The genomics of human development Understanding the trajectory of a developing human requires an understanding of how genes are regulated and expressed. Two papers now present a pooled approach using three levels of combinatorial indexing to examine the single-cell gene expression and chromatin landscapes from 15 organs in fetal samples. Cao et al. focus on measurements of RNA in broadly distributed cell types and provide insights into organ specificity. Domcke et al. examined the chromatin accessibility of cells from these organs and identify the regulatory elements that regulate gene expression. Together, these analyses generate comprehensive atlases of early human development. Science, this issue p. eaba7721, p. eaba7612 Gene expression is examined at the single-cell level during early human development. INTRODUCTION A reference atlas of human cell types is a major goal for the field. Here, we set out to generate single-cell atlases of both gene expression (this study) and chromatin accessibility (Domcke et al., this issue) using diverse human tissues obtained during midgestation. RATIONALE Contemporary knowledge of the molecular basis of in vivo human development mostly derives from a combination of human genetics, in vivo investigations of model organisms, and in vitro studies of differentiating human cell lines, rather than through direct investigations of developing human tissues. Several challenges have historically limited the study of developing human tissues at the molecular level, including limited access, tissue degradation, and cell type heterogeneity. For this and the companion study (Domcke et al., this issue), we were able to overcome these challenges. RESULTS We applied three-level single-cell combinatorial indexing for gene expression (sci-RNA-seq3) to 121 human fetal samples ranging from 72 to 129 days in estimated postconceptual age and representing 15 organs, altogether profiling 4 million single cells. We developed and applied a framework for quantifying cell type specificity, identifying 657 cell subtypes, which we preliminarily annotated based on cross-matching to mouse cell atlases. We identified and validated potentially circulating trophoblast-like and hepatoblast-like cells in unexpected tissues. Profiling gene expression in diverse tissues facilitated the cross-tissue analyses of broadly distributed cell types, including blood, endothelial, and epithelial cells. For blood cells, this yielded a multiorgan map of cell state trajectories from hematopoietic stem cells to all major sublineages. Multiple lines of evidence support the adrenal gland as a normal, albeit minor, site of erythropoiesis during fetal development. It was notably straightforward to integrate these human fetal data with a mouse embryonic cell atlas, despite differences in species and developmental stage. For some systems, this essentially permitted us to bridge gene expression dynamics from the embryonic to the fetal stages of mammalian development. CONCLUSION The single-cell data resource presented here is notable for its scale, its focus on human fetal development, the breadth of tissues analyzed, and the parallel generation of gene expression (this study) and chromatin accessibility data (Domcke et al., this issue). We furthermore consolidate the technical framework for individual laboratories to generate and analyze gene expression and chromatin accessibility data from millions of single cells. Looking forward, we envision that the somewhat narrow window of midgestational human development studied here will be complemented by additional atlases of earlier and later time points, as well as similarly comprehensive profiling and integration of data from model organisms. The continued development and application of methods for ascertaining gene expression and chromatin accessibility—in concert with spatial, epigenetic, proteomic, lineage history, and other information—will be necessary to obtain a comprehensive view of the temporal unfolding of human cell type diversity that begins at the single-cell zygote. An interactive website facilitates the exploration of these freely available data by tissue, cell type, or gene (descartes.brotmanbaty.org). A human cell atlas of fetal gene expression enables the exploration of in vivo gene expression across diverse cell types. We used a three-level combinatorial indexing assay (sci-RNA-seq3) to profile gene expression in ~4,000,000 single cells from 15 fetal organs. This rich resource enables, for example, the identification and annotation of cell types, cross-tissue integration of broadly distributed cell types (e.g., blood, endothelial, and epithelial), and interspecies integration of mouse embryonic and human fetal cell atlases. PCR, polymerase chain reaction. The gene expression program underlying the specification of human cell types is of fundamental interest. We generated human cell atlases of gene expression and chromatin accessibility in fetal tissues. For gene expression, we applied three-level combinatorial indexing to >110 samples representing 15 organs, ultimately profiling ~4 million single cells. We leveraged the literature and other atlases to identify and annotate hundreds of cell types and subtypes, both within and across tissues. Our analyses focused on organ-specific specializations of broadly distributed cell types (such as blood, endothelial, and epithelial), sites of fetal erythropoiesis (which notably included the adrenal gland), and integration with mouse developmental atlases (such as conserved specification of blood cells). These data represent a rich resource for the exploration of in vivo human gene expression in diverse tissues and cell types.

中文翻译：

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胎儿基因表达的人类细胞图谱

人类发展的基因组学 了解人类发展的轨迹需要了解基因的调控和表达方式。现在有两篇论文提出了一种合并方法，使用三个级别的组合索引来检查胎儿样本中 15 个器官的单细胞基因表达和染色质图谱。曹等人。专注于测量广泛分布的细胞类型中的 RNA，并提供对器官特异性的见解。多姆克等人。检查了来自这些器官的细胞的染色质可及性，并确定了调节基因表达的调节元件。这些分析共同生成了人类早期发展的综合图谱。科学，这个问题 p。eaba7721，第。eaba7612 在人类早期发育过程中在单细胞水平上检查基因表达。引言人类细胞类型的参考图谱是该领域的主要目标。在这里，我们开始使用在妊娠中期获得的不同人体组织生成基因表达（本研究）和染色质可及性（Domcke 等人，本期）的单细胞图谱。基本原理 人类体内发育的分子基础的现代知识主要来自人类遗传学、模型生物体内研究和分化人类细胞系的体外研究的组合，而不是通过对发育中的人体组织的直接研究。从历史上看，一些挑战限制了在分子水平上开发人体组织的研究，包括访问受限、组织降解和细胞类型异质性。对于本研究和配套研究（Domcke 等，本期），我们能够克服这些挑战。结果 我们将基因表达的三级单细胞组合索引 (sci-RNA-seq3) 应用于 121 个人类胎儿样本，估计受孕后年龄范围为 72 至 129 天，代表 15 个器官，总共分析了 400 万个单细胞。我们开发并应用了一个用于量化细胞类型特异性的框架，识别了 657 个细胞亚型，我们根据与小鼠细胞图谱的交叉匹配对其进行了初步注释。我们在意想不到的组织中鉴定并验证了潜在的循环滋养细胞样和肝细胞样细胞。分析不同组织中的基因表达有助于对广泛分布的细胞类型（包括血液、内皮细胞和上皮细胞）进行跨组织分析。对于血细胞，这产生了从造血干细胞到所有主要亚系的细胞状态轨迹的多器官图谱。多项证据支持肾上腺是胎儿发育过程中正常但微不足道的红细胞生成部位。尽管物种和发育阶段存在差异，但将这些人类胎儿数据与小鼠胚胎细胞图谱整合起来非常简单。对于某些系统，这基本上允许我们将基因表达动态从哺乳动物发育的胚胎阶段连接到胎儿阶段。结论 这里介绍的单细胞数据资源以其规模、对人类胎儿发育的关注、所分析的组织的广度以及基因表达的并行生成（本研究）和染色质可及性数据（Domcke 等人，这问题）。我们进一步巩固了各个实验室的技术框架，以从数百万个单细胞中生成和分析基因表达和染色质可及性数据。展望未来，我们设想这里研究的中期人类发育的有点狭窄的窗口将得到更早和更晚时间点的额外地图集的补充，以及来自模型生物的数据的类似综合分析和整合。确定基因表达和染色质可及性的方法的持续发展和应用——与空间、表观遗传、蛋白质组学、谱系历史和其他信息相一致——对于全面了解人类细胞类型多样性的时间展开是必要的在单细胞受精卵。一个交互式网站有助于按组织、细胞类型或基因 (descartes.brotmanbaty.org) 探索这些免费可用的数据。胎儿基因表达的人类细胞图谱使探索不同细胞类型的体内基因表达成为可能。我们使用三级组合索引分析 (sci-RNA-seq3) 来分析来自 15 个胎儿器官的约 4,000,000 个单细胞中的基因表达。例如，这种丰富的资源能够识别和注释细胞类型、广泛分布的细胞类型（例如血液、内皮细胞和上皮细胞）的跨组织整合，以及小鼠胚胎和人类胎儿细胞图谱的种间整合。PCR，聚合酶链反应。人类细胞类型规范背后的基因表达程序具有根本的意义。我们在胎儿组织中生成了基因表达和染色质可及性的人类细胞图谱。对于基因表达，我们对代表 15 个器官的 >110 个样本应用了三级组合索引，最终分析了约 400 万个单细胞。我们利用文献和其他地图集来识别和注释组织内和组织间的数百种细胞类型和亚型。我们的分析侧重于广泛分布的细胞类型（例如血液、内皮细胞和上皮细胞）的器官特异性特化、胎儿红细胞生成部位（尤其包括肾上腺）以及与小鼠发育图谱的整合（例如血液的保守规格）细胞）。这些数据代表了探索不同组织和细胞类型中体内人类基因表达的丰富资源。