Engineering Morphogenetic Factors for Enhanced Genetic Reprogramming

描述（由申请人提供） 摘要：哺乳动物细胞遗传重编程的最新进展正在挑战细胞分化的传统概念，并为再生医学提供新的途径。这些研究表明，细胞转录机制可以被操纵，以重新编程基因表达谱和重定向细胞行为。通过诱导多能干细胞（iPSC）开发的最新进展，遗传重编程的原理通常也适用于控制各种各样的细胞表型。事实上，许多细胞谱系是由一个单一的主调控转录因子，其中几个目前被用来控制细胞命运的基因和细胞为基础的治疗和其他生物技术应用。然而，目前用于遗传重编程的方法受到几种低效率的限制，包括低频率的重编程细胞、实现完全重编程的时间长以及新细胞表型的稳健性不足。为了解决这些限制，我们建议通过定向分子进化来增强重编程因子的内在特性。这代表了遗传重编程领域的一个新的概念方向，并利用了定向进化领域的最新进展来增强蛋白质功能。重要的是，这种方法广泛适用于提高任何转录调控机制的功效，因此将有利于生物医学研究的许多应用和领域。这项工作的成果将包括发现具有增强重编程能力的转录因子变体，深入了解形态发生转录因子的生物化学，以及开发控制细胞行为的新方法。此外，这项研究将推动定向进化领域进入遗传重编程和再生医学的新领域。这种技术的协同结合是将生物医学研究的变革性进展转化为人类健康的真实的惠益所必需的创新。 公共卫生相关性：科学家们正在研究使用天然蛋白质来协调复杂细胞行为的方法，包括组织修复。尽管这种方法显示出巨大的希望，但这些天然蛋白质通常不足以有效地指导细胞活性。该提案概述了一种新的独特技术，用于开发这些蛋白质的工程版本，这些蛋白质将更有效地为各种应用产生强大的细胞来源，包括治疗心血管疾病，神经退行性疾病和肌肉骨骼疾病。

DESCRIPTION (Provided by the applicant) Abstract: Recent advances in the genetic reprogramming of mammalian cells are challenging the traditional concepts of cell differentiation and providing new avenues for regenerative medicine. These studies have shown that cellular transcription machinery can be manipulated to reprogram gene expression profiles and redirect cell behavior. Highlighted by recent progress in the development of induced pluripotent stem cells (iPSCs), the principles of genetic reprogramming are also generally applicable to controlling a wide variety of cellular phenotypes. In fact, many cell lineages are defined by a single master regulatory transcription factor, several of which are currently being used to control cell fate for gene- and cell-based therapies and other biotechnological applications. However, current methods for genetic reprogramming are limited by several inefficiencies, including a low frequency of reprogrammed cells, long times to achieve full reprogramming, and insufficient robustness of new cell phenotypes. To address these limitations, we propose to enhance the intrinsic properties of the reprogramming factors through directed molecular evolution. This represents a new conceptual direction in the area of genetic reprogramming and capitalizes on the recent progress in the area of directed evolution for enhancing protein function. Importantly, this approach is broadly applicable to improving the efficacy of any transcriptional regulation machinery and therefore will be beneficial to numerous applications and fields of biomedical research. The outputs of this work will include the discovery of transcription factor variants with enhanced reprogramming capabilities, insights into the biochemistry of morphogenetic transcription factors, and the development of new methods for controlling cell behavior. Additionally, this study will propel the field of directed evolution into the new areas of genetic reprogramming and regenerative medicine. This synergistic incorporation of technologies represents the innovation that will be necessary to translate the transformative advances of biomedical research into real benefits for human health. Public Health Relevance: Scientists are investigating methods to use naturally occurring proteins to coordinate complex cell behaviors, including tissue repair. Although this approach shows tremendous promise, these natural proteins are often insufficient for effectively directing cell activity. This proposal outlines a new and unique technology for evolving engineered versions of these proteins that will be more effective in generating robust cell sources for a variety of applications, including the treatment of cardiovascular disease, neurodegenerative conditions, and musculoskeletal disorders.