Mechanisms of receptor protein tyrosine phosphatase signaling in Drosophila devel

受体蛋白酪氨酸磷酸酶（RPTPs）在神经系统中具有重要的功能 发育并参与代谢调节和致癌作用。然而，许多问题仍然存在 关于它们通过细胞外配体的调节及其下游信号传导机制。果蝇 果蝇提供了一个有吸引力的模型系统，在其中解决的RPTP功能在体内的机制。 LAR和PTP 69 D是RPTP的IIa型家族的两个果蝇成员，是R7所必需的 光感受器选择正确的突触目标层，并为幼虫运动神经元形成突触的 正确的大小在他们的目标肌肉。任一RPTP的突变产生强的和可量化的表型。 然而，R7光感受器和幼虫运动神经元在它们对特定的细胞因子的需求方面存在显著差异。 的结构特征。这项提案将研究如何使用新的信号机制LAR， 直接R7靶向不同于其在运动神经元突触生长中的作用模式和PTP 69 D信号传导。 一些RPTP的磷酸酶活性受二聚化负调控。初步数据 显示R7靶选择不需要LAR的磷酸酶活性，但需要结构域 介导LAR二聚化。本提案的第一个目的是检查PTP 69 D是否也有两个 不同的信号传导机制，以及它是否可以与运动神经元中的LAR互换。还将研究 强迫二聚化对LAR功能的影响，并开发一种体内可视化LAR二聚化的方法。 此外，将研究PTP 69 D同二聚化或异二聚化与LAR的重要性。 RPTP已显示调节细胞-细胞和细胞-基质粘附。在第二个目标中， 根据一项提议，与这些功能有关的分子将被检测是否存在遗传和物理相互作用 以确定LAR在R7光感受器中的磷酸酶非依赖性功能是否使用一个 这些机制。此外，遗传和生物化学方法将用于无偏筛选， 潜在的新分子，需要二聚化结构域与LAR相互作用，因此可能发挥作用， 在R7的LAR下游。 在运动神经元中控制LAR活性的配体在R7中不调节它。最终目的是 一个建议是筛选跨膜和分泌蛋白对R7靶向的影响，以鉴定 由靶神经元表达的LAR或PTP 69 D的候选配体。然后这些候选人将接受测试 因为它们能够结合并调节两种RPTP的功能。综合来看， 该提案将描述RPTP功能的非经典机制，并可能识别新的配体， 这类重要但知之甚少的受体的下游效应子。

Receptor protein tyrosine phosphatases (RPTPs) have important functions in nervous system development and are implicated in metabolic regulation and carcinogenesis. However, many questions remain regarding their regulation by extracellular ligands and their downstream signaling mechanisms. The fruit fly Drosophila offers an attractive model system in which to address the mechanisms of RPTP function in vivo. LAR and PTP69D, the two Drosophila members of the type IIa family of RPTPs, are required for R7 photoreceptors to select the correct synaptic target layer, and for larval motor neurons to form synapses of the correct size on their target muscles. Mutation of either RPTP produces a strong and quantifiable phenotype. However, R7 photoreceptors and larval motor neurons differ significantly in their requirements for specific structural features of LAR. This proposal will investigate how the novel signaling mechanism used by LAR to direct R7 targeting differs from its mode of action in motor neuron synapse growth and from PTP69D signaling. The phosphatase activity of some RPTPs is negatively regulated by dimerization. Preliminary data shows that R7 target selection does not require the phosphatase activity of LAR, but does require a domain that mediates LAR dimerization. The first aim of this proposal will examine whether PTP69D also has two distinct signaling mechanisms, and whether it is interchangeable with LAR in motor neurons. It will also study the effect of forced dimerization on LAR function and develop a method to visualize LAR dimerization in vivo. In addition, the importance of PTP69D homodimerization or heterodimerization with LAR will be investigated. RPTPs have been shown to regulate both cell-cell and cell-matrix adhesion. In the second aim of this proposal, molecules implicated in each of these functions will be tested for genetic and physical interactions with LAR to determine whether the phosphatase-independent function of LAR in R7 photoreceptors uses one of these mechanisms. In addition, both genetic and biochemical methods will be used in unbiased screens for potentially novel molecules that require the dimerization domain to interact with LAR and might therefore act downstream of LAR in R7. The ligands that control LAR activity in motor neurons do not regulate it in R7. The final aim of this proposal is to screen transmembrane and secreted proteins for an effect on R7 targeting, in order to identify candidate ligands for LAR or PTP69D expressed by the target neurons. These candidates will then be tested for their ability to bind to and regulate the function of both RPTPs. Taken together, the experiments in this proposal will characterize a non-canonical mechanism of RPTP function, and may identify new ligands and downstream effectors for this important but poorly understood class of receptors.