Kinetic Barriers of Transdifferentiation

摘要 人成纤维细胞向诱导神经元和其他细胞类型的转分化， 对细胞生物学的许多实际应用和基本理解产生重大影响。它 表明细胞类型的同一性在很大程度上是由细胞的转录决定和维持的 调控网络，它可以很容易地通过一些关键的转录因子重新布线，以指导相同的 基因组来表达不同的细胞类型。这些相对快速的转换效率低， 通常在几天内表现出来，这表明除了必要的条件之外， 必须满足转录因子以实现高效的细胞重编程。鉴定 这些动力学屏障将揭示细胞谱系重编程的重要机制 并产生高效的方法来产生许多不同类型的有用细胞， 可用的细胞如成纤维细胞。越来越多的证据表明， 人类和动物在生物医学研究中的许多方面。因此， 开发人类细胞模型，研究与任务高度相关的各种人类疾病 退伍军人事务部为此，我们显著改进了技术， 通过确定的因子（Ascl 1，Nurr 1， Lmx 1a和miR-124）。我们发现，细胞周期阻滞在G1期和p53基因敲低联合 与适当的细胞外环境显着提高了转化效率， 人原代成纤维细胞诱导多巴胺能神经元。在本建议中，我们会研究 细胞周期阻滞、p53敲低和细胞外环境的分子机制 影响了程序的重组从研究中获得的知识将有助于我们了解 细胞谱系决定的基本可塑性，并将提供机制的见解， 将人成纤维细胞直接转分化为可用于许多生物医学领域的细胞 对退伍军人事务部的使命很重要的研究。

Abstract The transdifferentiation of human fibroblasts to induced neurons and other cell types has significant impact on many practical applications and fundamental understanding of cell biology. It suggests that cell type identity is largely determined and maintained by the cell's transcription regulatory network, which can be readily rewired by a few key transcription factors to direct the same genome to express a different cell type. The low efficiency of these relatively fast conversions, which generally manifest themselves within days, suggests that conditions additional to the requisite transcription factors must be met to enable highly efficient cellular reprogramming. Identification of these kinetic barriers would reveal significant mechanistic insights into cell lineage reprogramming and produce highly efficient ways to generate many different types of useful cells from readily available cells such as fibroblasts. Increasing evidence indicates that there are significant differences between human and animals in many aspects of biomedical research. It is thus very important to develop human cell models to study various human diseases that are highly relevant to the missions of the Department of Veterans Affairs. To that end, we have significantly improved the technique to reprogram human fibroblasts to induced dopaminergic neurons by defined factors (Ascl1, Nurr1, Lmx1a and miR-124). We found that cell cycle arrest at G1 phase and p53 knockdown in conjunction with the appropriate extracellular environment dramatically increased the efficiency of converting human primary fibroblasts to induced dopaminergic neurons. In this proposal, we will study the molecular mechanisms by which cell cycle arrest, p53 knockdown and extracellular environment affect the reprogramming process. Knowledge gained from the study will help us to understand the fundamental plasticity of cell lineage determination and will provide mechanistic insights into the direct transdifferentiation of human fibroblasts to cells that are useful for many areas of biomedical research important for the mission of the Department of Veterans Affairs.