Mechanisms driving stem cell responses to injury in planarians

摘要 组织的成功再生需要干细胞可塑性、增殖性和 分化，以产生与先前存在的组织和器官整合的新细胞。路径 控制这些关键行为的机制已经确定，但损伤信号如何触发干细胞 再生所必需的细胞的增殖和分化仍然知之甚少。在大多数 模式生物，再生能力有限，干细胞稀缺，这使得很难 明确损伤后干细胞增殖和分化的调控机制。相比之下， 稻纵卷叶虫有丰富的干细胞，可被损伤和燃料激活 持续再生。与胚胎干细胞一样，脊椎动物干细胞也具有分化能力。 转化成任何类型的组织。这些多能干细胞可以很容易地被识别、监测、纯化和 在分子水平上彻底剖析。我们最近有了两个重要的发现，它们形成了 这项提议的基础。首先，任何类型的损伤似乎都能保护干细胞免受致命辐射的伤害， 因为它停止了细胞周期，更少的干细胞经历了凋亡。第二，我们首创了一种化学物质 方法有选择地切除单个器官，即咽部。咽部再生需要上调 紧随其后的干细胞离散亚群中保守的叉头转录因子FoxA的表达 这是一次定向伤害。我们发现，细胞外信号调节激酶(ERK)是这些变化的中心驱动因素 行为。ERK促进培养的干细胞分化，但在损伤后如何被激活尚不清楚 明白了。总而言之，这些发现确立了我们的中心假设，即损伤是同步的 细胞周期，使局部信号能够引导干细胞分化走向离散的细胞命运。在目标1中， 我们将确定损伤如何在辐射后诱导干细胞细胞周期停滞。我们将检测DNA 修复和测试损伤后上调的保守基因的功能。在目标2中，我们将剖析 干细胞纯化和单细胞测序驱动器官特异性再生的机制 器官丧失后细胞增殖。我们将鉴定这些细胞上富含的受体，并测试它们的功能 以确定它们是否作用于FoxA的上游。在目标3中，我们将确定上游 受体激活干细胞中的MAP激酶信号，结合RNAi，药理学和 生物化学。这项提议利用了我们用精确的侮辱来挑战干细胞的能力，提供了一种 透镜使灵活的干细胞反应在损伤和动态平衡的机制。 了解在生理相关疾病中控制干细胞行为的分子机制 背景将为再生医学技术的未来战略设计提供信息。

Abstract Successful regeneration of tissues requires transient increases in stem cell plasticity, proliferation, and differentiation, in order to produce new cells that integrate with preexisting tissues and organs. Pathways governing these critical behaviors have been identified, but how injury signals can trigger stem cell proliferationand differentiation of cells necessary for regeneration remains poorly understood. In most model organisms, regenerative capacity is limited and stem cells are scarce, which has made it difficult to pinpoint the mechanisms regulating stem cell proliferation and differentiation after injury. By contrast, the planarian flatworm Schmidtea mediterranea has abundant stem cells that are activated by injury and fuel continuous regeneration. Like embryonic stem cells, planarian stem cells have the capacity to differentiate into any type oftissue. These pluripotent stem cells can be readily identified, monitored, purified, and thoroughly profiled at themolecular level. We recently made two important discoveries that form the foundation of this proposal. First, injury of any type appears to protect stem cells from lethal radiation, because it halts the cell cycle and fewer stem cells undergo apoptosis. Second, we pioneered a chemical method to selectively remove a single organ,the pharynx. Pharynx regeneration requires the upregulation of the conserved Forkhead transcription factor FoxA in a discrete subset of stem cells immediately after this targeted injury. We find that the extracellular signal-regulated kinase (ERK) is a central driver of these behaviors. ERK promotes differentiation in cultured stem cells, but how it is activated after injury is poorly understood. Together, these findings establish our central hypothesis, which is that injury synchronizes the cell cycle, enabling local cues to channel stem cell differentiation toward discrete cell fates. In Aim 1, we will determine how injury induces cell cycle arrest in stemcells after radiation. We will examine DNA repair and test the function of conserved genes that are upregulatedafter injury. In Aim 2, we will dissect the mechanisms driving organ-specific regeneration by purification and single-cell sequencing of stem cells proliferating after organ loss. We will identify receptors enriched on these cells, and test their function in organ regeneration to determine if they act upstream of FoxA. In Aim 3, we will identify the upstream receptors that activate MAP kinase signaling in stem cells with combinations of RNAi, pharmacology and biochemistry. This proposal exploits our ability to challenge stem cells with precise insults, providing a lens into the mechanisms that enable flexible stem cell responses during injury and homeostasis. Understanding the molecular mechanisms that govern stem cell behavior in a physiologically-relevant context will inform the design of future strategies for regenerative medicine technologies.