272 results on '"John C. Marioni"'
Search Results
2. The dynamic genetic determinants of increased transcriptional divergence in spermatids
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Jasper Panten, Tobias Heinen, Christina Ernst, Nils Eling, Rebecca E. Wagner, Maja Satorius, John C. Marioni, Oliver Stegle, and Duncan T. Odom
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Science - Abstract
Abstract Cis-genetic effects are key determinants of transcriptional divergence in discrete tissues and cell types. However, how cis- and trans-effects act across continuous trajectories of cellular differentiation in vivo is poorly understood. Here, we quantify allele-specific expression during spermatogenic differentiation at single-cell resolution in an F1 hybrid mouse system, allowing for the comprehensive characterisation of cis- and trans-genetic effects, including their dynamics across cellular differentiation. Collectively, almost half of the genes subject to genetic regulation show evidence for dynamic cis-effects that vary during differentiation. Our system also allows us to robustly identify dynamic trans-effects, which are less pervasive than cis-effects. In aggregate, genetic effects were strongest in round spermatids, which parallels their increased transcriptional divergence we identified between species. Our approach provides a comprehensive quantification of the variability of genetic effects in vivo, and demonstrates a widely applicable strategy to dissect the impact of regulatory variants on gene regulation in dynamic systems.
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- 2024
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3. scSNV-seq: high-throughput phenotyping of single nucleotide variants by coupled single-cell genotyping and transcriptomics
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Sarah E. Cooper, Matthew A. Coelho, Magdalena E. Strauss, Aleksander M. Gontarczyk, Qianxin Wu, Mathew J. Garnett, John C. Marioni, and Andrew R. Bassett
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Single-cell CRISPR screen ,SNV ,GWAS ,Base editor ,Causal variant ,VUS ,Biology (General) ,QH301-705.5 ,Genetics ,QH426-470 - Abstract
Abstract CRISPR screens with single-cell transcriptomic readouts are a valuable tool to understand the effect of genetic perturbations including single nucleotide variants (SNVs) associated with diseases. Interpretation of these data is currently limited as genotypes cannot be accurately inferred from guide RNA identity alone. scSNV-seq overcomes this limitation by coupling single-cell genotyping and transcriptomics of the same cells enabling accurate and high-throughput screening of SNVs. Analysis of variants across the JAK1 gene with scSNV-seq demonstrates the importance of determining the precise genetic perturbation and accurately classifies clinically observed missense variants into three functional categories: benign, loss of function, and separation of function.
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- 2024
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4. BASiCS workflow: a step-by-step analysis of expression variability using single cell RNA sequencing data [version 2; peer review: 1 approved, 2 approved with reservations]
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Catalina A. Vallejos, Nils Eling, Alan O'Callaghan, and John C. Marioni
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single-cell RNA sequencing ,expression variability ,transcriptional noise ,differential expression testing ,scRNAseq ,Bayesian ,eng ,Medicine ,Science - Abstract
Cell-to-cell gene expression variability is an inherent feature of complex biological systems, such as immunity and development. Single-cell RNA sequencing is a powerful tool to quantify this heterogeneity, but it is prone to strong technical noise. In this article, we describe a step-by-step computational workflow that uses the BASiCS Bioconductor package to robustly quantify expression variability within and between known groups of cells (such as experimental conditions or cell types). BASiCS uses an integrated framework for data normalisation, technical noise quantification and downstream analyses, propagating statistical uncertainty across these steps. Within a single seemingly homogeneous cell population, BASiCS can identify highly variable genes that exhibit strong heterogeneity as well as lowly variable genes with stable expression. BASiCS also uses a probabilistic decision rule to identify changes in expression variability between cell populations, whilst avoiding confounding effects related to differences in technical noise or in overall abundance. Using a publicly available dataset, we guide users through a complete pipeline that includes preliminary steps for quality control, as well as data exploration using the scater and scran Bioconductor packages. The workflow is accompanied by a Docker image that ensures the reproducibility of our results.
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- 2024
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5. Time space and single-cell resolved tissue lineage trajectories and laterality of body plan at gastrulation
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Ran Wang, Xianfa Yang, Jiehui Chen, Lin Zhang, Jonathan A. Griffiths, Guizhong Cui, Yingying Chen, Yun Qian, Guangdun Peng, Jinsong Li, Liantang Wang, John C. Marioni, Patrick P. L. Tam, and Naihe Jing
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Science - Abstract
Abstract Understanding of the molecular drivers of lineage diversification and tissue patterning during primary germ layer development requires in-depth knowledge of the dynamic molecular trajectories of cell lineages across a series of developmental stages of gastrulation. Through computational modeling, we constructed at single-cell resolution, a spatio-temporal transcriptome of cell populations in the germ-layers of gastrula-stage mouse embryos. This molecular atlas enables the inference of molecular network activity underpinning the specification and differentiation of the germ-layer tissue lineages. Heterogeneity analysis of cellular composition at defined positions in the epiblast revealed progressive diversification of cell types. The single-cell transcriptome revealed an enhanced BMP signaling activity in the right-side mesoderm of late-gastrulation embryo. Perturbation of asymmetric BMP signaling activity at late gastrulation led to randomization of left-right molecular asymmetry in the lateral mesoderm of early-somite-stage embryo. These findings indicate the asymmetric BMP activity during gastrulation may be critical for the symmetry breaking process.
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- 2023
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6. Time-resolved single-cell RNAseq profiling identifies a novel Fabp5+ subpopulation of inflammatory myeloid cells with delayed cytotoxic profile in chronic spinal cord injury
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Regan Hamel, Luca Peruzzotti-Jametti, Katherine Ridley, Veronica Testa, Bryan Yu, David Rowitch, John C. Marioni, and Stefano Pluchino
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Single cell RNA sequencing ,Neuroinflammation ,Fatty acid binding protein 5 ,Spinal cord injury ,Fate-mapping myeloid cells ,Myeloid cells ,Science (General) ,Q1-390 ,Social sciences (General) ,H1-99 - Abstract
Traumatic spinal cord injuries (SCI) are a group of highly debilitating pathologies affecting thousands annually, and adversely affecting quality of life. Currently, no fully restorative therapies exist, and SCI still results in significant personal, societal and financial burdens. Inflammation plays a major role in the evolution of SCI, with myeloid cells, including bone marrow derived macrophages (BMDMs) and microglia (MG) being primary drivers of both early secondary pathogenesis and delayed wound healing events.The precise role of myeloid cell subsets is unclear as upon crossing the blood-spinal cord barrier, infiltrating bone marrow derived macrophages (BMDMs) may take on the morphology of resident microglia, and upregulate canonical microglia markers, thus making the two populations difficult to distinguish.Here, we used time-resolved scRNAseq and transgenic fate-mapping to chart the transcriptional profiles of tissue-resident and -infiltrating myeloid cells in a mouse model of thoracic contusion SCI.Our work identifies a novel subpopulation of foam cell-like inflammatory myeloid cells with increased expression of Fatty Acid Binding Protein 5 (Fabp5) and comprise both tissue-resident and -infiltrating cells. Fabp5+ inflammatory myeloid cells display a delayed cytotoxic profile that is predominant at the lesion epicentre and extends into the chronic phase of SCI.
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- 2023
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7. Single-cell multi-omics profiling links dynamic DNA methylation to cell fate decisions during mouse early organogenesis
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Stephen J. Clark, Ricard Argelaguet, Tim Lohoff, Felix Krueger, Deborah Drage, Berthold Göttgens, John C. Marioni, Jennifer Nichols, and Wolf Reik
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Biology (General) ,QH301-705.5 ,Genetics ,QH426-470 - Abstract
Abstract Background Perturbation of DNA methyltransferases (DNMTs) and of the active DNA demethylation pathway via ten-eleven translocation (TET) methylcytosine dioxygenases results in severe developmental defects and embryonic lethality. Dynamic control of DNA methylation is therefore vital for embryogenesis, yet the underlying mechanisms remain poorly understood. Results Here we report a single-cell transcriptomic atlas from Dnmt and Tet mutant mouse embryos during early organogenesis. We show that both the maintenance and de novo methyltransferase enzymes are dispensable for the formation of all major cell types at E8.5. However, DNA methyltransferases are required for silencing of prior or alternative cell fates such as pluripotency and extraembryonic programmes. Deletion of all three TET enzymes produces substantial lineage biases, in particular, a failure to generate primitive erythrocytes. Single-cell multi-omics profiling moreover reveals that this is linked to a failure to demethylate distal regulatory elements in Tet triple-knockout embryos. Conclusions This study provides a detailed analysis of the effects of perturbing DNA methylation on mouse organogenesis at a whole organism scale and affords new insights into the regulatory mechanisms of cell fate decisions.
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- 2022
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8. geneBasis: an iterative approach for unsupervised selection of targeted gene panels from scRNA-seq
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Alsu Missarova, Jaison Jain, Andrew Butler, Shila Ghazanfar, Tim Stuart, Maigan Brusko, Clive Wasserfall, Harry Nick, Todd Brusko, Mark Atkinson, Rahul Satija, and John C. Marioni
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Biology (General) ,QH301-705.5 ,Genetics ,QH426-470 - Abstract
Abstract scRNA-seq datasets are increasingly used to identify gene panels that can be probed using alternative technologies, such as spatial transcriptomics, where choosing the best subset of genes is vital. Existing methods are limited by a reliance on pre-existing cell type labels or by difficulties in identifying markers of rare cells. We introduce an iterative approach, geneBasis, for selecting an optimal gene panel, where each newly added gene captures the maximum distance between the true manifold and the manifold constructed using the currently selected gene panel. Our approach outperforms existing strategies and can resolve cell types and subtle cell state differences.
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- 2021
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9. Coordinated changes in gene expression kinetics underlie both mouse and human erythroid maturation
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Melania Barile, Ivan Imaz-Rosshandler, Isabella Inzani, Shila Ghazanfar, Jennifer Nichols, John C. Marioni, Carolina Guibentif, and Berthold Göttgens
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RNA velocity ,Gastrulation ,Erythropoiesis ,Gata1 ,Biology (General) ,QH301-705.5 ,Genetics ,QH426-470 - Abstract
Abstract Background Single-cell technologies are transforming biomedical research, including the recent demonstration that unspliced pre-mRNA present in single-cell RNA-Seq permits prediction of future expression states. Here we apply this RNA velocity concept to an extended timecourse dataset covering mouse gastrulation and early organogenesis. Results Intriguingly, RNA velocity correctly identifies epiblast cells as the starting point, but several trajectory predictions at later stages are inconsistent with both real-time ordering and existing knowledge. The most striking discrepancy concerns red blood cell maturation, with velocity-inferred trajectories opposing the true differentiation path. Investigating the underlying causes reveals a group of genes with a coordinated step-change in transcription, thus violating the assumptions behind current velocity analysis suites, which do not accommodate time-dependent changes in expression dynamics. Using scRNA-Seq analysis of chimeric mouse embryos lacking the major erythroid regulator Gata1, we show that genes with the step-changes in expression dynamics during erythroid differentiation fail to be upregulated in the mutant cells, thus underscoring the coordination of modulating transcription rate along a differentiation trajectory. In addition to the expected block in erythroid maturation, the Gata1-chimera dataset reveals induction of PU.1 and expansion of megakaryocyte progenitors. Finally, we show that erythropoiesis in human fetal liver is similarly characterized by a coordinated step-change in gene expression. Conclusions By identifying a limitation of the current velocity framework coupled with in vivo analysis of mutant cells, we reveal a coordinated step-change in gene expression kinetics during erythropoiesis, with likely implications for many other differentiation processes.
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- 2021
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10. Time-resolved single-cell analysis of Brca1 associated mammary tumourigenesis reveals aberrant differentiation of luminal progenitors
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Karsten Bach, Sara Pensa, Marija Zarocsinceva, Katarzyna Kania, Julie Stockis, Silvain Pinaud, Kyren A. Lazarus, Mona Shehata, Bruno M. Simões, Alice R. Greenhalgh, Sacha J. Howell, Robert B. Clarke, Carlos Caldas, Timotheus Y. F. Halim, John C. Marioni, and Walid T. Khaled
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Science - Abstract
BRCA1 driven breast cancer arises from luminal progenitor cells but how BRCA1 loss-of-function affects the luminal progenitor cell state during premalignant stages of the disease is still unclear. Here, the authors demonstrate an aberrant differentiation of luminal progenitors towards a partial secretory luminal cell phenotype that occurs in a Brca1 deficient mouse model of breast cancer at early stages of tumour initiation and in breast cells from BRCA1 carriers.
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- 2021
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11. Coagulation factor V is a T-cell inhibitor expressed by leukocytes in COVID-19
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Jun Wang, Prasanti Kotagiri, Paul A. Lyons, Rafia S. Al-Lamki, Federica Mescia, Laura Bergamaschi, Lorinda Turner, Michael D. Morgan, Fernando J. Calero-Nieto, Karsten Bach, Nicole Mende, Nicola K. Wilson, Emily R. Watts, Patrick H. Maxwell, Patrick F. Chinnery, Nathalie Kingston, Sofia Papadia, Kathleen E. Stirrups, Neil Walker, Ravindra K. Gupta, David K. Menon, Kieren Allinson, Sarah J. Aitken, Mark Toshner, Michael P. Weekes, James A. Nathan, Sarah R. Walmsley, Willem H. Ouwehand, Mary Kasanicki, Berthold Göttgens, John C. Marioni, Kenneth G.C. Smith, Jordan S. Pober, and John R. Bradley
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Immunology ,Microbiology ,Omics ,Transcriptomics ,Science - Abstract
Summary: Clotting Factor V (FV) is primarily synthesized in the liver and when cleaved by thrombin forms pro-coagulant Factor Va (FVa). Using whole blood RNAseq and scRNAseq of peripheral blood mononuclear cells, we find that FV mRNA is expressed in leukocytes, and identify neutrophils, monocytes, and T regulatory cells as sources of increased FV in hospitalized patients with COVID-19. Proteomic analysis confirms increased FV in circulating neutrophils in severe COVID-19, and immunofluorescence microscopy identifies FV in lung-infiltrating leukocytes in COVID-19 lung disease. Increased leukocyte FV expression in severe disease correlates with T-cell lymphopenia. Both plasma-derived and a cleavage resistant recombinant FV, but not thrombin cleaved FVa, suppress T-cell proliferation in vitro. Anticoagulants that reduce FV conversion to FVa, including heparin, may have the unintended consequence of suppressing the adaptive immune system.
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- 2022
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12. MOFA+: a statistical framework for comprehensive integration of multi-modal single-cell data
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Ricard Argelaguet, Damien Arnol, Danila Bredikhin, Yonatan Deloro, Britta Velten, John C. Marioni, and Oliver Stegle
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Single cell ,Multi-omics ,Data integration ,Factor analysis ,Biology (General) ,QH301-705.5 ,Genetics ,QH426-470 - Abstract
Abstract Technological advances have enabled the profiling of multiple molecular layers at single-cell resolution, assaying cells from multiple samples or conditions. Consequently, there is a growing need for computational strategies to analyze data from complex experimental designs that include multiple data modalities and multiple groups of samples. We present Multi-Omics Factor Analysis v2 (MOFA+), a statistical framework for the comprehensive and scalable integration of single-cell multi-modal data. MOFA+ reconstructs a low-dimensional representation of the data using computationally efficient variational inference and supports flexible sparsity constraints, allowing to jointly model variation across multiple sample groups and data modalities.
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- 2020
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13. A high-content RNAi screen reveals multiple roles for long noncoding RNAs in cell division
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Lovorka Stojic, Aaron T. L. Lun, Patrice Mascalchi, Christina Ernst, Aisling M. Redmond, Jasmin Mangei, Alexis R. Barr, Vicky Bousgouni, Chris Bakal, John C. Marioni, Duncan T. Odom, and Fanni Gergely
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Science - Abstract
Long noncoding RNAs (lncRNAs) regulate key steps of cell division. Here, the authors perform a comprehensive RNAi imaging screen targeting more than 2,000 human lncRNAs, and suggest a role of chromatin-associated linc00899 in regulation of cell division by suppressing the transcription of microtubule-binding protein TPPP/p25.
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- 2020
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14. Eleven grand challenges in single-cell data science
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David Lähnemann, Johannes Köster, Ewa Szczurek, Davis J. McCarthy, Stephanie C. Hicks, Mark D. Robinson, Catalina A. Vallejos, Kieran R. Campbell, Niko Beerenwinkel, Ahmed Mahfouz, Luca Pinello, Pavel Skums, Alexandros Stamatakis, Camille Stephan-Otto Attolini, Samuel Aparicio, Jasmijn Baaijens, Marleen Balvert, Buys de Barbanson, Antonio Cappuccio, Giacomo Corleone, Bas E. Dutilh, Maria Florescu, Victor Guryev, Rens Holmer, Katharina Jahn, Thamar Jessurun Lobo, Emma M. Keizer, Indu Khatri, Szymon M. Kielbasa, Jan O. Korbel, Alexey M. Kozlov, Tzu-Hao Kuo, Boudewijn P.F. Lelieveldt, Ion I. Mandoiu, John C. Marioni, Tobias Marschall, Felix Mölder, Amir Niknejad, Alicja Rączkowska, Marcel Reinders, Jeroen de Ridder, Antoine-Emmanuel Saliba, Antonios Somarakis, Oliver Stegle, Fabian J. Theis, Huan Yang, Alex Zelikovsky, Alice C. McHardy, Benjamin J. Raphael, Sohrab P. Shah, and Alexander Schönhuth
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Biology (General) ,QH301-705.5 ,Genetics ,QH426-470 - Abstract
Abstract The recent boom in microfluidics and combinatorial indexing strategies, combined with low sequencing costs, has empowered single-cell sequencing technology. Thousands—or even millions—of cells analyzed in a single experiment amount to a data revolution in single-cell biology and pose unique data science problems. Here, we outline eleven challenges that will be central to bringing this emerging field of single-cell data science forward. For each challenge, we highlight motivating research questions, review prior work, and formulate open problems. This compendium is for established researchers, newcomers, and students alike, highlighting interesting and rewarding problems for the coming years.
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- 2020
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15. Single-cell RNA-sequencing of differentiating iPS cells reveals dynamic genetic effects on gene expression
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Anna S. E. Cuomo, Daniel D. Seaton, Davis J. McCarthy, Iker Martinez, Marc Jan Bonder, Jose Garcia-Bernardo, Shradha Amatya, Pedro Madrigal, Abigail Isaacson, Florian Buettner, Andrew Knights, Kedar Nath Natarajan, HipSci Consortium, Ludovic Vallier, John C. Marioni, Mariya Chhatriwala, and Oliver Stegle
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Science - Abstract
Studying the genetic effects on early stages of human development is challenging due to a scarcity of biological material. Here, the authors utilise induced pluripotent stem cells from 125 donors to track gene expression changes and expression quantitative trait loci at single cell resolution during in vitro endoderm differentiation.
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- 2020
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16. Staged developmental mapping and X chromosome transcriptional dynamics during mouse spermatogenesis
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Christina Ernst, Nils Eling, Celia P. Martinez-Jimenez, John C. Marioni, and Duncan T. Odom
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Science - Abstract
The transcriptional regulation of murine spermatogenesis is not well understood. Here, the authors use single-cell and bulk RNA-Sequencing of juvenile and adult mice to characterise somatic and germ cell development, and chromatin profile the X chromosome to show that spermatid-specific genes are repressed by H3K9me3 during meiosis.
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- 2019
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17. EmptyDrops: distinguishing cells from empty droplets in droplet-based single-cell RNA sequencing data
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Aaron T. L. Lun, Samantha Riesenfeld, Tallulah Andrews, The Phuong Dao, Tomas Gomes, participants in the 1st Human Cell Atlas Jamboree, and John C. Marioni
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Single-cell transcriptomics ,Droplet-based protocols ,Empty droplets ,Cell detection ,Biology (General) ,QH301-705.5 ,Genetics ,QH426-470 - Abstract
Abstract Droplet-based single-cell RNA sequencing protocols have dramatically increased the throughput of single-cell transcriptomics studies. A key computational challenge when processing these data is to distinguish libraries for real cells from empty droplets. Here, we describe a new statistical method for calling cells from droplet-based data, based on detecting significant deviations from the expression profile of the ambient solution. Using simulations, we demonstrate that EmptyDrops has greater power than existing approaches while controlling the false discovery rate among detected cells. Our method also retains distinct cell types that would have been discarded by existing methods in several real data sets.
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- 2019
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18. SRPK1 maintains acute myeloid leukemia through effects on isoform usage of epigenetic regulators including BRD4
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Konstantinos Tzelepis, Etienne De Braekeleer, Demetrios Aspris, Isaia Barbieri, M. S. Vijayabaskar, Wen-Hsin Liu, Malgorzata Gozdecka, Emmanouil Metzakopian, Hamish D. Toop, Monika Dudek, Samuel C. Robson, Francisco Hermida-Prado, Yu Hsuen Yang, Roya Babaei-Jadidi, Dimitrios A. Garyfallos, Hannes Ponstingl, Joao M. L. Dias, Paolo Gallipoli, Michael Seiler, Silvia Buonamici, Binje Vick, Andrew J. Bannister, Roland Rad, Rab K. Prinjha, John C. Marioni, Brian Huntly, Jennifer Batson, Jonathan C. Morris, Cristina Pina, Allan Bradley, Irmela Jeremias, David O. Bates, Kosuke Yusa, Tony Kouzarides, and George S. Vassiliou
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Science - Abstract
SRPK1, a kinase involved in splicing regulation, is a potential therapeutic target for AML patients. Here, the authors show that SRPK1 inhibition changes isoform levels of key epigenetic regulators, including BRD4, and it has anti-tumor effects specifically against MLL-rearranged AML cells.
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- 2018
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19. CTCF maintains regulatory homeostasis of cancer pathways
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Sarah J. Aitken, Ximena Ibarra-Soria, Elissavet Kentepozidou, Paul Flicek, Christine Feig, John C. Marioni, and Duncan T. Odom
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CTCF ,Transcription ,Hemizygosity ,Cancer ,Chromatin state ,Chromatin architecture ,Biology (General) ,QH301-705.5 ,Genetics ,QH426-470 - Abstract
Abstract Background CTCF binding to DNA helps partition the mammalian genome into discrete structural and regulatory domains. Complete removal of CTCF from mammalian cells causes catastrophic genome dysregulation, likely due to widespread collapse of 3D chromatin looping and alterations to inter- and intra-TAD interactions within the nucleus. In contrast, Ctcf hemizygous mice with lifelong reduction of CTCF expression are viable, albeit with increased cancer incidence. Here, we exploit chronic Ctcf hemizygosity to reveal its homeostatic roles in maintaining genome function and integrity. Results We find that Ctcf hemizygous cells show modest but robust changes in almost a thousand sites of genomic CTCF occupancy; these are enriched for lower affinity binding events with weaker evolutionary conservation across the mouse lineage. Furthermore, we observe dysregulation of the expression of several hundred genes, which are concentrated in cancer-related pathways, and are caused by changes in transcriptional regulation. Chromatin structure is preserved but some loop interactions are destabilized; these are often found around differentially expressed genes and their enhancers. Importantly, the transcriptional alterations identified in vitro are recapitulated in mouse tumors and also in human cancers. Conclusions This multi-dimensional genomic and epigenomic profiling of a Ctcf hemizygous mouse model system shows that chronic depletion of CTCF dysregulates steady-state gene expression by subtly altering transcriptional regulation, changes which can also be observed in primary tumors.
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- 2018
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20. Single-Cell Transcriptomics Uncovers Zonation of Function in the Mesenchyme during Liver Fibrosis
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Ross Dobie, John R. Wilson-Kanamori, Beth E.P. Henderson, James R. Smith, Kylie P. Matchett, Jordan R. Portman, Karolina Wallenborg, Simone Picelli, Anna Zagorska, Swetha V. Pendem, Thomas E. Hudson, Minnie M. Wu, Grant R. Budas, David G. Breckenridge, Ewen M. Harrison, Damian J. Mole, Stephen J. Wigmore, Prakash Ramachandran, Chris P. Ponting, Sarah A. Teichmann, John C. Marioni, and Neil C. Henderson
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Biology (General) ,QH301-705.5 - Abstract
Summary: Iterative liver injury results in progressive fibrosis disrupting hepatic architecture, regeneration potential, and liver function. Hepatic stellate cells (HSCs) are a major source of pathological matrix during fibrosis and are thought to be a functionally homogeneous population. Here, we use single-cell RNA sequencing to deconvolve the hepatic mesenchyme in healthy and fibrotic mouse liver, revealing spatial zonation of HSCs across the hepatic lobule. Furthermore, we show that HSCs partition into topographically diametric lobule regions, designated portal vein-associated HSCs (PaHSCs) and central vein-associated HSCs (CaHSCs). Importantly we uncover functional zonation, identifying CaHSCs as the dominant pathogenic collagen-producing cells in a mouse model of centrilobular fibrosis. Finally, we identify LPAR1 as a therapeutic target on collagen-producing CaHSCs, demonstrating that blockade of LPAR1 inhibits liver fibrosis in a rodent NASH model. Taken together, our work illustrates the power of single-cell transcriptomics to resolve the key collagen-producing cells driving liver fibrosis with high precision. : Dobie et al. use scRNA-seq to reveal spatial and functional zonation of hepatic stellate cells (HSCs) across the hepatic lobule, identifying central vein-associated HSCs as the dominant pathogenic collagen-producing cells during centrilobular injury-induced fibrosis. This illustrates the power of scRNA-seq to resolve the key collagen-producing cells driving liver fibrosis. Keywords: liver fibrosis, mesenchyme, hepatic stellate cells, single-cell RNA sequencing, zonation
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- 2019
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21. Detection and removal of barcode swapping in single-cell RNA-seq data
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Jonathan A. Griffiths, Arianne C. Richard, Karsten Bach, Aaron T. L. Lun, and John C. Marioni
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Science - Abstract
DNA barcode swapping results in mislabelling of sequencing reads between multiplexed samples. Here, the authors investigate the severity and consequences of barcode swapping for single-cell RNA-seq data, and develop a computational method to exclude swapped reads.
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- 2018
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22. CpG island composition differences are a source of gene expression noise indicative of promoter responsiveness
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Michael D. Morgan and John C. Marioni
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Gene expression noise ,Single cell ,Promoter response ,Biology (General) ,QH301-705.5 ,Genetics ,QH426-470 - Abstract
Abstract Background Population phenotypic variation can arise from genetic differences between individuals, or from cellular heterogeneity in an isogenic group of cells or organisms. The emergence of gene expression differences between genetically identical cells is referred to as gene expression noise, the sources of which are not well understood. Results In this work, by studying gene expression noise between multiple cell lineages and mammalian species, we find consistent evidence of a role for CpG islands as sources of gene expression noise. Variation in noise among CpG island promoters can be partially attributed to differences in island size, in which short islands have noisier gene expression. Building on these findings, we investigate the potential for short CpG islands to act as fast response elements to environmental stimuli. Specifically, we find that these islands are enriched amongst primary response genes in SWI/SNF-independent stimuli, suggesting that expression noise is an indicator of promoter responsiveness. Conclusions Thus, through the integration of single-cell RNA expression profiling, chromatin landscape and temporal gene expression dynamics, we have uncovered a role for short CpG island promoters as fast response elements.
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- 2018
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23. scNMT-seq enables joint profiling of chromatin accessibility DNA methylation and transcription in single cells
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Stephen J. Clark, Ricard Argelaguet, Chantriolnt-Andreas Kapourani, Thomas M. Stubbs, Heather J. Lee, Celia Alda-Catalinas, Felix Krueger, Guido Sanguinetti, Gavin Kelsey, John C. Marioni, Oliver Stegle, and Wolf Reik
- Subjects
Science - Abstract
Relationships between DNA methylation and transcription, and methylation and DNA accessibility can be probed but interrogating all three in the same single cells has not been possible. Here, the authors report the first single-cell method for parallel chromatin accessibility, DNA methylation and transcriptome profiling.
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- 2018
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24. Differentiation dynamics of mammary epithelial cells revealed by single-cell RNA sequencing
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Karsten Bach, Sara Pensa, Marta Grzelak, James Hadfield, David J. Adams, John C. Marioni, and Walid T. Khaled
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Science - Abstract
There is a need to understand how mammary epithelial cells respond to changes at various developmental stages. Here, the authors use single-cell RNA sequencing of mammary epithelial cells at different adult developmental stages, identifying different cell types and charting their developmental trajectory.
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- 2017
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25. Isolation and Comparative Transcriptome Analysis of Human Fetal and iPSC-Derived Cone Photoreceptor Cells
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Emily Welby, Jorn Lakowski, Valentina Di Foggia, Dimitri Budinger, Anai Gonzalez-Cordero, Aaron T.L. Lun, Michael Epstein, Aara Patel, Elisa Cuevas, Kamil Kruczek, Arifa Naeem, Federico Minneci, Mike Hubank, David T. Jones, John C. Marioni, Robin R. Ali, and Jane C. Sowden
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Medicine (General) ,R5-920 ,Biology (General) ,QH301-705.5 - Abstract
Summary: Loss of cone photoreceptors, crucial for daylight vision, has the greatest impact on sight in retinal degeneration. Transplantation of stem cell-derived L/M-opsin cones, which form 90% of the human cone population, could provide a feasible therapy to restore vision. However, transcriptomic similarities between fetal and stem cell-derived cones remain to be defined, in addition to development of cone cell purification strategies. Here, we report an analysis of the human L/M-opsin cone photoreceptor transcriptome using an AAV2/9.pR2.1:GFP reporter. This led to the identification of a cone-enriched gene signature, which we used to demonstrate similar gene expression between fetal and stem cell-derived cones. We then defined a cluster of differentiation marker combination that, when used for cell sorting, significantly enriches for cone photoreceptors from the fetal retina and stem cell-derived retinal organoids, respectively. These data may facilitate more efficient isolation of human stem cell-derived cones for use in clinical transplantation studies. : Welby et al. define a cone-enriched gene signature within a human fetal L/M-opsin cone population, which is used as a baseline reference to demonstrate similar cone gene expression between bona fide and stem cell-derived L/M-opsin cone cells. Furthermore, profiling of cell surface molecules in human fetal cones led to the generation of a cluster of differentiation marker panel, which provides enrichment of fetal and stem cell-derived cones. Keywords: retinal dystrophies, cone photoreceptor cells, transcriptome, human pluripotent stem cells, retinal organoids, cell surface markers, cell transplantation therapy
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- 2017
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26. f-scLVM: scalable and versatile factor analysis for single-cell RNA-seq
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Florian Buettner, Naruemon Pratanwanich, Davis J. McCarthy, John C. Marioni, and Oliver Stegle
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Single-cell RNA-seq ,Sparse factor analysis ,Gene set annotations ,Biology (General) ,QH301-705.5 ,Genetics ,QH426-470 - Abstract
Abstract Single-cell RNA-sequencing (scRNA-seq) allows studying heterogeneity in gene expression in large cell populations. Such heterogeneity can arise due to technical or biological factors, making decomposing sources of variation difficult. We here describe f-scLVM (factorial single-cell latent variable model), a method based on factor analysis that uses pathway annotations to guide the inference of interpretable factors underpinning the heterogeneity. Our model jointly estimates the relevance of individual factors, refines gene set annotations, and infers factors without annotation. In applications to multiple scRNA-seq datasets, we find that f-scLVM robustly decomposes scRNA-seq datasets into interpretable components, thereby facilitating the identification of novel subpopulations.
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- 2017
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27. Mosaic autosomal aneuploidies are detectable from single-cell RNAseq data
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Jonathan A. Griffiths, Antonio Scialdone, and John C. Marioni
- Subjects
Aneuploidy detection ,Single-cell ,Copy-number ,RNAseq ,Biotechnology ,TP248.13-248.65 ,Genetics ,QH426-470 - Abstract
Abstract Background Aneuploidies are copy number variants that affect entire chromosomes. They are seen commonly in cancer, embryonic stem cells, human embryos, and in various trisomic diseases. Aneuploidies frequently affect only a subset of cells in a sample; this is known as “mosaic” aneuploidy. A cell that harbours an aneuploidy exhibits disrupted gene expression patterns which can alter its behaviour. However, detection of aneuploidies using conventional single-cell DNA-sequencing protocols is slow and expensive. Methods We have developed a method that uses chromosome-wide expression imbalances to identify aneuploidies from single-cell RNA-seq data. The method provides quantitative aneuploidy calls, and is integrated into an R software package available on GitHub and as an Additional file of this manuscript. Results We validate our approach using data with known copy number, identifying the vast majority of aneuploidies with a low rate of false discovery. We show further support for the method’s efficacy by exploiting allele-specific gene expression levels, and differential expression analyses. Conclusions The method is quick and easy to apply, straightforward to interpret, and represents a substantial cost saving compared to single-cell genome sequencing techniques. However, the method is less well suited to data where gene expression is highly variable. The results obtained from the method can be used to investigate the consequences of aneuploidy itself, or to exclude aneuploidy-affected expression values from conventional scRNA-seq data analysis.
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- 2017
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28. Interplay of cis and trans mechanisms driving transcription factor binding and gene expression evolution
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Emily S. Wong, Bianca M. Schmitt, Anastasiya Kazachenka, David Thybert, Aisling Redmond, Frances Connor, Tim F. Rayner, Christine Feig, Anne C. Ferguson-Smith, John C. Marioni, Duncan T. Odom, and Paul Flicek
- Subjects
Science - Abstract
“Variation in the noncoding regulatory sequences in the genome plays important roles in human disease and evolution. Here, the authors use F1 mouse hybrids to shed light on the regulatory mechanisms mediating transcription factor binding, chromatin state and gene expression in mammalian cells.”
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- 2017
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29. Impact of Alternative Splicing on the Human Proteome
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Yansheng Liu, Mar Gonzàlez-Porta, Sergio Santos, Alvis Brazma, John C. Marioni, Ruedi Aebersold, Ashok R. Venkitaraman, and Vihandha O. Wickramasinghe
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Biology (General) ,QH301-705.5 - Abstract
Summary: Alternative splicing is a critical determinant of genome complexity and, by implication, is assumed to engender proteomic diversity. This notion has not been experimentally tested in a targeted, quantitative manner. Here, we have developed an integrative approach to ask whether perturbations in mRNA splicing patterns alter the composition of the proteome. We integrate RNA sequencing (RNA-seq) (to comprehensively report intron retention, differential transcript usage, and gene expression) with a data-independent acquisition (DIA) method, SWATH-MS (sequential window acquisition of all theoretical spectra-mass spectrometry), to capture an unbiased, quantitative snapshot of the impact of constitutive and alternative splicing events on the proteome. Whereas intron retention is accompanied by decreased protein abundance, alterations in differential transcript usage and gene expression alter protein abundance proportionate to transcript levels. Our findings illustrate how RNA splicing links isoform expression in the human transcriptome with proteomic diversity and provides a foundation for studying perturbations associated with human diseases. : Liu et al. have developed an integrative approach to ask whether perturbations in mRNA splicing patterns alter the composition of the proteome. Their findings illustrate how RNA splicing links isoform expression in the human transcriptome with proteomic diversity and provides a foundation for studying perturbations associated with human diseases. Keywords: alternative splicing, proteomics, RNA
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- 2017
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30. Single-Cell Landscape of Transcriptional Heterogeneity and Cell Fate Decisions during Mouse Early Gastrulation
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Hisham Mohammed, Irene Hernando-Herraez, Aurora Savino, Antonio Scialdone, Iain Macaulay, Carla Mulas, Tamir Chandra, Thierry Voet, Wendy Dean, Jennifer Nichols, John C. Marioni, and Wolf Reik
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Biology (General) ,QH301-705.5 - Abstract
Summary: The mouse inner cell mass (ICM) segregates into the epiblast and primitive endoderm (PrE) lineages coincident with implantation of the embryo. The epiblast subsequently undergoes considerable expansion of cell numbers prior to gastrulation. To investigate underlying regulatory principles, we performed systematic single-cell RNA sequencing (seq) of conceptuses from E3.5 to E6.5. The epiblast shows reactivation and subsequent inactivation of the X chromosome, with Zfp57 expression associated with reactivation and inactivation together with other candidate regulators. At E6.5, the transition from epiblast to primitive streak is linked with decreased expression of polycomb subunits, suggesting a key regulatory role. Notably, our analyses suggest elevated transcriptional noise at E3.5 and within the non-committed epiblast at E6.5, coinciding with exit from pluripotency. By contrast, E6.5 primitive streak cells became highly synchronized and exhibit a shortened G1 cell-cycle phase, consistent with accelerated proliferation. Our study systematically charts transcriptional noise and uncovers molecular processes associated with early lineage decisions. : Mohammed et al. chart mouse embryonic development from implantation to early gastrulation at single-cell resolution. They describe regulatory processes associated with lineage commitment. An increased level of transcriptional noise is observed prior to lineage commitment, an observation that provides fresh insights into cell fate decision-making processes. Keywords: gastrulation, embryo, single-cell RNA-seq, epiblast, primitive endoderm, primitive streak, X-chromosome, transcriptional noise
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- 2017
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31. Flipping between Polycomb repressed and active transcriptional states introduces noise in gene expression
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Gozde Kar, Jong Kyoung Kim, Aleksandra A. Kolodziejczyk, Kedar Nath Natarajan, Elena Torlai Triglia, Borbala Mifsud, Sarah Elderkin, John C. Marioni, Ana Pombo, and Sarah A. Teichmann
- Subjects
Science - Abstract
Polycomb repressive complexes modify histones but it is unclear how changes in chromatin states alter kinetics of transcription. Here, the authors use single-cell RNAseq and ChIPseq to find that actively transcribed genes with Polycomb marks have greater cell-to-cell variation in expression.
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- 2017
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32. Maturing Human CD127+ CCR7+ PDL1+ Dendritic Cells Express AIRE in the Absence of Tissue Restricted Antigens
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Joannah R. Fergusson, Michael D. Morgan, Melanie Bruchard, Leonie Huitema, Balthasar A. Heesters, Vincent van Unen, Jan Piet van Hamburg, Nicole N. van der Wel, Daisy Picavet, Frits Koning, Sander W. Tas, Mark S. Anderson, John C. Marioni, Georg A. Holländer, and Hergen Spits
- Subjects
dendritic cells ,AIRE ,PDL1 ,maturation ,tissue restricted antigen ,Immunologic diseases. Allergy ,RC581-607 - Abstract
Expression of the Autoimmune regulator (AIRE) outside of the thymus has long been suggested in both humans and mice, but the cellular source in humans has remained undefined. Here we identify AIRE expression in human tonsils and extensively analyzed these “extra-thymic AIRE expressing cells” (eTACs) using combinations of flow cytometry, CyTOF and single cell RNA-sequencing. We identified AIRE+ cells as dendritic cells (DCs) with a mature and migratory phenotype including high levels of antigen presenting molecules and costimulatory molecules, and specific expression of CD127, CCR7, and PDL1. These cells also possessed the ability to stimulate and re-stimulate T cells and displayed reduced responses to toll-like receptor (TLR) agonists compared to conventional DCs. While expression of AIRE was enriched within CCR7+CD127+ DCs, single-cell RNA sequencing revealed expression of AIRE to be transient, rather than stable, and associated with the differentiation to a mature phenotype. The role of AIRE in central tolerance induction within the thymus is well-established, however our study shows that AIRE expression within the periphery is not associated with an enriched expression of tissue-restricted antigens (TRAs). This unexpected finding, suggestive of wider functions of AIRE, may provide an explanation for the non-autoimmune symptoms of APECED patients who lack functional AIRE.
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- 2019
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33. Publisher Correction: Single-cell RNA-sequencing of differentiating iPS cells reveals dynamic genetic effects on gene expression
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Anna S. E. Cuomo, Daniel D. Seaton, Davis J. McCarthy, Iker Martinez, Marc Jan Bonder, Jose Garcia-Bernardo, Shradha Amatya, Pedro Madrigal, Abigail Isaacson, Florian Buettner, Andrew Knights, Kedar Nath Natarajan, HipSci Consortium, Ludovic Vallier, John C. Marioni, Mariya Chhatriwala, and Oliver Stegle
- Subjects
Science - Abstract
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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- 2020
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34. Genome-wide Bisulfite Sequencing in Zygotes Identifies Demethylation Targets and Maps the Contribution of TET3 Oxidation
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Julian R. Peat, Wendy Dean, Stephen J. Clark, Felix Krueger, Sébastien A. Smallwood, Gabriella Ficz, Jong Kyoung Kim, John C. Marioni, Timothy A. Hore, and Wolf Reik
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Biology (General) ,QH301-705.5 - Abstract
Fertilization triggers global erasure of paternal 5-methylcytosine as part of epigenetic reprogramming during the transition from gametic specialization to totipotency. This involves oxidation by TET3, but our understanding of its targets and the wider context of demethylation is limited to a small fraction of the genome. We employed an optimized bisulfite strategy to generate genome-wide methylation profiles of control and TET3-deficient zygotes, using SNPs to access paternal alleles. This revealed that in addition to pervasive removal from intergenic sequences and most retrotransposons, gene bodies constitute a major target of zygotic demethylation. Methylation loss is associated with zygotic genome activation and at gene bodies is also linked to increased transcriptional noise in early development. Our data map the primary contribution of oxidative demethylation to a subset of gene bodies and intergenic sequences and implicate redundant pathways at many loci. Unexpectedly, we demonstrate that TET3 activity also protects certain CpG islands against methylation buildup.
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- 2014
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35. A step-by-step workflow for low-level analysis of single-cell RNA-seq data with Bioconductor [version 2; referees: 1 approved, 4 approved with reservations]
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Aaron T.L. Lun, Davis J. McCarthy, and John C. Marioni
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Bioinformatics ,Genomics ,Medicine ,Science - Abstract
Single-cell RNA sequencing (scRNA-seq) is widely used to profile the transcriptome of individual cells. This provides biological resolution that cannot be matched by bulk RNA sequencing, at the cost of increased technical noise and data complexity. The differences between scRNA-seq and bulk RNA-seq data mean that the analysis of the former cannot be performed by recycling bioinformatics pipelines for the latter. Rather, dedicated single-cell methods are required at various steps to exploit the cellular resolution while accounting for technical noise. This article describes a computational workflow for low-level analyses of scRNA-seq data, based primarily on software packages from the open-source Bioconductor project. It covers basic steps including quality control, data exploration and normalization, as well as more complex procedures such as cell cycle phase assignment, identification of highly variable and correlated genes, clustering into subpopulations and marker gene detection. Analyses were demonstrated on gene-level count data from several publicly available datasets involving haematopoietic stem cells, brain-derived cells, T-helper cells and mouse embryonic stem cells. This will provide a range of usage scenarios from which readers can construct their own analysis pipelines.
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- 2016
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36. Structure and evolutionary history of a large family of NLR proteins in the zebrafish
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Kerstin Howe, Philipp H. Schiffer, Julia Zielinski, Thomas Wiehe, Gavin K. Laird, John C. Marioni, Onuralp Soylemez, Fyodor Kondrashov, and Maria Leptin
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nacht ,b30.2 ,spry ,gene conversion ,innate immune system ,genome evolution ,Biology (General) ,QH301-705.5 - Abstract
Multicellular eukaryotes have evolved a range of mechanisms for immune recognition. A widespread family involved in innate immunity are the NACHT-domain and leucine-rich-repeat-containing (NLR) proteins. Mammals have small numbers of NLR proteins, whereas in some species, mostly those without adaptive immune systems, NLRs have expanded into very large families. We describe a family of nearly 400 NLR proteins encoded in the zebrafish genome. The proteins share a defining overall structure, which arose in fishes after a fusion of the core NLR domains with a B30.2 domain, but can be subdivided into four groups based on their NACHT domains. Gene conversion acting differentially on the NACHT and B30.2 domains has shaped the family and created the groups. Evidence of positive selection in the B30.2 domain indicates that this domain rather than the leucine-rich repeats acts as the pathogen recognition module. In an unusual chromosomal organization, the majority of the genes are located on one chromosome arm, interspersed with other large multigene families, including a new family encoding zinc-finger proteins. The NLR-B30.2 proteins represent a new family with diversity in the specific recognition module that is present in fishes in spite of the parallel existence of an adaptive immune system.
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- 2016
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37. BASiCS workflow: a step-by-step analysis of expression variability using single cell RNA sequencing data [version 2; peer review: 3 approved with reservations]
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Alan O'Callaghan, Nils Eling, John C. Marioni, and Catalina A. Vallejos
- Subjects
Software Tool Article ,Articles ,single-cell RNA sequencing ,expression variability ,transcriptional noise ,differential expression testing ,scRNAseq ,Bayesian ,bioinformatics ,heterogeneity - Abstract
Cell-to-cell gene expression variability is an inherent feature of complex biological systems, such as immunity and development. Single-cell RNA sequencing is a powerful tool to quantify this heterogeneity, but it is prone to strong technical noise. In this article, we describe a step-by-step computational workflow that uses the BASiCS Bioconductor package to robustly quantify expression variability within and between known groups of cells (such as experimental conditions or cell types). BASiCS uses an integrated framework for data normalisation, technical noise quantification and downstream analyses, propagating statistical uncertainty across these steps. Within a single seemingly homogeneous cell population, BASiCS can identify highly variable genes that exhibit strong heterogeneity as well as lowly variable genes with stable expression. BASiCS also uses a probabilistic decision rule to identify changes in expression variability between cell populations, whilst avoiding confounding effects related to differences in technical noise or in overall abundance. Using a publicly available dataset, we guide users through a complete pipeline that includes preliminary steps for quality control, as well as data exploration using the scater and scran Bioconductor packages. The workflow is accompanied by a Docker image that ensures the reproducibility of our results.
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- 2024
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38. Correction: Corrigendum: Characterizing noise structure in single-cell RNA-seq distinguishes genuine from technical stochastic allelic expression
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Jong Kyoung Kim, Aleksandra A. Kolodziejczyk, Tomislav Ilicic, Sarah A. Teichmann, and John C. Marioni
- Subjects
Science - Abstract
Nature Communications 6: Article number: 8687 (2015); Published: 22 October 2015; Updated: 11 January 2016. The original version of this Article contained an error in the spelling of the author Tomislav Ilicic, which was incorrectly given as Tomislav Illicic. This has now been corrected in both the PDF and HTML versions of the Article.
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- 2016
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39. RNAget: an API to securely retrieve RNA quantifications.
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Sean Upchurch, Emilio Palumbo, Jeremy Adams, David Bujold, Guillaume Bourque, Jared Nedzel, Keenan Graham, Meenakshi S. Kagda, Pedro Assis, Benjamin C. Hitz, Emilio Righi, Roderic Guigó, Barbara J. Wold, Alvis Brazma, Julia Burchard, Joe Capka, Michael Cherry, Laura Clarke, Brian Craft, Manolis Dermitzakis, Mark Diekhans, John Dursi, Michael Sean Fitzsimons, Zac Flaming, Romina Garrido, Alfred Gil, Paul Godden, Matt Green, Mitch Guttman, Brian Haas, Max Haeussler, Bo Li, Sten Linnarsson, Adam Lipski, David Liu, Simonne Longerich, David Lougheed, Jonathan Manning, John C. Marioni, Christopher Meyer, Stephen B. Montgomery, Alyssa Morrow, Alfonso Muñoz-Pomer Fuentes, Jared L. Nedzel, David Nguyen, Kevin Osborn, Francis Ouellette, Irene Papatheodorou, Dmitri D. Pervouchine, Arun K. Ramani, Jordi Rambla, Bashir Sadjad, David Steinberg, Jeremiah Talkar, Timothy Tickle, Kathy Tzeng, Saman Vaisipour, Sean Watford, Barbara Wold, Zhenyu Zhang, and Jing Zhu
- Published
- 2023
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40. Expression Atlas update: gene and protein expression in multiple species.
- Author
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Pablo A. Moreno, Silvie Fexova, Nancy George, Jonathan R. Manning, Zhichao Miao, Suhaib Mohammed, Alfonso Muñoz-Pomer Fuentes, Anja Füllgrabe, Yalan Bi, Natassja Bush, Haider Iqbal, Upendra Kumbham, Andrey Solovyev, Lingyun Zhao, Ananth Prakash, David García-Seisdedos, Deepti Jaiswal Kundu, Shengbo Wang, Mathias Walzer, Laura Clarke, David Osumi-Sutherland, Marcela Karey Tello-Ruiz, Sunita Kumari, Doreen Ware, Jana Eliasova, Mark J. Arends, Martijn C. Nawijn, Kerstin B. Meyer, Tony Burdett, John C. Marioni, Sarah A. Teichmann, Juan Antonio Vizcaíno, Alvis Brazma, and Irene Papatheodorou
- Published
- 2022
- Full Text
- View/download PDF
41. BASiCS workflow: a step-by-step analysis of expression variability using single cell RNA sequencing data [version 1; peer review: 3 approved with reservations]
- Author
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Alan O'Callaghan, Nils Eling, John C. Marioni, and Catalina A. Vallejos
- Subjects
Software Tool Article ,Articles ,single-cell RNA sequencing ,expression variability ,transcriptional noise ,differential expression testing ,scRNAseq ,Bayesian ,bioinformatics ,heterogeneity - Abstract
Cell-to-cell gene expression variability is an inherent feature of complex biological systems, such as immunity and development. Single-cell RNA sequencing is a powerful tool to quantify this heterogeneity, but it is prone to strong technical noise. In this article, we describe a step-by-step computational workflow that uses the BASiCS Bioconductor package to robustly quantify expression variability within and between known groups of cells (such as experimental conditions or cell types). BASiCS uses an integrated framework for data normalisation, technical noise quantification and downstream analyses, propagating statistical uncertainty across these steps. Within a single seemingly homogeneous cell population, BASiCS can identify highly variable genes that exhibit strong heterogeneity as well as lowly variable genes with stable expression. BASiCS also uses a probabilistic decision rule to identify changes in expression variability between cell populations, whilst avoiding confounding effects related to differences in technical noise or in overall abundance. Using a publicly available dataset, we guide users through a complete pipeline that includes preliminary steps for quality control, as well as data exploration using the scater and scran Bioconductor packages. The workflow is accompanied by a Docker image that ensures the reproducibility of our results.
- Published
- 2022
- Full Text
- View/download PDF
42. Expression Atlas update: from tissues to single cells.
- Author
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Irene Papatheodorou, Pablo A. Moreno, Jonathan R. Manning, Alfonso Muñoz-Pomer Fuentes, Nancy George, Silvie Fexova, Nuno A. Fonseca, Anja Füllgrabe, Matthew Green 0005, Ni Huang, Laura Huerta, Haider Iqbal, Monica Jianu, Suhaib Mohammed, Lingyun Zhao, Andrew F. Jarnuczak, Simon Jupp, John C. Marioni, Kerstin B. Meyer, Robert Petryszak, Cesar Augusto Prada Medina, Carlos Talavera-López, Sarah A. Teichmann, Juan Antonio Vizcaíno, and Alvis Brazma
- Published
- 2020
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43. A single-cell molecular map of mouse gastrulation and early organogenesis.
- Author
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Blanca Pijuan-Sala, Jonathan A. Griffiths, Carolina Guibentif, Tom W. Hiscock, Wajid Jawaid, Fernando J. Calero-Nieto, Carla Mulas, Ximena Ibarra-Soria, Richard C. V. Tyser, Debbie Lee Lian Ho, Wolf Reik, Shankar Srinivas, Benjamin D. Simons, Jennifer Nichols, John C. Marioni, and Berthold Göttgens
- Published
- 2019
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44. The human body at cellular resolution: the NIH Human Biomolecular Atlas Program.
- Author
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Michael P. Snyder, Shin Lin, Amanda Posgai, Mark Atkinson, Aviv Regev, Jennifer Rood, Orit Rozenblatt-Rosen, Leslie Gaffney, Anna Hupalowska, Rahul Satija, Nils Gehlenborg, Jay Shendure, Julia Laskin, Pehr Harbury, Nicholas A. Nystrom, Jonathan C. Silverstein, Ziv Bar-Joseph, Kun Zhang 0020, Katy Börner, Yiing Lin, Richard Conroy, Dena Procaccini, Ananda L. Roy, Ajay Pillai, Marishka Brown, Zorina S. Galis, Long Cai, Cole Trapnell, Dana Jackson, Garry P. Nolan, William James Greenleaf, Sylvia K. Plevritis, Sara Ahadi, Stephanie A. Nevins, Hayan Lee, Christian Martijn Schuerch, Sarah Black, Vishal Gautham Venkataraaman, Ed Esplin, Aaron Horning, Amir Bahmani, Xin Sun, Sanjay Jain 0006, James S. Hagood, Gloria Pryhuber, Peter V. Kharchenko, Bernd Bodenmiller, Todd Brusko, Michael Clare-Salzler, Harry Nick, Kevin Otto 0001, Clive Wasserfall, Marda Jorgensen, Maigan Brusko, Sergio Maffioletti, Richard M. Caprioli, Jeffrey M. Spraggins, Danielle Gutierrez, Nathan Heath Patterson, Elizabeth K. Neumann, Raymond Harris, Mark P. de Caestecker, Agnes B. Fogo, Raf Van de Plas, Ken Lau, Guo-Cheng Yuan, Qian Zhu, Ruben Dries, Peng Yin, Sinem K. Saka, Jocelyn Y. Kishi, Yu Wang, Isabel Goldaracena, Dong Hye Ye, Kristin E. Burnum-Johnson, Paul D. Piehowski, Charles Ansong, Ying Zhu, Tushar Desai, Jay Mulye, Peter Chou, Monica Nagendran, Sarah A. Teichmann, Benedict Paten, Robert F. Murphy, Jian Ma 0004, Vladimir Yu. Kiselev, Carl Kingsford, Allyson Ricarte, Maria Keays, Sushma Anand Akoju, Matthew Ruffalo, Margaret Vella, Chuck McCallum, Leonard E. Cross, Samuel H. Friedman, Randy W. Heiland, Bruce William Herr II, Paul Macklin, Ellen M. Quardokus, Lisel Record, James P. Sluka, Griffin M. Weber, Philip D. Blood, Alexander Ropelewski, William Shirey, Robin M. Scibek, Paula M. Mabee, W. Christopher Lenhardt, Kimberly Robasky, Stavros Michailidis, John C. Marioni, Andrew Butler, Tim Stuart, Eyal Fisher, Shila Ghazanfar, Gökcen Eraslan, Tommaso Biancalani, Eeshit D. Vaishnav, Pothur Srinivas, Aaron Pawlyk, Salvatore Sechi, Elizabeth L. Wilder, and James Anderson
- Published
- 2019
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45. Stabilized mosaic single-cell data integration using unshared features
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Shila Ghazanfar, Carolina Guibentif, and John C. Marioni
- Subjects
Biomedical Engineering ,Molecular Medicine ,Bioengineering ,Applied Microbiology and Biotechnology ,Biotechnology - Abstract
Currently available single-cell omics technologies capture many unique features with different biological information content. Data integration aims to place cells, captured with different technologies, onto a common embedding to facilitate downstream analytical tasks. Current horizontal data integration techniques use a set of common features, thereby ignoring non-overlapping features and losing information. Here we introduce StabMap, a mosaic data integration technique that stabilizes mapping of single-cell data by exploiting the non-overlapping features. StabMap first infers a mosaic data topology based on shared features, then projects all cells onto supervised or unsupervised reference coordinates by traversing shortest paths along the topology. We show that StabMap performs well in various simulation contexts, facilitates ‘multi-hop’ mosaic data integration where some datasets do not share any features and enables the use of spatial gene expression features for mapping dissociated single-cell data onto a spatial transcriptomic reference.
- Published
- 2023
46. High-throughput phenotyping of single nucleotide variants by linking transcriptomes to genotypes in single cells
- Author
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Sarah E. Cooper, Matthew A. Coelho, Magdalena E. Strauss, Aleksander M. Gontarczyk, Qianxin Wu, Mathew J. Garnett, John C. Marioni, and Andrew R. Bassett
- Abstract
CRISPR screens with single-cell transcriptomic readouts are a valuable tool to understand the effect of genetic perturbations, but are currently limited because genotypes are inferred from the guide RNA identity. We have developed a technique that couples single-cell genotyping to transcriptomics of the same cells to enable screening for the effects of single nucleotide variants. Analysis of variants tiling across theJAK1gene demonstrates the importance of determining the precise genetic perturbation and classifies missense variants into three functional categories.
- Published
- 2023
47. A Human Breast Cell Atlas Mapping the Homeostatic Cellular Shifts in the Adult Breast
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Austin D. Reed, Sara Pensa, Adi Steif, Jack Stenning, Daniel J. Kunz, Peng He, Alecia-Jane Twigger, Katarzyna Kania, Rachel Barrow-McGee, Iain Goulding, Jennifer J. Gomm, Louise Jones, John C. Marioni, and Walid T. Khaled
- Abstract
One of the barriers for breast cancer prevention and treatment is our poor understanding of the dynamic cellular shifts that naturally occur within the breast and how these changes contribute to tumour initiation. In this study we report the use of single cell RNA sequencing (scRNAseq) to compile a Human Breast Cell Atlas (HBCA) assembled from 55 donors that had undergone reduction mammoplasties or risk reduction mammoplasties. The data from more than 800,000 cells identified 41 cell subclusters distributed across the epithelial, immune, and stromal compartments. We found that the contribution of these different clusters varied according to the natural history of the tissue. Breast cancer risk modulating factors such as age, parity, and germline mutation affected the homeostatic cellular state of the breast in different ways however, none of the changes observed were restricted to any one cell type. Remarkably, we also found that immune cells fromBRCA1/2carriers had a distinct gene expression signature indicative of potential immune exhaustion. This suggests that immune escape mechanisms could manifest in non-cancerous tissues during very early stages of tumour initiation. Therefore, the Atlas presented here provides the research community with a rich resource that can be used as a reference for studies on the origins of breast cancer which could inform novel approaches for early detection and prevention.
- Published
- 2023
48. Time, Space and Single-Cell Resolved Molecular Trajectory of Cell Populations and the Laterality of the Body Plan at Gastrulation
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Ran Wang, Xianfa Yang, Jiehui Chen, Lin Zhang, Jonathan A. Griffiths, Guizhong Cui, Yingying Chen, Yun Qian, Guangdun Peng, Jinsong Li, Liantang Wang, John C. Marioni, Patrick P.L. Tam, and Naihe Jing
- Abstract
Understanding of the molecular drivers of lineage diversification and tissue patterning during primary germ layer development requires in-depth knowledge of the dynamic molecular trajectories of cell lineages across a series of developmental stages of gastrulation1–7. Through computational modeling, we constructed at single-cell resolution a spatio-temporal compendium of the molecular trajectories of germ-layer derivatives in gastrula-stage mouse embryos. This molecular atlas infers the developmental trajectories of single-cell populations and the molecular network activity underpinning the specification and differentiation of the germ-layer lineages. Analysis of the heterogeneity of cellular composition of cell populations at defined positions in the epiblast revealed progressive diversification of cell types, mirroring the process of lineage allocation during gastrulation. A novel observation is the difference in the contribution of cells on contralateral sides of the epiblast to mesoderm derivatives of the early organogenesis embryo, and the enhanced BMP signaling activity in right-side mesoderm of E7.5 embryo. Perturbation of BMP signaling activity at late gastrulation led to randomization of left-right (L-R) molecular asymmetry in the lateral mesoderm of early-somite-stage embryo. Our findings indicate the asymmetric BMP activity during gastrulation may be critical for the symmetry breaking process associated with specification of L-R body asymmetry ahead of the acquisition of functionality of the L-R organizer.
- Published
- 2023
49. Data from Contributions to Drug Resistance in Glioblastoma Derived from Malignant Cells in the Sub-Ependymal Zone
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Colin Watts, Simon Tavaré, John C. Marioni, Christina Curtis, Ashok R. Venkitaraman, Vincent P. Collins, Karen D. Howarth, Nicola-Jane Francis, Richard Heywood, Stephen J. Price, Suzan Ber, Anestis Touloumis, Andrea Sottoriva, Inmaculada Spiteri, and Sara G.M. Piccirillo
- Abstract
Glioblastoma, the most common and aggressive adult brain tumor, is characterized by extreme phenotypic diversity and treatment failure. Through fluorescence-guided resection, we identified fluorescent tissue in the sub-ependymal zone (SEZ) of patients with glioblastoma. Histologic analysis and genomic characterization revealed that the SEZ harbors malignant cells with tumor-initiating capacity, analogous to cells isolated from the fluorescent tumor mass (T). We observed resistance to supramaximal chemotherapy doses along with differential patterns of drug response between T and SEZ in the same tumor. Our results reveal novel insights into glioblastoma growth dynamics, with implications for understanding and limiting treatment resistance. Cancer Res; 75(1); 194–202. ©2014 AACR.
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- 2023
50. Supplementary Methods, Figures 1 - 19, Tables 1 - 6 from Contributions to Drug Resistance in Glioblastoma Derived from Malignant Cells in the Sub-Ependymal Zone
- Author
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Colin Watts, Simon Tavaré, John C. Marioni, Christina Curtis, Ashok R. Venkitaraman, Vincent P. Collins, Karen D. Howarth, Nicola-Jane Francis, Richard Heywood, Stephen J. Price, Suzan Ber, Anestis Touloumis, Andrea Sottoriva, Inmaculada Spiteri, and Sara G.M. Piccirillo
- Abstract
Supplementary experimental procedures: this file contains an expanded materials and methods section on 5-ALA administration and sample collection, cell lines propagation, immunofluorescence and in vivo assays, genomic and molecular clock analysis, fluorescence in situ hybridization, drug concentration used in the proliferation assay and MGMT promoter methylation analysis. Supplementary Figure S1: this figure illustrates the identification of the sub-ependymal zone (SEZ) in two GB patients included in this study. Supplementary Figure S2: this figure shows the sampling and morphology of the SEZ tissue. Supplementary Figure S3: this figure summarizes the results of real time analysis for markers of glial, precursor, stem cells and proliferation in paired T and SEZ from three GB patients. Supplementary Figure S4: this figure shows an example of common copy number aberration breakpoints between the SEZ and the corresponding T in each of the GB patients included in this study. Supplementary Figure S5: this figure summarizes the fluorescence in situ hybridization (FISH) results for EGFR, MET and PTEN in three GB patients. Supplementary Figure S6: this figure shows the copy number aberration breakpoints between T and SEZ and the corresponding cell lines in patient sp14. Supplementary Figure S7: this figure shows the results of growth curve analysis of tumor-initiating cells (TICs) derived from matched T and SEZ of five GBs. Supplementary Figure S8: this figure summarizes the results of clonogenic index between three paired T and SEZ cells and multipotency of SEZ cells. Supplementary Figure S9: this figure shows the immunofluorescence results for stem cell markers on T and SEZ cells in two GBs. Supplementary Figure S10: this figure summarizes the results of cumulative Kaplan-Meier survival analysis from five additional GBs. Supplementary Figure S11: this figure shows an analysis of the in vivo properties of TICs isolated from the SEZ of three GBs. Supplementary Figure S12: this figure shows the results of the cell proliferation assay of 7 paired TICs using 50μM of Temozolomide (TMZ). Supplementary Figure S13: this figure summarizes the cell proliferation data of additional 20 TICs treated with TMZ for dose escalation analysis. Supplementary Figure S14: this figure shows the results of cell proliferation assay of additional 4 paired TICs treated with TMZ, Cisplatin and Cediranib. Supplementary Figure S15: this figure shows a gel image of MGMT promoter status analysis for T and SEZ of the analyzed 7 paired TICs. Supplementary Figure S16: this figure summarizes the immunofluorescence results for vascular endothelial growth factor receptor 2 (Vegfr2) on T and SEZ cells from three GBs. Supplementary Figure S17: this figure summarizes the cell proliferation data of three control lines. Supplementary Figure S18: this figure illustrates a model of residual disease in human GB and summarizes the cardinal features of T and SEZ. Supplementary Figure S19: this figure shows the results of growth curve analysis of two SEZ in absence of fluorescence. Supplementary Table S1: this table summarizes the results of the histological features of T and SEZ tissues. Supplementary Table S2: this table provides a summary of the samples used for the genomic analysis and of the clinical information available for the eleven GB patients included in this study. Supplementary Table S3: this table shows the gene ontology (GO) terms whose genes are differentially expressed between SEZ and T. Supplementary Table S4: this table provides a summary of the results of MGMT methylation analysis by pyrosequencing in sixteen GBs. Supplementary Table S5: this table summarizes the overall survival of the eight GB patients whose TICs were used for the cell proliferation assay and includes the MGMT methylation by pyrosequencing. Supplementary Table S6: this table shows the gene expression data of PDGF and VEGF receptors in T and SEZ.
- Published
- 2023
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