Kinetic Barriers of Transdifferentiation

摘要 人成纤维细胞向诱导神经元和其他类型细胞的转分化 对许多实际应用和对细胞生物学的基本理解产生重大影响。它 表明细胞类型的一致性在很大程度上是由细胞的转录决定和维持的 调控网络，它可以很容易地被几个关键的转录因子重新连接起来，以指导相同的 基因组表达一种不同的细胞类型。这些相对较快的转换效率较低， 通常在几天内显现，表明除了必要条件之外的条件 必须满足转录因子的要求才能实现高效的细胞重新编程。身份识别 这些动力学障碍将揭示对细胞谱系重新编程的重大机械论见解 并产生高效的方法来从容易地产生许多不同类型的有用细胞 可利用的细胞，如成纤维细胞。越来越多的证据表明， 人与动物之间在生物医学研究的许多方面。因此，非常重要的是 开发人体细胞模型以研究与任务高度相关的各种人类疾病 退伍军人事务部的。为此，我们对技术进行了显著改进，以 根据确定的因子(Ascl1，Nurr1， Lmx1a和miR-124)。我们发现细胞周期停滞于G1期与P53基因下调有关 与适当的胞外环境极大地提高了转换效率 人原代成纤维细胞诱导多巴胺能神经元。在这项建议中，我们将研究 细胞周期停滞、P53基因敲除与细胞外环境的分子机制 影响重新编程过程。从研究中获得的知识将有助于我们理解 细胞谱系决定的基本可塑性，并将提供对 人成纤维细胞直接转分化为对许多生物医学领域有用的细胞 研究对退伍军人事务部的使命非常重要。

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.