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传统的细胞分化概念，并为再生医学提供了新的途径。这些研究表明，细胞转录机制可以被操纵来重新编程基因表达谱并改变细胞行为。最近在诱导多能干细胞(IPSCs)的开发方面取得的进展突显了基因重编程的原理也普遍适用于控制各种细胞表型。事实上，许多细胞系由单一的主调控转录因子决定，其中几个目前被用于控制基于基因和细胞的治疗和其他生物技术应用的细胞命运。然而，目前的遗传重编程方法受到一些低效方法的限制，包括重编程细胞的频率低，实现完全重编程的时间长，以及新细胞表型的稳健性不够。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属性。这代表了基因重新编程领域的一个新的概念方向，并利用了在增强蛋白质功能的定向进化领域的最新进展。重要的是，这种方法广泛适用于提高任何转录调控机制的效率，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的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新编程和再生医学的新领域。这种技术的协同结合代表了将生物医学研究的变革性进展转化为对人类健康的真正好处所必需的创新。 与公共卫生相关：科学家正在研究使用自然产生的蛋白质来协调复杂的细胞行为的方法，包括组织修复。尽管这种方法显示出巨大的前景，但这些天然蛋白质往往不足以有效地指导细胞活动。这项提案概述了一种新的独特技术，用于进化这些蛋白质的工程化版本，将更有效地为各种应用产生强大的细胞源，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.