A protodermal miR394 signal defines a region of stem cell competence in the Arabidopsis shoot meristem

植物在其整个生命过程中形成新的器官，在极端情况下可以持续数千年以上，通过其生长中心分生组织中的多能干细胞的活动。揭示维持分生组织活动的机制是理解植物生长和发育的核心。为了维持茎的生长，尽管所有的分生组织细胞分裂并且分化细胞离开分生组织，但是茎分生组织的空间组织（其产生所有的气生器官）必须被稳定地保存。干细胞龛位于茎分生组织中心的最外层细胞层，而分化发生在周围区域。因此，茎分生组织代表了生物学中的一个基本问题，即在分裂的细胞群体中维持功能组织。在传统的遗传筛选中，没有发现将干细胞能力固定在植物顶端的因素，可能是由于遗传冗余。为了规避这一限制，我们进行了几个遗传增强子和抑制筛选新的干细胞调节剂在模式植物拟南芥在我们的准备工作，这一建议。我们通过对一个突变体的分析发现，原表皮（茎分生组织的最外层）在干细胞调控中起着迄今为止未知的信号中心的作用。它将microRNA miR 394以梯度形式提供给底层干细胞，其中miR 394抑制F-Box蛋白叶子卷曲反应（LCR）。这种抑制对于通过转录因子WUSCHEL（WUS）维持干细胞是必要的，该转录因子从干细胞下方向上移动。这些发现的含义是，原表皮作为最外层的细胞层的茎分生组织可以作为一个稳定的来源图案的信息，虽然其居民细胞可能会发生变化。向内（miR 394）和向外（WUS）的干细胞群体的相对两极产生的信号的相互作用提供了一个新的概念，干细胞龛是如何稳定地锚定在自组织芽分生组织。该提案的目标是研究miR 394如何调节茎分生组织中干细胞感受态区域的分子机制。我们将探索miR 394功能的空间、时间和剂量学参数，将miR 394信号传导整合到茎分生组织的调控网络中，并研究LCR如何抑制干细胞命运的分子机制。从这个项目中获得的结果也将用于与弗莱克教授（瓦赫宁根）合作，开发芽分生组织维持的数学模型。该项目不仅将解决干细胞生物学中的一个基本问题，而且有可能有助于我们理解经济上重要的植物中基于干细胞的再生策略。

Plants form new organs throughout their life, which in extreme cases can last more than a thousands years, by the activity of pluripotent stem cells in their growth centers, the meristems. Unraveling the mechanisms that maintain meristem activity is central to understand plant growth and development. To maintain shoot growth, the spatial organization of the shoot meristem, which gives rise to all aerial organs, must be stably preserved despite the fact that all meristem cells divide and differentiating cells leave the meristem. The stem cell niche is positioned at the outermost cell layers in the shoot meristem center, whereas a differentiation occurs in the surrounding areas. The shoot meristem thus represents a fundamental problem in biology, the maintenance of a functional organisation in a dividing cell population. In conventional genetic screens, factors fixing stem cell competence to the tip of the plant have not been discovered, presumably due to genetic redundancy. To circumvent this limitation, we performed several genetic enhancer and suppressor screens for novel stem cell regulators in the model plant Arabidopsis thaliana in our preparatory work for this proposal. We discovered through the analysis of one mutant that the protoderm (the outermost layer of the shoot meristem) functions as a hitherto unknown signaling center in stem cell regulation. It supplies the microRNA miR394 in a gradient to the underlying stem cells, where miR394 inhibits the F-Box protein LEAF CURLING RESPONSIVENESS (LCR). This repression is necessary to enable stem cell maintenance through the transcription factor WUSCHEL (WUS), which moves upward from underneath the stem cells. The implication of these findings is that the protoderm as the outermost cell layer of the shoot meristem can act as a stable source of patterning information, although its resident cells may change. The interaction of inward (miR394) and outward (WUS) signals produced from opposite poles of the stem cell population provides a novel concept for how the stem cell niche is stably anchored in the self organizing shoot meristem. The goal of this proposal is to investigate the molecular mechanisms of how miR394 regulates the region of stem cell competence in the shoot meristem. We will explore the spatial, temporal, and dosimetric parameters of miR394 function, integrate miR394 signaling into the regulatory network of the shoot meristem, and investigate the molecular mechanism of how LCR inhibits stem cell fate. The results obtained from this project will also be used in collaboration with Prof. Fleck (Wageningen) to develop mathematical models for shoot meristem maintenance. This project will not only address a fundamental question in stem cell biology, but also has the potential to contribute to our understanding of stem cell-based regeneration strategies in economically important plants.