Regulation of neuronal motility

描述（来自申请人的摘要）： 发育中的胚胎是建立神经元的重要步骤， 连接，神经系统功能的要求。 虽然大量的导向分子及其 神经元受体已被鉴定，但在细胞水平上知之甚少 关于受体激活如何引起细胞形状的变化， 是轴突生长所必需的神经元粘附受体的研究 其他迁移细胞表明细胞牵引力的产生 通过细胞与环境接触部位的力可能起着中心作用， 在调节细胞易位中的作用。本提案的总体目标是确定 在生长的神经元中产生牵引力的机制， 以神经元免疫球蛋白（IG）L1为模型， 受体的我们的初步数据表明，L1家族成员与 细胞骨架中产生力的成分。根据这一观察，我们 提出三个主要目标：1）描述监管和结构 L1-细胞骨架相互作用的要求和由此产生的细胞 牵引力2)为了定量L1磷酸化的动力学， 已知调节细胞骨架接头结合的细胞质尾区 蛋白质锚定B 3)在基质上使用体外神经突生长试验 用L1配体包被，以确定被鉴定为必需的结构域 上述1和2中的L1功能是L1介导的轴突生长所必需的。使用 高分辨率视频显微镜和光学梯度激光的组合 陷阱（激光镊子）拉上结合到细胞表面L1的微观珠，我们 可以监测L1功能的调节。这些方法允许 检测L1-细胞骨架相互作用，具有高的空间和时间 准确性，使我们能够直接检查L1功能是如何被调制的 配体活化、交联和磷酸化。的基因突变 人类编码L1会导致复杂的神经系统疾病，包括 痉挛性截瘫和智力低下，这表明L1在神经系统中起着重要作用。 在神经发育中的重要作用。通过表征 L1在细胞和生物物理水平上的功能，我们可以开始发展一种 详细了解指导神经元生长的机制。 此外，我们从研究粘附受体中获得的信息， 神经元可能会告知我们对各种细胞迁移的理解， 其他系统，从免疫系统功能到肿瘤细胞转移。

DESCRIPTION (From the Applicant's Abstract): The guidance of growing neurons in the developing embryo is an essential step in the establishment of neuronal connections, a requirement for nervous system function. Although a large number of guidance molecules and their neuronal receptors have been identified, little is known at the cellular level about how receptor activation gives rise to the changes in cell shape that are required for guided axon growth. The study of adhesion receptors in neurons and other migrating cells has suggested that the generation of cellular traction forces through sites of cell contact with the environment may play a central role in the regulation of cell translocation. The overall aim of this proposal is to define the mechanism underlying the generation of traction forces in the growing neuron, focusing on the neuronal immunoglobulin (Ig) protein L1 as a model adhesion receptor. Our preliminary data suggest that L1 family members interact with force-generating components of the cytoskeleton. Based on this observation, we propose three major aims: 1) To characterize the regulation and structural requirements of L1-cytoskeleton interactions and the resulting cellular traction forces. 2) To quantify the kinetics of L1 phosphorylation at a site in the cytoplasmic tail known to regulate the binding of the cytoskeleton linker protein ankyrinB. 3) To use in vitro neurite outgrowth assays on substrates coated with L1 ligands to determine how domains identified as being essential to L1 function in 1 and 2 above are required for L1-mediated axon growth. Using a combination of high-resolution video microscopy and an optical gradient laser trap (laser tweezers) to pull on microscopic beads bound to cell-surface L1, we can monitor the regulation of L1 function. These approaches permit the detection of L1-cytosksleton interactions with high spatial and temporal accuracy, allowing us to examine directly how L1 function is modulated by ligand activation, crosslinking and phosphorylation. Mutations in the gene encoding L1 in humans leads to a complex of neurological disorders including spastic paraplegia and mental retardation, suggesting that L1 plays an important role in neural development. By characterizing L1 function at a cellular and biophysical level, we can begin to develop a detailed understanding of the mechanisms underlying directed neuronal growth. Moreover, the information that we gain from studying adhesion receptors in neurons is likely to inform our understanding of cell migration in a variety of other systems, ranging from immune system function to tumor cell metastasis.