Biophysical Regulation of Axial Polarity During Regeneration

描述（由申请人提供）：（1）干细胞和再生研究的临床应用不仅需要了解从头组织合成，还需要了解调控芽基形态的模式化机制。该项目旨在阐明生物物理表观遗传变化控制的调控网络，这些变化使芽基极性相对于现有组织定向。我们的假设是，离子运输的H，K ATP酶转运蛋白和间隙连接通讯（GJC）一起调节前-后极性在涡虫再生的传统信号上游。目的1是通过对再生蠕虫的H，K ATP酶（完全）和GJC活性抑制的微阵列分析，鉴定和表征离子转运下游的转录靶点。将使用特定标准选择候选物，通过表达分析进行验证，并在RNAi敲除后使用原位杂交和表型的分子标记分析进行功能表征。目的2是建立和测试模型，概述了离子转运蛋白用于控制再生组织中的轴向身份的机制。离子转运下游的初步转录网络将通过识别网络节点来生成，使用定量PCR来测试单个候选物的RNAi敲低如何影响所有其他候选物的表达。然后将生成和测试由该网络预测的假设，以阐明再生极性建立期间对所涉及的离子转运蛋白的活性的转录和信号响应。长期目标是编译和验证系统级再生网络模型，整合生物物理和生化信号。这有可能用作形态发生的预测模型，以帮助将基础再生研究转化为实际的生物医学治疗。(2)干细胞和再生研究的前景是最终取代受伤、衰老或癌症患者失去或受损的组织、器官和肢体。虽然新组织的再生是至关重要的，但要使该组织发挥功能，它必须与现有组织形成适当的模式：这项研究旨在通过学习再生扁形虫如何确定截肢后是否形成新的头部和/或尾部，来发现新旧组织如何交流以调节这种模式。

DESCRIPTION (provided by applicant): (1) Clinical applications arising from stem cell and regeneration research will require an understanding not only of de novo tissue synthesis but also of the patterning mechanisms regulating blastema morphology. This project seeks to elucidate the regulatory networks controlled by biophysical epigenetic changes which orient blastema polarity relative to existing tissues. Our hypothesis is that ion transport by the H,K ATPase transporter and gap junctional communication (GJC) together regulate anterior-posterior polarity during planarian regeneration upstream of traditional signaling. Aim 1 is to identify and characterize transcriptional targets downstream of ion transport, through microarray analyses of regenerating worms inhibited for H,K ATPase (completed) and GJC activity. Candidates will be selected using specific criteria, validated by expression analyses, and functionally characterized following RNAi knockdown using in situ hybridization and molecular marker analyses of phenotypes. Aim 2 is to build and test models which outline the mechanisms ion transporters use to control axial identity in regenerating tissue. A preliminary transcriptional network downstream of ion transport will be generated by identifying network nodes, using quantitative PCR to test how RNAi knockdown of a single candidate affects the expression of all other candidates. Hypotheses predicted by this network will then be generated and tested, to elucidate the transcriptional and signaling response to the activity of the implicated ion transporters during the establishment of regenerative polarity. The long-term goal is to compile and validate a systems-level regeneration network model integrating biophysical and biochemical signaling. This has the potential for use as a predictive model of morphogenesis to aid in the translation of basic regeneration research into practical biomedical therapies. (2) The promise of stem cell and regeneration research is ultimately to replace lost or damaged tissues, organs and limbs in patients suffering from injury, aging or cancer. Although the regeneration of new tissue is critical, for that tissue to be functional, it must be properly patterned in relationship to the existing tissues: This research aims to discover how new and old tissues communicate to regulate this pattering, by learning how regenerating flatworms determine whether to form a new head and/or tail following amputation.