Single cell dynamics on a whole organism scale

项目总结 哺乳动物器官的功能是由单个细胞的行为和动态维持的。这个 系统地绘制每种细胞类型的时间动态图的能力对于理解许多 哺乳动物在发育过程中经历的生物变化的各个方面。然而，传统的方法是 受吞吐量不足和可测量的蜂窝内容物范围有限的限制。而当 单细胞基因组技术已经被开发用来表征细胞状态的异质性 分辨率，几乎所有这样的方法都只在单个时间点捕获静态快照，同时具有时态和 细胞分离过程中丢失的空间信息。在这里，拟议的项目旨在开发新的方法 这使得能够全面地查看整个生命周期中的单细胞时空动态 哺乳动物有机体。具体地说，我将在高通量单细胞rna-seq平台(sci-rna- SEQ)，开发了一种同时分析转录组、表观基因组和细胞时间组的新方法 数百万个细胞中的每一个细胞的动力学(例如，增殖、凋亡)。这项技术将被用于 研究衰老如何通过系统地监测单个细胞来调节整个哺乳动物身体的状态 在幼年和老年小鼠的广泛组织中的状态动力学。这种方法将是强大的 因为我们不仅可以可视化每种细胞类型的体内增殖和凋亡行为，而且还可以 剖析其与内部转录组/表观基因组状态的关系。除了内部分子 程序中，细胞状态动态由组织结构的各个方面控制，如细胞-细胞相互作用和 细胞外基质丰度。为了以高吞吐量和准确性描述单细胞微环境，我们 将开发一种名为“微组织序列”的新技术，用于共同分析单细胞分子内容物、细胞 空间相互作用和细胞外基质(ECM)蛋白质在一个 单次实验。我们将使用这项技术来询问细胞微环境是如何调节的 不同年龄组小鼠的组织尺度细胞状态动力学。总体而言，拟议项目将 建立一个全面剖析单细胞时空动力学的技术框架 整个哺乳动物有机体的规模是前所未有的。通过分析特定于单元状态的动态行为 在老鼠的整个寿命中，这些技术和实验将独特地使准确的建模成为可能 在单个细胞中，哺乳动物系统随年龄的维持和崩溃的精致程序 决议。这些多管齐下的方法也为理解全球分子打开了一个新的范式 在衰老过程中调节细胞状态和动态的程序，从而通知潜在的途径来延缓 衰老过程以及合理设计有效的治疗方法来恢复患者的组织稳态 患有与衰老相关的疾病。

PROJECT SUMMARY The functions of mammalian organs are maintained by the behaviors and dynamics of individual cells. The ability to systematically map each cell type's temporal dynamics is central to the understanding of many aspects of biological changes that mammals undergo in development. However, conventional methods are restricted by inadequate throughput and the limited range of cellular contents that can be measured. While single-cell genomic techniques have been developed to characterize cell state heterogeneity with high resolution, nearly all such methods capture only a static snapshot at a single time point, with both temporal and spatial information lost during cell isolation. Herein, the proposed projects aim to develop novel methodologies that enable a comprehensive view of single-cell spatiotemporal dynamics across the lifespan of an entire mammalian organism. Specifically, I will expand on the high-throughput single-cell RNA-seq platform (sci-RNA- seq), to develop a novel method for concurrently profiling transcriptome, epigenome, and cellular temporal dynamics (e.g., proliferation, apoptosis) in each of millions of cells. The technique will be employed to investigate how aging regulates the status of a whole mammalian body by systematically monitoring single cell state dynamics across a broad range of tissues in young and aged mice. This approach will be powerful because we can not only visualize in-vivo proliferation and apoptosis behaviors of each cell type but also dissect its connection with internal transcriptome/epigenome states. In addition to the internal molecular programs, cell state dynamics are controlled by aspects of tissue architecture such as cell-cell interactions and extracellular matrix abundance. To profile single cell microenvironment with high throughput and accuracy, we will develop a novel technique called "microtissue-seq", for co-profiling single-cell molecular contents, cellular spatial interactions, and extracellular matrix (ECM) proteins across tens of thousands of spatial locations in a single experiment. We will employ this technique to interrogate how cellular microenvironment regulates organismal-scale cell state dynamics in different age groups of mice. Overall, the proposed projects will establish a technical framework for comprehensive profiling single-cell spatiotemporal dynamics at an unprecedented scale of a whole mammalian organism. By profiling cell-state specific dynamic behaviors across the lifespan of mice, these technologies and experiments would uniquely enable accurate modeling of the exquisite program underlying mammalian system maintenance and breakdown with age at single cell resolution. These multi-pronged approaches also open a new paradigm for understanding the global molecular programs regulating cell states and dynamics during aging, thereby informing potential pathways to delay the aging process as well as the rational design of effective therapies to restore tissue homeostasis for patients with aging-related diseases.