Decoding the bridges and barriers to cellular reprogramming and lineage identity

解码细胞重编程和谱系身份的桥梁和障碍

• 批准号: 9790532
• 负责人: Rajan Jain
• 金额: $ 114.26万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-19 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9790532
• 关键词: Adopted; Automobile Driving; Back; Biological; Biology; CRISPR/Cas technology; Cardiac; Cardiac Myocytes; Cell Fate Control; Cells; Computer Analysis; DNA; Development; Developmental Biology; Elements; Exposure to; Frequencies; Gene Delivery; Gene Expression; Genes; Genetic; Genetic Screening; Genetic Transcription; Genomics; Goals; Human body; Image; Individual; Knowledge; Maps; Measures; Methodology; Methods; Molecular; Molecular Profiling; Nature; Pathway interactions; Pharmaceutical Preparations; Phenotype; Population; RNA; Regenerative Medicine; Research; Rest; Series; Sorting - Cell Movement; Source; Techniques; Technology; Thinking; Time; Viral; Work; base; cell transformation; cell type; clinical application; experimental study; innovation; interest; new technology; response; screening; small molecule; tool; transdifferentiation; translational approach

Project Summary The human body is composed of a large number of cell types. These types are generally quite stable in the sense that cells of one type, once established, typically do not switch to other cell types. A major goal for regenerative medicine and our understanding of cell type in general is to discover if and how we can force cells to switch from one type to another. Recent results over roughly the last decade have shown that it is in principle possible to convert cells from one type by forcing them to turn on small sets of genes. However, in the vast majority of cases, we have no idea what these sets of genes are out of the many thousands of potential ones, and so our understanding has largely been dictated by picking candidates based on prior knowledge. Furthermore, even when these sets of genes are identified, the efficiency of interconversion of cell type is very low, with only a small percentage of source cells converting to the target type. Our proposed research tackles both of these challenges using a combination of new concepts of cell identity and new technology for tracing individual cells back in time. For cell identity, the approach most common in the field is to use profiles of which genes are on or off in any particular cell type to determine lineage-specific factors. However, while these genes are lineage-specific, they may not be lineage-determining in the sense that they may not drive a cell to a particular type per se. We have developed a methodology we call PerturbID that uses a series of systematic perturbations to identify specific genes that turn on and off in response, which we have shown have the capacity to interconvert cells. We propose to use PerturbID to identify and validate candidates for cell type transformation across a range of cell types, ultimately arriving at a set of general principles for cellular reprogramming. This will have applications for regenerative medicine as well as across biology as a whole. The other major problem, of efficiency, has remained mysterious because nobody currently knows why some cells are capable of reprogramming while the vast majority are not. The challenge is the lack of tools for retrospective profiling of cells: how do we rewind time to profile the cells that will ultimately adopt a different fate? We have developed tools for performing this retrospective profiling. We will apply this “time machine” methodology to the problem of inefficient reprogramming to determine the unique signature of cells primed for cell type conversion, and will perform genetic screens to isolate pathways capable of manipulating this frequency. Together, our work will transform our concepts of cell type and will have enormous practical implications for its application in regenerative medicine.

项目概要 人体由大量细胞类型组成。这些类型一般都相当稳定 感觉一种类型的细胞一旦建立，通常不会转换为其他细胞类型。一个主要目标是 再生医学和我们对细胞类型的一般理解是为了发现我们是否以及如何能够迫使细胞 从一种类型切换到另一种类型。大约过去十年的最新结果表明，它处于 通过迫使细胞开启一小组基因，可以将细胞从一种类型转变为一种原理。然而，在 绝大多数情况下，我们不知道这些基因组在数千个潜在基因中是什么 因此，我们的理解很大程度上取决于根据先验知识挑选候选人。 此外，即使这些基因组被识别出来，细胞类型相互转换的效率也非常高。 低，只有一小部分源单元格转换为目标类型。 我们提出的研究结合了细胞身份的新概念来解决这两个挑战 以及及时追踪单个细胞的新技术。对于细胞识别，最常见的方法是 领域是使用任何特定细胞类型中基因开启或关闭的概况来确定谱系特异性 因素。然而，虽然这些基因是谱系特异性的，但它们在某种意义上可能不是谱系决定性的。 它们可能不会将细胞本身驱动为特定类型。我们开发了一种称为 PerturbID 的方法 它使用一系列系统扰动来识别响应中打开和关闭的特定基因， 我们已经证明了具有相互转换细胞的能力。我们建议使用 PerturbID 来识别和验证 跨一系列细胞类型进行细胞类型转化的候选者，最终得到一组通用的 细胞重编程原理。这将应用于再生医学以及跨领域 生物学作为一个整体。另一个主要问题，即效率，仍然是个谜，因为目前没有人 知道为什么有些细胞能够重新编程，而绝大多数细胞却不能。挑战在于缺乏 用于细胞回顾性分析的工具：我们如何倒带时间来分析最终采用细胞分析的细胞 不同的命运？我们开发了用于执行这种回顾性分析的工具。我们将应用这个“时间 机器”方法来解决低效重编程问题，以确定细胞的独特特征 为细胞类型转换做好准备，并将进行遗传筛选以分离能够操纵的途径 这个频率。我们的工作将共同改变我们对细胞类型的概念，并将具有巨大的实际意义 及其在再生医学中的应用的影响。