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新生组织合成，还需要了解胚泡形态的模式机制。本项目旨在阐明由生物物理表观遗传变化控制的调控网络，这些变化使胚泡相对于现有组织的极性取向。我们的假设是，在涡虫再生过程中，H、K atp酶转运体的离子运输和间隙连接通讯（GJC）共同调节了传统信号上游的前后极性。目的1是通过对H，K atp酶（已完成）和GJC活性抑制的再生蠕虫的微阵列分析，鉴定和表征离子运输下游的转录靶点。候选人将使用特定标准进行选择，通过表达分析进行验证，并使用原位杂交和表型分子标记分析进行RNAi敲除后的功能表征。目的2是建立和测试模型，概述离子转运体在再生组织中用于控制轴向身份的机制。通过识别网络节点，将生成离子运输下游的初步转录网络，使用定量PCR测试RNAi敲低单个候选基因如何影响所有其他候选基因的表达。然后将生成和测试该网络预测的假设，以阐明在再生极性建立过程中对相关离子转运体活性的转录和信号反应。长期目标是编译和验证整合生物物理和生化信号的系统级再生网络模型。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。尽管新组织的再生至关重要，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。本研究旨在通过了解再生扁虫如何决定在截肢后是否形成新的头部和/或尾巴，来发现新组织和旧组织如何沟通以调节这种模式。

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.