Div-Seq: Single-nucleus RNA-Seq reveals dynamics of rare adult newborn neurons

Habib, Naomi; Li, Yinqing; Heidenreich, Matthias; Swiech, Lukasz; Avraham‐Davidi, Inbal; Trombetta, John J.; Hession, Cynthia C.; Zhang, Feng; Regev, Aviv

doi:10.1126/science.aad7038

Cited by 503 publications

(570 citation statements)

References 58 publications

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“…With constantly improving technology for the analysis of single cells (65,66), it will soon be possible to evaluate the brain's full antiviral innate immune response. Additionally, in combination with genetically modified animals, and perhaps in vivo imaging (40), it will be possible to dissect the cellular mechanisms by which viruses are detected in the brain as well as the immune responses that are elicited.…”

Section: Future Directionsmentioning

confidence: 99%

The brain parenchyma has a type I interferon response that can limit virus spread

Drokhlyansky

Ayturk

Soh

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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The brain has a tightly regulated environment that protects neurons and limits inflammation, designated "immune privilege." However, there is not an absolute lack of an immune response. We tested the ability of the brain to initiate an innate immune response to a virus, which was directly injected into the brain parenchyma, and to determine whether this response could limit viral spread. We injected vesicular stomatitis virus (VSV), a transsynaptic tracer, or naturally occurring VSV-derived defective interfering particles (DIPs), into the caudate-putamen (CP) and scored for an innate immune response and inhibition of virus spread. We found that the brain parenchyma has a functional type I interferon (IFN) response that can limit VSV spread at both the inoculation site and among synaptically connected neurons. Furthermore, we characterized the response of microglia to VSV infection and found that infected microglia produced type I IFN and uninfected microglia induced an innate immune response following virus injection.

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Section: Future Directionsmentioning

confidence: 99%

The brain parenchyma has a type I interferon response that can limit virus spread

Drokhlyansky

Ayturk

Soh

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…High-throughput singlecell transcriptome [2][3][4] and chromatin accessibility profiling assays [5][6][7] have dramatically improved our ability to uncover latent cell types within complex tissues and dynamic cell states during differentiation. These same possibilities extend to DNA methylation; a covalent addition of a methyl group to cytosine bases in the genome that largely serves in a repressive role 8 .…”

Section: Mainmentioning

confidence: 99%

Scalable and efficient single-cell DNA methylation sequencing by combinatorial indexing

Mulqueen

Pokholok

Norberg

et al. 2017

Preprint

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Here we present a novel method: single-cell combinatorial indexing for methylation analysis (sci-MET), which is the first highly scalable assay for whole genome methylation profiling of single cells. We use sci-MET to produce 2,697 total single-cell bisulfite sequencing libraries and achieve read alignment rates of 69 ± 7%, comparable to those of bulk cell methods. As a proof of concept, we applied sci-MET to successfully deconvolve the cellular identity of a mixture of three human cell lines.. CC-BY-NC-ND 4.0 International license It is made available under a (which was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.The copyright holder for this preprint . http://dx.doi.org/10.1101/157230 doi: bioRxiv preprint first posted online Jun. 28, 2017; MainThe fundamental challenge of identifying and characterizing the molecular properties of every cell type in the body has recently entered the realm of possibility 1 . High-throughput singlecell transcriptome [2][3][4] and chromatin accessibility profiling assays [5][6][7] have dramatically improved our ability to uncover latent cell types within complex tissues and dynamic cell states during differentiation. These same possibilities extend to DNA methylation; a covalent addition of a methyl group to cytosine bases in the genome that largely serves in a repressive role 8 . DNA methylation occurs at a high rate in cytosine-guanine dinucleotides (CG), with cytosine methylation in non-CG sites (CH) occurring rarely and only in select tissues 9 . Both CG and CH methylation have cell type-specificity and are the subject of active modification in developing tissues 8 . DNA methylation can be probed at base-pair resolution using the deaminating chemistry of sodium bisulfite treatment before or after the generation of sequencing libraries (e.g. as in whole genome bisulfite sequencing, WGBS) 10,11 . Despite the comprehensive nature of these assays, key aspects of methylation architecture and dynamics remain elusive, with cell type heterogeneity at the forefront of this challenge.Recent work has optimized bisulfite sequencing to decrease input requirements to the single cell level (scWGBS) [12][13][14][15] . These assays have provided unique insights into environmental effects on methylome dynamics 13 , have uncovered new cell-types that form during hematopoesis 14 , and been coupled with scRNA-seq to directly study the relationship of DNA methylation to transcription 15 . However, these methods use a parallelized library generation strategy in which the WGBS protocol must be carried out on each cell in its own reaction vessel and thus remain low-throughput. Existing platforms to assess the transcriptome of single cells in high-throughput largely rely on droplet-based microfluidics strategies by sequestering single cells into individual droplets along with a substrate harboring barcoded oligonucleotides for cell-level indexing 2,16 . This approach is not readily adaptable to genome-wide DNA methylation pro...

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“…Recent progress have used scRNA-seq to examine detailed biology processes in newborn neuron generation and synaptic regulation [33,34]. However, there are still challenges for the application of scRNA-seq in the brain, given the highly myelinated and complicated cellular-cellular connection of the tissue.…”

Section: Conclusion Remarksmentioning

confidence: 99%

“…For instance, the making of fully expression profile of neural mRNA including dendritic and axonal mRNA is still difficult [35][36][37], although a previous TIVA approach based on a light-activated mRNA-capture reagent was designed to harvest mRNA from cells in their natural environment [38]. Another replacement strategy is to represent whole neuron RNA by their neuronal nuclei RNA as several studies have demonstrated that the nucleis contain enough amount of mRNA for analysing [34,39,40]. Additionally, as most cells in the body are under control of the circadian transcriptional system, it would introduce unwanted biological variations if this factor is ignored in the experimental design.…”

mentioning

confidence: 99%

Decoding nervous system by single‐cell RNA sequencing

Wang

2017

Quant. Biol.

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BackgroundMammalian brain are composed of a large number of specialized cell types with diverse molecular composition, functions and differentiation potentials. The application of recently developed single‐cell RNA sequencing (scRNA‐seq) technology in this filed has provided us new insights about this sophisticated system, deepened our understanding of the cell type diversity and led to the discovery of novel cell types.ResultsHere we review recent progresses of applying this technology on studying brain cell heterogeneity, adult neurogenesis as well as brain tumors, then we discuss some current limitations and future directions of using scRNA‐seq on the investagation of nervous system.ConclusionsWe believe the application of single‐cell RNA sequencing in neuroscience will accelerate the progress of big brain projects.

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Div-Seq: Single-nucleus RNA-Seq reveals dynamics of rare adult newborn neurons

Cited by 503 publications

References 58 publications

The brain parenchyma has a type I interferon response that can limit virus spread

The brain parenchyma has a type I interferon response that can limit virus spread

Scalable and efficient single-cell DNA methylation sequencing by combinatorial indexing

Decoding nervous system by single‐cell RNA sequencing

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