NEURONAL CYTOSKELETAL REMODELLING

在一个过程的顶端，神经突延伸涉及到转化为 从生长锥型组织到神经突的细胞质 轴.这一转变涉及以下方面的高度动态互动： 微管和肌动蛋白丝，其中有一个高度动态的 微管穿过生长锥中致密的肌动蛋白网络 薄片后来微管稳定形成神经突轴。我们 假设微管相关蛋白（MAPs）介导了这一过程 通过最初与肌动蛋白丝结合， 生长锥薄片，然后用这些微管， 最终形成轴。 我们认为，在这种情况下， 肌动蛋白/微管结合是由磷酸化状态的 地图该模型基于Kosik实验室的观察， 缺乏主要肌动蛋白丝凝胶化的恶性黑素瘤细胞系（M2） 蛋白质，肌动蛋白结合蛋白（ABP-280）可以被MAP 2c拯救 显微注射这些细胞表现出皮质胶凝缺陷， 通过显微注射肌动蛋白胶凝蛋白和 微量注射MAP 2c。MAP 2c还诱导两种不同的 结构-一个扁平的薄层富含肌动蛋白和多个过程丰富 在微管中。与成熟的MAP 2和tau相比，MAP 2c最 强烈诱导形态学变化，并在 神经元发育 在这里，我们将研究信号转导通路在调节 MAP 2c的组织属性。最近，科西克实验室发现， 生长因子NT-3能显著促进生长锥形成 并在培养的神经元中扩散，也增加了 MAP 2和肌动蛋白。这项规定将通过共同注射研究， MAP同种型和多种rac和rho蛋白。的影响因素 在这些实验中，tau和MAP 2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- 在原代培养的神经元中的免疫沉淀。 该建议的另一个目的是获得临床医生的培训 科学家我们准备了几辆车， 额外的实验室培训和继续支出的一小部分 临床相关事务的时间

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.