Integrative single-cell analysis of transcriptional and epigenetic states in the human adult brain

Detailed characterization of the cell types in the human brain requires scalable experimental approaches to examine multiple aspects of the molecular state of individual cells, as well as computational integration of the data to produce unified cell-state annotations. Here we report improved high-throughput methods for single-nucleus droplet-based sequencing (snDrop-seq) and single-cell transposome hypersensitive site sequencing (scTHS-seq). We used each method to acquire nuclear transcriptomic and DNA accessibility maps for >60,000 single cells from human adult visual cortex, frontal cortex, and cerebellum. Integration of these data revealed regulatory elements and transcription factors that underlie cell-type distinctions, providing a basis for the study of complex processes in the brain, such as genetic programs that coordinate adult remyelination. We also mapped disease-associated risk variants to specific cellular populations, which provided insights into normal and pathogenic cellular processes in the human brain. This integrative multi-omics approach permits more detailed single-cell interrogation of complex organs and tissues.

[1]  M. Mattson.  Pathways towards and away from Alzheimer's disease , 2004, Nature.

[2]  M. Wegner,et al.  Sox9 and Sox10 influence survival and migration of oligodendrocyte precursors in the spinal cord by regulating PDGF receptor α expression , 2008, Development.

[3]  F. Collins,et al.  Potential etiologic and functional implications of genome-wide association loci for human diseases and traits , 2009, Proceedings of the National Academy of Sciences.

[4]  Z. Werb,et al.  Sox10 directs neural stem cells toward the oligodendrocyte lineage by decreasing Suppressor of Fused expression , 2010, Proceedings of the National Academy of Sciences.

[5]  D. Herr,et al.  FTY720 (fingolimod) efficacy in an animal model of multiple sclerosis requires astrocyte sphingosine 1-phosphate receptor 1 (S1P1) modulation , 2010, Proceedings of the National Academy of Sciences.

[6]  Timothy J. Durham,et al.  "Systematic" , 1966, Comput. J..

[7]  L. Feuk,et al.  Total RNA sequencing reveals nascent transcription and widespread co-transcriptional splicing in the human brain , 2011, Nature Structural &Molecular Biology.

[8]  R. Douglas Fields,et al.  Control of Local Protein Synthesis and Initial Events in Myelination by Action Potentials , 2011, Science.

[9]  Allan R. Jones,et al.  Large-Scale Cellular-Resolution Gene Profiling in Human Neocortex Reveals Species-Specific Molecular Signatures , 2012, Cell.

[10]  Allan R. Jones,et al.  An anatomically comprehensive atlas of the adult human brain transcriptome , 2012, Nature.

[11]  Data production leads,et al.  An integrated encyclopedia of DNA elements in the human genome , 2012 .

[12]  Shane J. Neph,et al.  Systematic Localization of Common Disease-Associated Variation in Regulatory DNA , 2012, Science.

[13]  ENCODEConsortium,et al.  An Integrated Encyclopedia of DNA Elements in the Human Genome , 2012, Nature.

[14]  H. Monyer,et al.  Bergmann Glial AMPA Receptors Are Required for Fine Motor Coordination , 2012, Science.

[15]  Yasuyuki Kihara,et al.  Fingolimod: Direct CNS effects of sphingosine 1-phosphate (S1P) receptor modulation and implications in multiple sclerosis therapy , 2013, Journal of the Neurological Sciences.

[16]  Kun Zhang,et al.  Massively parallel polymerase cloning and genome sequencing of single cells using nanoliter microwells , 2013, Nature Biotechnology.

[17]  F. Gage,et al.  RNA-sequencing from single nuclei , 2013, Proceedings of the National Academy of Sciences.

[18]  Buhm Han,et al.  Chromatin marks identify critical cell types for fine mapping complex trait variants , 2012 .

[19]  J. Rubenstein,et al.  Subcortical origins of human and monkey neocortical interneurons , 2013, Nature Neuroscience.

[20]  E. Klann,et al.  Suppression of eIF2α kinases alleviates AD-related synaptic plasticity and spatial memory deficits , 2013, Nature Neuroscience.

[21]  Jan H Lui,et al.  Non-epithelial stem cells and cortical interneuron production in the human ganglionic eminences , 2013, Nature Neuroscience.

[22]  L. Tran,et al.  Integrated Systems Approach Identifies Genetic Nodes and Networks in Late-Onset Alzheimer’s Disease , 2013, Cell.

[23]  F. Rossi,et al.  Origin, lineage and function of cerebellar glia , 2013, Progress in Neurobiology.

[24]  D. Attwell,et al.  Neuregulin and BDNF Induce a Switch to NMDA Receptor-Dependent Myelination by Oligodendrocytes , 2013, PLoS biology.

[25]  Howard Y. Chang,et al.  Transposition of native chromatin for fast and sensitive epigenomic profiling of open chromatin, DNA-binding proteins and nucleosome position , 2013, Nature Methods.

[26]  T. Butts,et al.  Development of the cerebellum: simple steps to make a ‘little brain’ , 2014, Development.

[27]  Cole Trapnell,et al.  The dynamics and regulators of cell fate decisions are revealed by pseudotemporal ordering of single cells , 2014, Nature Biotechnology.

[28]  T. Maniatis,et al.  An RNA-Sequencing Transcriptome and Splicing Database of Glia, Neurons, and Vascular Cells of the Cerebral Cortex , 2014, The Journal of Neuroscience.

[29]  S. Linnarsson,et al.  Cell types in the mouse cortex and hippocampus revealed by single-cell RNA-seq , 2015, Science.

[30]  Ethan K. Scott,et al.  Neuronal activity biases axon selection for myelination in vivo , 2015, Nature Neuroscience.

[31]  Andrew C. Adey,et al.  Multiplex single-cell profiling of chromatin accessibility by combinatorial cellular indexing , 2015, Science.

[32]  J. Sklar,et al.  Genome-wide Detection of DNase I Hypersensitive Sites in Single Cells and FFPE Samples , 2015, Nature.

[33]  S. Quake,et al.  A survey of human brain transcriptome diversity at the single cell level , 2015, Proceedings of the National Academy of Sciences.

[34]  Allon M. Klein,et al.  Droplet Barcoding for Single-Cell Transcriptomics Applied to Embryonic Stem Cells , 2015, Cell.

[35]  Howard Y. Chang,et al.  Single-cell chromatin accessibility reveals principles of regulatory variation , 2015, Nature.

[36]  Evan Z. Macosko,et al.  Highly Parallel Genome-wide Expression Profiling of Individual Cells Using Nanoliter Droplets , 2015, Cell.

[37]  Qing-Yu He,et al.  ChIPseeker: an R/Bioconductor package for ChIP peak annotation, comparison and visualization , 2015, Bioinform..

[38]  G. von Heijne,et al.  Tissue-based map of the human proteome , 2015, Science.

[39]  D. Weitz,et al.  Single-cell ChIP-seq reveals cell subpopulations defined by chromatin state , 2015, Nature Biotechnology.

[40]  S. P. Fodor,et al.  Combinatorial labeling of single cells for gene expression cytometry , 2015, Science.

[41]  Gwendolyn E. Kaeser,et al.  Genomic mosaicism with increased amyloid precursor protein (APP) gene copy number in single neurons from sporadic Alzheimer's disease brains , 2015, eLife.

[42]  R. Reynolds,et al.  Neuronal activity regulates remyelination via glutamate signalling to oligodendrocyte progenitors , 2015, Nature Communications.

[43]  Michael Q. Zhang,et al.  Integrative analysis of 111 reference human epigenomes , 2015, Nature.

[44]  Staci A. Sorensen,et al.  Adult Mouse Cortical Cell Taxonomy Revealed by Single Cell Transcriptomics , 2016 .

[45]  R. Rocha,et al.  MEF2C haploinsufficiency syndrome: Report of a new MEF2C mutation and review. , 2016, European journal of medical genetics.

[46]  Grace X. Y. Zheng,et al.  Massively parallel digital transcriptional profiling of single cells , 2016, bioRxiv.

[47]  Howard Y. Chang,et al.  Lineage-specific and single cell chromatin accessibility charts human hematopoiesis and leukemia evolution , 2016, Nature Genetics.

[48]  S. Goldman,et al.  Dual regulatory switch through interactions of Tcf7l2/Tcf4 with stage-specific partners propels oligodendroglial maturation , 2016, Nature Communications.

[49]  Lars E. Borm,et al.  Molecular Diversity of Midbrain Development in Mouse, Human, and Stem Cells , 2016, Cell.

[50]  E. Chang,et al.  Purification and Characterization of Progenitor and Mature Human Astrocytes Reveals Transcriptional and Functional Differences with Mouse , 2016, Neuron.

[51]  Cynthia C. Hession,et al.  Div-Seq: Single-nucleus RNA-Seq reveals dynamics of rare adult newborn neurons , 2016, Science.

[52]  Kun Zhang,et al.  Characterization of chromatin accessibility with a transposome hypersensitive sites sequencing (THS-seq) assay , 2016, Genome Biology.

[53]  Fabian J. Theis,et al.  destiny: diffusion maps for large-scale single-cell data in R , 2015, Bioinform..

[54]  M. Ronaghi,et al.  Neuronal subtypes and diversity revealed by single-nucleus RNA sequencing of the human brain , 2016, Science.

[55]  David J. Arenillas,et al.  JASPAR 2016: a major expansion and update of the open-access database of transcription factor binding profiles , 2015, Nucleic Acids Res..

[56]  Jens Hjerling-Leffler,et al.  Oligodendrocyte heterogeneity in the mouse juvenile and adult central nervous system , 2016, Science.

[57]  Sara B. Linker,et al.  Using single nuclei for RNA-seq to capture the transcriptome of postmortem neurons , 2016, Nature Protocols.

[58]  Conor Fitzpatrick,et al.  Nuclear RNA-seq of single neurons reveals molecular signatures of activation , 2016, Nature communications.

[59]  Matt Thomson,et al.  Low Dimensionality in Gene Expression Data Enables the Accurate Extraction of Transcriptional Programs from Shallow Sequencing. , 2016, Cell systems.

[60]  Baptiste N. Jaeger,et al.  An environment-dependent transcriptional network specifies human microglia identity , 2017, Science.

[61]  William Stafford Noble,et al.  Massively multiplex single-cell Hi-C , 2016, Nature Methods.

[62]  Kun Zhang,et al.  A comparative strategy for single-nucleus and single-cell transcriptomes confirms accuracy in predicted cell-type expression from nuclear RNA , 2017, Scientific Reports.

[63]  Amirali Kia,et al.  Improved genome sequencing using an engineered transposase , 2017, BMC Biotechnology.