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Stereo-sequencing: 3D Transcriptome of the mouse embryo at gastrulation

ABSTRACT: To glean an appreciation of the holistic genetic activity in the gastrulating mouse embryo, we performed a genome-wide spatial transcriptome analysis (Stereo-seq), using a low-cell number sequencing protocol on laser microdissected samples of epiblast cells with retained positional address. The 3D transcriptome reveals that (i) the epiblast is partitioned into transcription domains corresponding to regions of epiblast where cells are endowed specifically with ectoderm and mesendoderm potency, (ii) novel lineage markers are identified as genes expressed in epiblast domains populated by cells displaying different lineage fates, (iii) functionally related gene regulatory circuitry and signaling pathways are acting in concert in the transcriptional domains, and (iv) the spatial information provides reference zipcodes for mapping the prospective address of cell samples from different embryos and stem cell lines. The quantified expression data can also be visualized as “3D digitized whole mount in situ hybridization” of all the expressed transcripts in the epiblast.

ORGANISM(S): Mus musculus

PROVIDER: GSE65924 | GEO | 2016/03/24

SECONDARY ACCESSION(S): PRJNA275415

REPOSITORIES: GEO

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Project description:To glean an appreciation of the holistic genetic activity in the gastrulating mouse embryo, we performed a genome-wide spatial transcriptome analysis (Stereo-seq), using a low-cell number sequencing protocol on laser microdissected samples of epiblast cells with retained positional address. The 3D transcriptome reveals that (i) the epiblast is partitioned into transcription domains corresponding to regions of epiblast where cells are endowed specifically with ectoderm and mesendoderm potency, (ii) novel lineage markers are identified as genes expressed in epiblast domains populated by cells displaying different lineage fates, (iii) functionally related gene regulatory circuitry and signaling pathways are acting in concert in the transcriptional domains, and (iv) the spatial information provides reference zipcodes for mapping the prospective address of cell samples from different embryos and stem cell lines. The quantified expression data can also be visualized as â3D digitized whole mount in situ hybridizationâ of all the expressed transcripts in the epiblast. (i) By using laser-microdissection, we carried out transcriptome profiling on embryo sections at a high resolution of ~20 cells per sample with the spatial information preserved. We then constructed a comprehensive spatial transcriptome map in the mid-gastrulation embryo that is visualized in a 3D embryonic model based on the sequencing data. Embryo position (A/L/P/R) and section (1-11) descriptors: A stands for laser capture microdissected samples from the anterior epiblast of the embryo; P for posterior; L for the left lateral epiblast of the embryo; R for the right lateral. The section is collected from distal to proximal, and the section 1 to 11 is the cryosection order, covering the whole embryonic part of a late mid-streak embryo. Section 1 is the most distal section and 11 is the most proximal section. (ii) Additional samples are RNA-seq data of 70 single cells from E7.0 mouse embryo. These 70 samples were randomly picked from the anterior or posterior embryonic half.

Project description:Aging significantly elevates the risk for Alzheimer’s disease (AD), contributing to the accumulation of AD pathologies, such as amyloid-β (Aβ), inflammation, and oxidative stress. The human prefrontal cortex (PFC) is highly vulnerable to the impacts of both aging and AD. Unveiling and understanding the molecular alterations in PFC associated with normal aging (NA) and AD is essential for elucidating the mechanisms of AD progression and developing novel therapeutics for this devastating disease. In this study, for the first time, we employed a cutting-edge spatial transcriptome platform, STOmics® SpaTial Enhanced Resolution Omics-sequencing (Stereo-seq), to generate the first comprehensive, subcellular resolution spatial transcriptome atlas of the human PFC from six AD cases at various neuropathological stages and six age, sex, and ethnicity matched controls. Our analyses revealed distinct transcriptional alterations across six neocortex layers, highlighted the AD-associated disruptions in laminar architecture, and identified changes in layer-to-layer interactions as AD progresses. Further, throughout the progression from NA to various stages of AD, we discovered specific genes that were significantly upregulated in neurons experiencing high stress and in nearby non-neuronal cells, compared to cells distant from the source of stress. Notably, the cell-cell interactions between the neurons under the high stress and adjacent glial cells that promote Aβ clearance and neuroprotection were diminished in AD in response to stressors compared to NA. Through cell-type specific gene co-expression analysis, we identified three modules in excitatory and inhibitory neurons associated with neuronal protection, protein dephosphorylation, and negative regulation of Aβ plaque formation. These modules negatively correlated with AD progression, indicating a reduced capacity for toxic substance clearance in AD subject samples. Moreover, we have discovered a novel transcription factor, ZNF460, that regulates all three modules, establishing it as a potential new therapeutic target for AD. Overall, utilizing the latest spatial transcriptome platform, our study developed the first transcriptome-wide atlas with subcellular resolution for assessing the molecular alterations in the human PFC due to AD. This atlas sheds light on the potential mechanisms underlying the progression from NA to AD.

Project description:The generation of properly functioning gametes in vitro, a key goal in developmental/reproductive biology, requires multi-step reconstitutions of complex germ cell development. Based on the logic of primordial germ cell (PGC)-specification, we demonstrate here the generation of PGC-like cells (PGCLCs) in mice with robust capacity for spermatogenesis from embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) through epiblast-like cells (EpiLCs), a cellular state highly similar to pre-gastrulating epiblasts, but distinct from epiblast stem cells (EpiSCs). The global transcription profiles, epigenetic reprogramming, and cellular dynamics during PGCLC induction from EpiLCs are a meticulous capture of those associated with PGC specification from the epiblasts. Furthermore, we identify Integrin-beta 3 and SSEA1 as markers that purify PGCLCs with spermatogenic capacity free from tumorigenic undifferentiated cells. With the reconstitution of PGC specification pathway from the naive inner cell mass state, our study defines a paradigm for the essential step of in vitro gametogenesis. We performed this analysis to reveal the characters of the cells that we created in this study, epiblast-like cells (EpiLCs) and primordial germ cells-like cells (PGCLCs). Because EpiLCs were induced from embryonic stem cells (ESCs), and equivalent to pre-gastrulating epiblast (embryonic day [E] 5.5-6.0) in vivo (embryo), ESCs and epiblast were included in this analysis. Epiblast stem cells (EpiSCs) are a culture cell type derived from epiblast, and were also included. PGCLCs were supposed to be equivalent to E9.5 PGCs based on reporter fluorescent transgene expressions and epigenetic properties, and therefore E9.5 PGCs were also inckuded in this analysis. Because epiblast and E9.5 PGCs are of a small number of cells in embryos (a few hundred to thousand cells), cDNAs were amplified with a quantitative global PCR method (Kurimoto et al., 2006, Nucleic Acids Research) for microarray analyses. We took two biological replicate for each cell type.

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Stereo-sequencing: 3D Transcriptome of the mouse embryo at gastrulation

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