Regulation of Appendage Regeneration in Zebrafish

描述（由首席研究员提供）：成年硬骨鱼和尾形两栖动物有能力再生整个被截肢的附肢。与之形成鲜明对比的是，哺乳动物四肢的再生愈合仅限于趾尖。在尾尾动物的肢体再生和硬骨鱼的鳍再生过程中，再生过程受到精确的调控，只有适当的结构才会被替换。附属物再生的一个经典和未解释的问题是受伤区域的成年细胞如何保留或识别完成这一过程所必需的位置信息。维持这种“位置记忆”的核心问题是：首先，在再生过程中，细胞在多大程度上保留了它们的谱系限制和分化状态？也就是说，一个谱系的分化的成年细胞是否能够为一个或多个其他谱系做出贡献，还是采取一种分化程度较低的形式？第二，哪些特定的分子程序对于保留和调节这些细胞内的位置信息至关重要？在这里，我们建议定义新的细胞和分子调节机制，以维持位置信息并指导斑马鱼鳍，包括骨，结缔组织间质，表皮，血管，神经和色素细胞的再生更新。在这个建议中，我们描述了一种位置记忆的编程方法，使用1)新的技术来追踪成年鳍细胞的谱系；2)基于新结果的候选基因检测；3)突变筛选和定位克隆的正向遗传方法。通过这种方法，我们将验证分化的成年附属物组织表达区域特异性调节因子的假设，这些调节因子维持对再生保真度很重要的位置信息。我们的分子遗传学方法将增加对再生器官发生过程中活跃的调节机制的理解，并为理解甚至可能改变大多数人体器官再生能力的现有限制提供重要的视角。

DESCRIPTION (provided by Principal Investigator): Adult teleost fish and urodele amphibians have the capacity to regenerate entire amputated appendages. In striking contrast, regenerative healing of mammalian limbs is limited to the very tips of digits. During limb regeneration in urodeles and fin regeneration in teleosts, regeneration is precisely regulated such that only the appropriate structures are replaced. One of the classic and unexplained questions of appendage regeneration is how adult cells in the injured area retain or recognize the positional information necessary to accomplish this. The central questions by which this "positional memory" is maintained are these. First, to what extent do cells retain their lineage restriction and state of differentiation during regeneration? That is, are differentiated adult cells of one lineage able to contribute to one or more other lineages, or to assume a less differentiated form? Second, which specific molecular programs are essential for retaining and regulating positional information within these cells? Here, we propose to define new cellular and molecular regulatory mechanisms that maintain positional information and instruct regenerative renewal of zebrafish fins, complex organs containing bone, connective tissue mesenchyme, epidermis, blood vessels, nerves, and pigment cells. In this proposal, we describe a programmatic approach to positional memory, using 1) new technology for the lineage tracing of adult fin cells; 2) candidate gene testing based on new results; and 3) a forward genetic approach of mutagenesis screening and positional cloning. With this approach, we will test the hypothesis that differentiated adult appendage tissues express region-specific profiles of regulatory factors that maintain positional information important for regenerative fidelity. Our molecular genetic approach will increase understanding of regulatory mechanisms active during regenerative organogenesis, and provide important perspective for comprehending, and perhaps changing, the existing limitations in regenerative capacity of most human organs. PUBLIC HEALTH RELEVANCE: Our molecular genetic approach will increase understanding of regulatory mechanisms active during regenerative organogenesis, and provide important perspective for comprehending, and perhaps changing, the existing limitations in regenerative capacity of most human organs.