Cell-Cell Signaling in Embryonic and Retinal Development

描述（由申请人提供）：这项工作的长期目标是阐明细胞整合来自多个信号通路的指令并以适当的方式响应的机制。反映了精确协调的信号传导在所有生物过程中发挥的关键作用，这些途径的不受控制或失调的活性导致人类的肿瘤发生和/或发育缺陷。我们的方法是利用遗传上易于处理的果蝇系统，以发现新的策略，保守的信号转导级联收敛于核转录因子网络，以指导细胞增殖，命运的规范，分化，形态发生和生存在视网膜发育。由于发育信号机制在进化中高度保守，因此对果蝇中使用的分子电路的了解将从根本上促进我们对细胞命运如何指定和维持以及为什么失调会导致哺乳动物癌症和疾病的理解。这项建议的目标是探索一种新的信号整合机制，我们正在进行的转录调控网络，编排视网膜的规范和发展的研究建议。特别是Eyes absent（Eya），通过其作为转录因子和蛋白酪氨酸磷酸酶（PTP）的不寻常的双重功能，具有通过指导下游基因表达模式来响应上游信号输入和通过其磷酸酶功能影响这些相同途径的活性的潜力。虽然受损的Eya PTP功能与人类疾病有关，包括眼部缺陷，但Eya PTP活性的正常生物学功能及其与Eya介导的转录调控的关系仍然知之甚少。该提案描述了一个多方面的方法相结合的遗传，生物化学和细胞生物学检测，以解决这一假设，即适当的协调和调节Eya的两个基本功能，一个作为核转录因子和一个作为磷酸酪氨酸信号网络的组成部分，是至关重要的视网膜发育。成功完成拟议的研究计划将提供对Eya PTP功能所需的细胞和发育背景的深入了解，并应揭示Eya，视网膜决定基因网络和其他对眼睛发育至关重要的信号模块之间的新的串扰机制。具体目标是通过探索与Abelson（Abl）和Src 64酪氨酸激酶的遗传和生化相互作用来阐明Eyes abscess（Eya）参与的磷酸酪氨酸信号传导途径，以研究在体外表达筛选中鉴定的新结合伴侣的贡献。Eyes abscess的两种功能是转录因子和磷酸酶，并检验核质之间的眼功能缺失的空间划分有助于视网膜正常发育的假设。由于我们正在研究的蛋白质和途径在哺乳动物中具有保守的功能，因此我们的工作揭示的新信号策略将促进对人类发育和疾病的理解。公共卫生相关性：基因功能的不适当调节是包括癌症在内的广泛的人类发育异常和疾病的基础。这项研究的目的是揭示在眼睛正常发育过程中调节关键决策的细胞机制，作为理解这些关键控制丧失时出现的疾病和缺陷的重要第一步，并最终开发有效的治疗干预措施。

DESCRIPTION (provided by applicant): The long-term objectives of this work are to elucidate the mechanisms whereby cells integrate instructions from multiple signaling pathways and respond in a context-appropriate manner. Reflecting the key role that precisely orchestrated signaling plays in all biological processes, uncontrolled or dysregulated activity of these pathways leads to tumorigenesis and/or developmental defects in humans. Our approach is to exploit the genetically tractable Drosophila system to uncover novel strategies whereby conserved signal transduction cascades converge on nuclear transcription factor networks to direct cell proliferation, fate specification, differentiation, morphogenesis and survival during retinal development. Because developmental signaling mechanisms have been highly conserved in evolution, knowledge of the molecular circuitries used in Drosophila will fundamentally advance our understanding of how cell fates are designated and maintained, and why misregulation results in cancer and disease in mammals. The goal of this proposal is to explore a novel mechanism of signal integration suggested by our ongoing study of transcriptional regulatory networks that orchestrate retinal specification and development. Specifically Eyes absent (Eya), through its unusual dual functionality as transcription factor and protein tyrosine phosphatase (PTP), has the potential both to respond to upstream signaling inputs by directing downstream patterns of gene expression and to influence the activity of these same pathways through its phosphatase function. Although impaired Eya PTP function has been implicated in human disease, including ocular defects, the normal biological function of Eya's PTP activity, and its relationship to Eya-mediated transcriptional regulation, remains poorly understood. This proposal describes a multi-faceted approach combining genetic, biochemical, and cell biological assays to address the hypothesis that proper coordination and regulation of Eya's two essential functions, one as a nuclear transcription factor and one as a component of phosphotyrosine signaling networks, is critical for retinal development. Successful completion of the proposed research plan will provide insight into the cellular and developmental contexts in which Eya's PTP function is required and should uncover novel mechanisms of cross-talk between Eya, the retinal determination gene network and other signaling modules critical for eye development. The specific aims are to elucidate the phosphotyrosine signaling pathways in which Eyes absent (Eya) participates by exploring genetic and biochemical interactions with the Abelson (Abl) and Src64 tyrosine kinases, to investigate the contribution of new binding partners identified in an in vitro expression screen with respect to Eyes absent's two functions as transcription factor and phosphatase, and to test the hypothesis that spatial partitioning of Eyes absent function between nucleus and cytoplasm contributes to proper retinal development. Because the proteins and pathways we are studying have conserved functions in mammals, the new signaling strategies revealed by our work will advance understanding of human development and disease. PUBLIC HEALTH RELEVANCE: Inappropriate regulation of gene function underlies a broad spectrum of human developmental abnormalities and diseases, including cancer. The goal of this research is to reveal the cellular mechanisms that regulate critical decisions during normal development of the eye, as an essential first step toward understanding the diseases and defects that arise when these key controls are lost and eventually toward developing effective therapeutic interventions.