CAREER: Linking systemic stem cell activation to vertebrate limb regeneration

本奖项全部或部分由《2021年美国救援计划法案》（公法117- 2）资助。蝾螈在截肢后可以完全再生它们的腿。尽管经过了几个世纪的研究，它们是如何再生的仍然是个谜。人类和其他哺乳动物的肢体再生能力非常有限，仅限于手指和脚趾的尖端。与此同时，一些无脊椎动物，比如涡虫，几乎身体的每个部位都能再生。再生能力的巨大差异也没有得到很好的理解。这个项目旨在填补关于蝾螈如何再生四肢的缺失信息。它还将讨论蝾螈的这些过程与其他超级再生动物的再生过程和哺乳动物的损伤反应有多相似。最近在蝾螈身上发现了一种对截肢的系统性反应，其中全身的一些细胞被激活以产生新的细胞。这些细胞的身份和功能将被解决。最近的数据还表明，神经系统传递的信号被远处的细胞用来感知截肢。该项目将测试在神经中检测到的几种分子，以确定它们是否对全身损伤反应是必需的。它还将解决在蝾螈血液中传播截肢损伤信号的分子的作用。最后，该项目将在蝾螈环境下研究涡虫和小鼠的全身损伤反应的重要分子。总之，这些实验将有助于理解系统反应如何与身体部位的局部再生联系在一起。它们还将填补关于再生过程进化的知识空白，并将为未来的再生医学方法提供信息。该项目还将通过新课程和实验室交换计划培养许多本科生。最后，它将通过每周采访来自不同背景的科学家的网络纪录片向公众开放。随着近年来各种分子遗传学工具在蝾螈中的应用，截肢反应和随后的肢体再生终于可以在机制水平上理解。该项目旨在确定系统祖细胞激活（定义为细胞周期再进入）与蝾螈肢体再生之间的联系。系统活化细胞的命运和身份将利用体内整合逆转录病毒、转基因蝾螈和单细胞RNA-seq进行研究。神经和循环系统在系统激活中的具体作用也将通过实验进行检验，使用功能获得和功能丧失的方法，并基于来自周围神经和反应组织以及血浆蛋白质组学的RNA-seq数据。该项目还将测试先前确定的因子（如蝾螈系统中的EGR1、wnt和HGF）在纯肠动物和小鼠的损伤反应中可能发挥的保守作用。总之，这些实验将为一个正在发展的模型提供信息，在这个模型中，系统性损伤反应可能代表了在进化过程中局部损伤部位不同阐述的祖先过程，在现存物种的大范围再生结果中表现出来。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

This award is funded in whole or in part under the American Rescue Plan Act of 2021 (Public Law 117- 2). Salamanders can completely regenerate their legs following amputation throughout life. How they regenerate is still mysterious despite centuries of research. Humans and other mammals have very limited natural limb regeneration abilities, restricted to just the very tips of the fingers and toes. Meanwhile, some invertebrates, like planarians, can regrow almost every body part. The huge variation in regenerative ability is also not well understood. This project aims to fill in missing information about how salamanders regenerate limbs. It will also address how similar these processes in salamanders are to regenerative processes in other super-regenerators and to injury responses in mammals. A systemic response to amputation was recently discovered in axolotl salamanders in which some cells throughout the body are activated to make new cells. The identity and function of these cells will be addressed. Recent data also indicates the nervous system transmits signals that distant cells use to sense an amputation. This project will test several molecules detected in nerves to determine if they are required for the body-wide injury response. It will also address the role of molecules that travel in the blood in transmitting amputation injury signals in axolotls. Finally, the project will examine molecules important for body-wide injury responses in planarians and mice in the axolotl context. Together, these experiments will build understanding of how systemic responses are linked to local regeneration of a body part. They will also fill in gaps in knowledge about evolution of regenerative processes, and they will inform future regenerative medicine approaches. This project will also train many undergraduate students in a new course and through a laboratory exchange program. Finally, it will reach out to the public with a weekly web documentary that interviews scientists from diverse backgrounds.With the recent application of a variety of molecular genetic tools in axolotl, amputation responses and subsequent limb regeneration can finally be understood at a mechanistic level. This project seeks to define the connections between systemic progenitor cell activation, defined as cell-cycle re-entry, and limb regeneration in axolotl. The fate and identity of systemically-activated cells will be investigated using integrative retroviruses in vivo as well as transgenic axolotls and single-cell RNA-seq. The specific roles of the nervous and circulatory systems in systemic activation will also be examined experimentally, using both gain-of-function and loss-of-function approaches, and building on RNA-seq data from peripheral nerves and responding tissues as well as plasma proteomics. The project will also test for possible conserved roles of previously identified factors in injury responses in both planarians and mice, such as EGR1, Wnts, and HGF, in the axolotl system. Together, these experiments will inform a developing model in which systemic injury responses may represent ancestral processes that have been differentially elaborated upon at the local injury site during evolutionary time, manifesting in a large range of regenerative outcomes in extant species.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.