Improving cell fate conversion by tracking cells and RNA over time and space

 描述(申请人提供)：体外操纵细胞命运，无论是通过核重新编程还是分化，仍然是一个不精确和低效的过程。提高体外细胞命运转换的准确性和效率对再生医学具有重要意义。为了更好地控制细胞的命运转换，重要的是确定驱动单个细胞达到理想谱系的关键基因或分子途径，并将它们与处于不希望看到的状态的细胞中的基因或途径的活动进行比较。这将产生一个候选基因/路径的简短列表，可以干预这些候选基因/路径，以实现更精确的细胞命运转换。为此，沿着时间轴对齐单个细胞的完整转录组信息，并在后期将分子特征与细胞表型联系起来是至关重要的。在这个项目中，我们将开发基于细胞谱系追踪和单细胞全转录组测序的策略，并将其应用于使用新的人类细胞去分化模型来研究细胞去分化的转录调控，这对心血管疾病的治疗具有明确的意义。第一个目标是建立一个基于随机荧光标记、单细胞分离和转录组分析的单细胞谱系追踪技术平台。在第二个目标中，我们将基于部分去分化和再分化的概念来研究两种细胞命运转换过程。

 DESCRIPTION (provided by applicant): In vitro manipulation of cell fate, either through nuclear reprogramming or differentiation, remains to be an imprecise and inefficient process. Improving the accuracy and efficiency of in vitro cell fate conversion is of great interest to regenerative medicine. In order to gain a better control of cell fate conversion, it is important t identify the key genes or molecular pathways that drive individual cells to the desirable lineage, and to contrast them with the activities of the genes or pathways in the cells at the undesirable states. This will generate a short list of candidate genes/pathways that can be intervened for a more precise cell fate conversion. For this purpose, it is crucial to align the full transcriptome information of single cells along the time axis, and to connect the molecular signatures to the cellular phenotypes at the later stages. In this project, we will develop strategies based on cell lineage tracing and single-cell full transcriptome sequencing, and will apply it to the investigatin of the transcriptional regulation underlying cell dedifferentiation using novel models of human cells dedifferentiation with clear implications for the treatment of cardiovascular diseases. The first aim involves establishing a technical platform for single cell lineage tracing based on stochastic fluorescent labeling, single- cell isolation and transcriptome analysis. In the second aim we will investigate two cell-fate conversion procedures based on the concept of partial de-differentiation and re-differentiation.