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功能 并与代谢调节和癌症的发生有关。然而，许多问题仍然存在。 关于细胞外配体对它们的调节及其下游信号机制。果蝇 果蝇为研究RPTP在体内的作用机制提供了一个有吸引力的模型系统。 Lar和PTP69D是R7所必需的IIa型RPTP家族中的两个果蝇成员 光感受器选择正确的突触靶层，并为幼虫运动神经元形成突触的 目标肌肉的大小正确。任何一种RPTP的突变都会产生一种强大的、可量化的表型。 然而，R7光感受器和幼虫运动神经元在对特定的 LAR的结构特点。该提案将调查LAR使用的新信令机制如何 直接靶向R7与其在运动神经元突触生长和PTP69D信号转导中的作用模式不同。 一些RPTPs的磷酸酶活性受二聚化的负调控。初步数据 表明R7靶选择不需要LAR的磷酸酶活性，但确实需要一个结构域 这调节了LAR的二聚化。这项提案的第一个目标将检查PTP69D是否也有两个 不同的信号机制，以及它是否可以与运动神经元中的LAR互换。它还将研究 研究强制二聚对LAR功能的影响，并建立一种体内可视化LAR二聚的方法。 此外，还将研究PTP69D与LAR的均二聚或异二聚的重要性。 RPTPs已被证明可以调节细胞与细胞和细胞与基质的黏附。在此的第二个目标中 提议，涉及这些功能的分子将接受遗传和物理相互作用的测试 用LAR来确定LAR在R7光感受器中的磷酸酶非依赖性功能是否使用了 这些机制中。此外，遗传和生化方法都将用于无偏见的筛查 潜在的新型分子，需要二聚化结构域与LAR相互作用，因此可能起作用 在R7的LAR下游。 控制运动神经元中LAR活性的配体在R7中不调节它。这样做的最终目的是 建议筛选跨膜和分泌蛋白对R7靶向的影响，以确定 靶神经元表达的LAR或PTP69D的候选配体。然后，这些候选人将接受测试 它们与两个RPTP结合并调节其功能的能力。综上所述，这个实验 提案将描述RPTP功能的非规范机制，并可能确定新的配体和 这类重要但知之甚少的受体的下游效应器。

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.