NEURONAL CYTOSKELETAL REMODELLING

在过程的顶端，轴突伸长涉及到 从生长锥型组织到轴突的细胞质 竖井。这种转变涉及到高度动态的相互作用， 微管和肌动蛋白细丝中有一池高度动态的 微管穿透生长锥体内致密的肌动蛋白网络 薄层。后来，微管稳定下来，形成轴突。我们 假设微管相关蛋白(MAP)参与了这一过程 通过最初与肌动蛋白细丝结合的转变，来组织 生长锥片层，然后是那些将 最终形成竖井。我们建议在这方面对地图进行监管 肌动蛋白/微管结合是由肌动蛋白的磷酸化状态介导的。 地图。该模型基于Kosik实验室的观察结果， 恶性黑色素瘤细胞系(M2)缺乏肌动蛋白细丝凝胶 蛋白、肌动蛋白结合蛋白(ABP-280)可被MAP2c拯救 微量注射。这些细胞显示出皮质凝胶化的缺陷 通过显微注射肌动蛋白凝胶蛋白和通过 微量注射MAP2c。MAP2c还诱导形成两个截然不同的 结构--一个扁平的富含肌动蛋白和多个突起的板层 在微管中。与成熟的MAP2和tau相比，MAP2c最 强烈诱导形态变化，并在很早的时候就表达出来 神经元发育。 在这里，我们将研究信号转导通路在调节 MAP2c的组织特性。最近，Kosik实验室展示了 生长因子NT-3，能显著促进生长锥的形成 并在培养的神经元中传播，也增加了 MAP2和肌动蛋白。这一规定将通过联合注入 MAP异构体和各种RAC和RHO蛋白。对经济的影响 在这些实验中tau和MAP2与肌动蛋白的相互作用将是 通过直接免疫标记、电子显微镜和共... 原代培养神经元的免疫沉淀。 这项提议的另一个目的是获得临床医生的培训。 科学家。我们已经安排了几辆车，我可以通过这些车 额外的实验室培训，并继续花费很小的比例 我在临床相关问题上的时间。

At the tip of a process, neurite elongation involves the conversion of the cytoplasm from a growth cone type organization to that of a neurite shaft. This transition involves a highly dynamic interaction between microtubules and actin filaments in which a pool of highly dynamic microtubules penetrate the dense actin network in the growth cone lamella. Later microtubules stabilize to form the neurite shaft. We hypothesize that microtubule-associated proteins (MAPs) mediate this transition by initially associating with the actin filaments, to organize the growth cone lamellae, and then with those microtubules that will ultimately form the shaft. We suggest that regulation of MAPs in this actin/microtubule binding is mediated by the phosphorylation state of the MAP. This model is based on observations from the Kosik lab that a malignant melanoma cell line (M2) lacking a major actin filament gelation protein, actin binding protein (ABP-280) can be rescued by MAP2c microinjections. These cells show defects in cortical gelation that are visibly corrected by microinjection of actin gelation proteins and by microinjection of MAP2c. MAP2c also induces the formation of two distinct structures--a flattened lamella rich in actin and multiple processes rich in microtubules. In comparison to mature MAP2 and tau, MAP2c most strongly induces morphologic changes, and is expressed very early in neuronal development. Here we will study the role of signal transduction pathways in regulating the organizational properties of MAP2c. Recently the Kosik lab has shown the growth factor NT-3, which can markedly enhance growth cone formation and spreading in cultured neurons, also increased the association between MAP2 and actin. This regulation will be studied by the co-injection of MAP isoforms and a variety of rac and rho proteins. The effects on the interaction of tau and MAP2 with actin in these experiments will be determined by direct immunolabeling, electron microscopy, and co- immunoprecipitations in the primary cultured neurons. Another aim of this proposal is to obtain training as a clinician scientist. We have set-up several vehicles via which I may obtain additional laboratory training and continue spending a small proportion of my time with clinically related matters.