Dissecting Ret receptor signaling in space-filling dendrite patterning in Drosophila

解析果蝇空间填充树突图案中的 Ret 受体信号传导

• 批准号: 397556468
• 负责人: Professor Dr. Peter Soba
• 金额: --
• 依托单位: Abteilung Molekulare Hirnphysiologie und Verhaltensforschung
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2018
• 资助国家: 德国
• 起止时间: 2017-12-31 至 2020-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/397556468?language=en
• 关键词: Dissecting; Ret; receptor; signaling; space

The conserved receptor tyrosine kinase Ret (Rearranged during transfection) plays a major role in nervous system development, maintenance and disease. Although Ret has been heavily studied, its function and signaling in nervous system development are only incompletely understood in vivo. We have recently shown that Ret plays a major role in dendrite growth, dynamics and adhesion in a subset of Drosophila sensory neurons. The Drosophila larval peripheral nervous system has been a powerful model to study molecular mechanisms of dendrite development in vivo, featuring highly stereotyped neuronal morphologies and accessibility to confocal live imaging microscopy. Sensory dendrites of these neurons grow in a 2-dimensional environment between an epidermal cell layer and the extracellular matrix (ECM). The most complex class IV (C4da) neurons cover the entire larval body wall with their dendrites in a complete yet non-redundant manner, and their development critically relies on Ret function. We have shown that Ret is required for dendrite-ECM adhesion in C4da neurons by forming a functional complex with integrins and downstream signaling by the small GTPase Rac1. However, Ret dependent dendrite growth and dynamics are relying on other so far unidentified molecular cues. By candidate screening and genome wide analysis of Ret deficient C4da neurons we have identified novel ligand and downstream adaptor proteins linking Ret function to TGFß signaling. We therefore hypothesize that Drosophila Ret is a TGFß-like receptor mediating C4da neuron dendrite growth in response to a novel extracellular ligand. We will investigate these novel interactors by genetic loss and gain of function analyses, cellular and biochemical assays. First, we aim to analyze the function of the novel putative Ret ligand in C4da neuron dendrite development by generating CRISPR/Cas9 mediated knockout and knock-in lines. We will extensively investigate its developmental expression and function in C4da neuron dendrite growth. Second, we will address the link between Ret and TGFß and receptor tyrosine kinase signaling in vivo and in vitro to investigate ligand-receptor interaction and the relevant signaling pathway. Lastly, we will analyze the role of the identified intracellular adaptor proteins in Ret dependent C4da neuron dendrite development to get molecular and cellular insight into the downstream signaling processes. Taken together, our studies will address a novel Ret dependent signaling mechanism required for space-filling dendrite growth.

保守的受体酪氨酸激酶Ret（转染期间重排）在神经系统发育、维持和疾病中发挥重要作用。虽然Ret已经被大量研究，但其在神经系统发育中的功能和信号传导仅在体内不完全了解。我们最近发现，Ret在果蝇感觉神经元的树突生长，动力学和粘附中起着重要作用。果蝇幼虫的周围神经系统已成为一个强大的模型来研究树突发育的分子机制在体内，具有高度定型的神经元形态和可访问的共聚焦实时成像显微镜。这些神经元的感觉树突生长在表皮细胞层和细胞外基质（ECM）之间的二维环境中。最复杂的IV类（C4da）神经元以完整而非冗余的方式用树突覆盖整个幼虫体壁，它们的发育严重依赖于Ret功能。我们已经表明，Ret是C4da神经元树突-ECM粘附所必需的，通过与整合素形成功能复合物和小GTdR Rac 1的下游信号传导。然而，Ret依赖的树突生长和动力学依赖于其他迄今未识别的分子线索。通过对Ret缺陷型C4da神经元的候选筛选和全基因组分析，我们已经鉴定了将Ret功能与TGF β 1信号传导连接的新型配体和下游衔接蛋白。因此，我们假设果蝇Ret是一个TGF β 1样受体介导的C4da神经元树突生长的一种新的细胞外配体。我们将研究这些新的相互作用基因的功能分析，细胞和生化检测的遗传损失和增益。首先，我们的目标是通过产生CRISPR/Cas9介导的敲除和敲入系来分析新型推定的Ret配体在C4da神经元树突发育中的功能。我们将广泛研究其在C4da神经元树突生长中的发育表达和功能。其次，我们将在体内和体外研究Ret和TGF β 1与受体酪氨酸激酶信号之间的联系，以研究配体-受体相互作用和相关的信号通路。最后，我们将分析识别的细胞内接头蛋白在Ret依赖的C4da神经元树突发育中的作用，以获得对下游信号传导过程的分子和细胞见解。总之，我们的研究将解决一个新的Ret依赖的信号机制所需的空间填充树突生长。

项目成果

Maintenance of cell type-specific connectivity and circuit function requires Tao kinase

• DOI: 10.1038/s41467-019-11408-1
• 发表时间: 2019-07
• 期刊: Nature Communications
• 影响因子: 16.6
• 作者: Federico Tenedini;Maria Sáez González;Chun Hu;Lisa H. Pedersen;Mabel Matamala Petruzzi;Bettina Spitzweck;Denan Wang;Melanie Richter;Meike Petersen;E. Szpotowicz;M. Schweizer;S. Sigrist;Froylan Calderón de Anda;P. Soba