTAC, ICGC-TCGA DREAM Challenges, and the 1000 Genomes SV Project. Here, we demonstrate GenomeVIP's ability to provide high-confidence annotated somatic, germline, and de novo variants of potential biological significance using publicly available data sets.
Assuntos
Computação em Nuvem , Variação Genética , Genoma Humano , Genômica/métodos , Neoplasias/genética , Software , Bases de Dados Genéticas , Sequenciamento de Nucleotídeos em Larga Escala/métodos , HumanosAssuntos
Diabetes Mellitus Tipo 1/congênito , Diarreia/genética , Doenças Genéticas Ligadas ao Cromossomo X/genética , Doenças do Sistema Imunitário/congênito , Fator de Transcrição STAT1/genética , Diabetes Mellitus Tipo 1/genética , Feminino , Mutação com Ganho de Função , Mutação em Linhagem Germinativa , Humanos , Doenças do Sistema Imunitário/genética , MosaicismoRESUMO
While most dizygotic twins have a dichorionic placenta, rare cases of dizygotic twins with a monochorionic placenta have been reported. The monochorionic placenta in dizygotic twins allows in utero exchange of embryonic cells, resulting in chimerism in the twins. In practice, this chimerism is incidentally identified in mixed ABO blood types or in the presence of cells with a discordant sex chromosome. Here, we applied whole-genome sequencing to one triplet and one twin family to precisely understand their zygotic compositions, using millions of genomic variants as barcodes of zygotic origins. Peripheral blood showed asymmetrical contributions from two sister zygotes, where one of the zygotes was the major clone in both twins. Single-cell RNA sequencing of peripheral blood tissues further showed differential contributions from the two sister zygotes across blood cell types. In contrast, buccal tissues were pure in genetic composition, suggesting that in utero cellular exchanges were confined to the blood tissues. Our study illustrates the cellular history of twinning during human development, which is critical for managing the health of chimeric individuals in the era of genomic medicine.
Assuntos
Gêmeos Dizigóticos , Sequenciamento Completo do Genoma , Zigoto , Humanos , Feminino , Gêmeos Dizigóticos/genética , Zigoto/metabolismo , Gravidez , Quimerismo , Placenta/metabolismo , Masculino , Quimera/genética , Gêmeos Monozigóticos/genéticaRESUMO
Somatic cells accumulate genomic alterations with age; however, our understanding of mitochondrial DNA (mtDNA) mosaicism remains limited. Here we investigated the genomes of 2,096 clones derived from three cell types across 31 donors, identifying 6,451 mtDNA variants with heteroplasmy levels of â³0.3%. While the majority of these variants were unique to individual clones, suggesting stochastic acquisition with age, 409 variants (6%) were shared across multiple embryonic lineages, indicating their origin from heteroplasmy in fertilized eggs. The mutational spectrum exhibited replication-strand bias, implicating mtDNA replication as a major mutational process. We evaluated the mtDNA mutation rate (5.0 × 10-8 per base pair) and a turnover frequency of 10-20 per year, which are fundamental components shaping the landscape of mtDNA mosaicism over a lifetime. The expansion of mtDNA-truncating mutations toward homoplasmy was substantially suppressed. Our findings provide comprehensive insights into the origins, dynamics and functional consequences of mtDNA mosaicism in human somatic cells.
Assuntos
DNA Mitocondrial , Mosaicismo , Mutação , Humanos , DNA Mitocondrial/genética , Heteroplasmia/genética , Taxa de Mutação , Mitocôndrias/genética , Genoma Mitocondrial , Replicação do DNA/genética , Feminino , MasculinoRESUMO
Multiple myeloma (MM) is characterized by the uncontrolled proliferation of plasma cells. Despite recent treatment advances, it is still incurable as disease progression is not fully understood. To investigate MM and its immune environment, we apply single cell RNA and linked-read whole genome sequencing to profile 29 longitudinal samples at different disease stages from 14 patients. Here, we collect 17,267 plasma cells and 57,719 immune cells, discovering patient-specific plasma cell profiles and immune cell expression changes. Patients with the same genetic alterations tend to have both plasma cells and immune cells clustered together. By integrating bulk genomics and single cell mapping, we track plasma cell subpopulations across disease stages and find three patterns: stability (from precancer to diagnosis), and gain or loss (from diagnosis to relapse). In multiple patients, we detect "B cell-featured" plasma cell subpopulations that cluster closely with B cells, implicating their cell of origin. We validate AP-1 complex differential expression (JUN and FOS) in plasma cell subpopulations using CyTOF-based protein assays, and integrated analysis of single-cell RNA and CyTOF data reveals AP-1 downstream targets (IL6 and IL1B) potentially leading to inflammation regulation. Our work represents a longitudinal investigation for tumor and microenvironment during MM progression and paves the way for expanding treatment options.
Assuntos
Linfócitos B/metabolismo , Regulação Neoplásica da Expressão Gênica/genética , Mieloma Múltiplo/genética , Mieloma Múltiplo/imunologia , Recidiva Local de Neoplasia/genética , Microambiente Tumoral/imunologia , Idoso , Linfócitos B/citologia , Linfócitos B/imunologia , Linhagem da Célula , Evolução Clonal/genética , Estudos de Coortes , Progressão da Doença , Feminino , Regulação Neoplásica da Expressão Gênica/imunologia , Haplótipos , Humanos , Interleucina-1beta/sangue , Interleucina-6/sangue , Masculino , Espectrometria de Massas , Pessoa de Meia-Idade , Família Multigênica , Mieloma Múltiplo/sangue , Mieloma Múltiplo/patologia , Mutação , Recidiva Local de Neoplasia/sangue , Recidiva Local de Neoplasia/imunologia , Proteínas Proto-Oncogênicas c-fos/sangue , Proteínas Proto-Oncogênicas c-jun/sangue , RNA-Seq , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Análise de Célula ÚnicaRESUMO
Multiple myeloma is a plasma cell blood cancer with frequent chromosomal translocations leading to gene fusions. To determine the clinical relevance of fusion events, we detect gene fusions from a cohort of 742 patients from the Multiple Myeloma Research Foundation CoMMpass Study. Patients with multiple clinic visits enable us to track tumor and fusion evolution, and cases with matching peripheral blood and bone marrow samples allow us to evaluate the concordance of fusion calls in patients with high tumor burden. We examine the joint upregulation of WHSC1 and FGFR3 in samples with t(4;14)-related fusions, and we illustrate a method for detecting fusions from single cell RNA-seq. We report fusions at MYC and a neighboring gene, PVT1, which are related to MYC translocations and associated with divergent progression-free survival patterns. Finally, we find that 4% of patients may be eligible for targeted fusion therapies, including three with an NTRK1 fusion.
Assuntos
Fusão Gênica/genética , Histona-Lisina N-Metiltransferase/genética , Mieloma Múltiplo/genética , Proteínas Proto-Oncogênicas c-myc/genética , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/genética , Proteínas Repressoras/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Variações do Número de Cópias de DNA/genética , Perfilação da Expressão Gênica/métodos , Histona-Lisina N-Metiltransferase/biossíntese , Humanos , Imunoglobulinas/genética , Pessoa de Meia-Idade , Intervalo Livre de Progressão , RNA Longo não Codificante/genética , RNA-Seq/métodos , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/biossíntese , Receptor trkA/genética , Proteínas Repressoras/biossínteseRESUMO
Non-coding mutations can create splice sites, however the true extent of how such somatic non-coding mutations affect RNA splicing are largely unexplored. Here we use the MiSplice pipeline to analyze 783 cancer cases with WGS data and 9494 cases with WES data, discovering 562 non-coding mutations that lead to splicing alterations. Notably, most of these mutations create new exons. Introns associated with new exon creation are significantly larger than the genome-wide average intron size. We find that some mutation-induced splicing alterations are located in genes important in tumorigenesis (ATRX, BCOR, CDKN2B, MAP3K1, MAP3K4, MDM2, SMAD4, STK11, TP53 etc.), often leading to truncated proteins and affecting gene expression. The pattern emerging from these exon-creating mutations suggests that splice sites created by non-coding mutations interact with pre-existing potential splice sites that originally lacked a suitable splicing pair to induce new exon formation. Our study suggests the importance of investigating biological and clinical consequences of noncoding splice-inducing mutations that were previously neglected by conventional annotation pipelines. MiSplice will be useful for automatically annotating the splicing impact of coding and non-coding mutations in future large-scale analyses.
Assuntos
Neoplasias/genética , Precursores de RNA/genética , Sítios de Splice de RNA , Splicing de RNA , Quinases Proteína-Quinases Ativadas por AMP , Inibidor de Quinase Dependente de Ciclina p15/genética , Inibidor de Quinase Dependente de Ciclina p15/metabolismo , Bases de Dados Genéticas , Éxons , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Íntrons , MAP Quinase Quinase Quinase 1/genética , MAP Quinase Quinase Quinase 1/metabolismo , MAP Quinase Quinase Quinase 4/genética , MAP Quinase Quinase Quinase 4/metabolismo , Mutação , Neoplasias/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-mdm2/genética , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , RNA não Traduzido , RNA-Seq , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Proteína Smad4/genética , Proteína Smad4/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Sequenciamento do Exoma , Proteína Nuclear Ligada ao X/genética , Proteína Nuclear Ligada ao X/metabolismoRESUMO
Mitochondria are essential cellular organelles that play critical roles in cancer. Here, as part of the International Cancer Genome Consortium/The Cancer Genome Atlas Pan-Cancer Analysis of Whole Genomes Consortium, which aggregated whole-genome sequencing data from 2,658 cancers across 38 tumor types, we performed a multidimensional, integrated characterization of mitochondrial genomes and related RNA sequencing data. Our analysis presents the most definitive mutational landscape of mitochondrial genomes and identifies several hypermutated cases. Truncating mutations are markedly enriched in kidney, colorectal and thyroid cancers, suggesting oncogenic effects with the activation of signaling pathways. We find frequent somatic nuclear transfers of mitochondrial DNA, some of which disrupt therapeutic target genes. Mitochondrial copy number varies greatly within and across cancers and correlates with clinical variables. Co-expression analysis highlights the function of mitochondrial genes in oxidative phosphorylation, DNA repair and the cell cycle, and shows their connections with clinically actionable genes. Our study lays a foundation for translating mitochondrial biology into clinical applications.
Assuntos
Variações do Número de Cópias de DNA , Genoma Humano/genética , Genoma Mitocondrial/genética , Neoplasias/genética , Sequenciamento Completo do Genoma , Ciclo Celular/genética , Reparo do DNA/genética , DNA Mitocondrial/genética , Humanos , Mutação , Fosforilação Oxidativa , Análise de Sequência de RNARESUMO
Activation of PI3K signaling is frequently observed in triple-negative breast cancer (TNBC), yet PI3K inhibitors have shown limited clinical activity. To investigate intrinsic and adaptive mechanisms of resistance, we analyzed a panel of patient-derived xenograft models of TNBC with varying responsiveness to buparlisib, a pan-PI3K inhibitor. In a subset of patient-derived xenografts, resistance was associated with incomplete inhibition of PI3K signaling and upregulated MAPK/MEK signaling in response to buparlisib. Outlier phosphoproteome and kinome analyses identified novel candidates functionally important to buparlisib resistance, including NEK9 and MAP2K4. Knockdown of NEK9 or MAP2K4 reduced both baseline and feedback MAPK/MEK signaling and showed synthetic lethality with buparlisib in vitro A complex in/del frameshift in PIK3CA decreased sensitivity to buparlisib via NEK9/MAP2K4-dependent mechanisms. In summary, our study supports a role for NEK9 and MAP2K4 in mediating buparlisib resistance and demonstrates the value of unbiased omic analyses in uncovering resistance mechanisms to targeted therapy.Significance: Integrative phosphoproteogenomic analysis is used to determine intrinsic resistance mechanisms of triple-negative breast tumors to PI3K inhibition. Cancer Res; 78(10); 2732-46. ©2018 AACR.
Assuntos
Aminopiridinas/farmacologia , Antineoplásicos/farmacologia , Classe I de Fosfatidilinositol 3-Quinases/antagonistas & inibidores , MAP Quinase Quinase 4/genética , Morfolinas/farmacologia , Quinases Relacionadas a NIMA/genética , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Animais , Linhagem Celular Tumoral , Classe I de Fosfatidilinositol 3-Quinases/genética , Feminino , Humanos , Espectrometria de Massas , Camundongos , Proteômica/métodos , Interferência de RNA , RNA Interferente Pequeno/genética , Transdução de Sinais/genética , Neoplasias de Mama Triplo Negativas/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Bacteria have shown a remarkable ability to overcome drug therapy if there is a failure to achieve sustained bactericidal concentration or if there is a reduction in activity in situ. The latter can be caused by localized acidity, a phenomenon that can occur as a result of the combined actions of bacterial metabolism and the host immune response. Nanoparticles (NP) have shown promise in treating bacterial infections, but a significant challenge has been to develop antibacterial NPs that may be suitable for systemic administration. Herein we develop drug-encapsulated, pH-responsive, surface charge-switching poly(D,L-lactic-co-glycolic acid)-b-poly(L-histidine)-b-poly(ethylene glycol) (PLGA-PLH-PEG) nanoparticles for treating bacterial infections. These NP drug carriers are designed to shield nontarget interactions at pH 7.4 but bind avidly to bacteria in acidity, delivering drugs and mitigating in part the loss of drug activity with declining pH. The mechanism involves pH-sensitive NP surface charge switching, which is achieved by selective protonation of the imidazole groups of PLH at low pH. NP binding studies demonstrate pH-sensitive NP binding to bacteria with a 3.5 ± 0.2- to 5.8 ± 0.1-fold increase in binding to bacteria at pH 6.0 compared to 7.4. Further, PLGA-PLH-PEG-encapsulated vancomycin demonstrates reduced loss of efficacy at low pH, with an increase in minimum inhibitory concentration of 1.3-fold as compared to 2.0-fold and 2.3-fold for free and PLGA-PEG-encapsulated vancomycin, respectively. The PLGA-PLH-PEG NPs described herein are a first step toward developing systemically administered drug carriers that can target and potentially treat Gram-positive, Gram-negative, or polymicrobial infections associated with acidity.