NEUROTROPHIN RECEPTOR INTERACTIONS

75kD(P75)的功能相互作用的特定模型 酪氨酸激酶(Trk)神经营养素受体将进行测试。模型 提出p75的功能形式是二聚体，即二聚化 受磷酸化调节，神经营养因子作用于p75和trk 独立于受体，神经营养因子激活p75的过程如下 关于协同构象转变的Monod模型，以及 P75的活性构象优先与trk二聚体结合，便于 Trk受体的二聚化和激活。另外，p75有可能 通过调节逆行轴突促进逆行轴突信号传递 活性神经营养因子/trk复合体的运输将被研究。装订 P75受体在不同细胞系中表达的研究和生化研究 将进行非神经元和神经元细胞系的检测 受体二聚化、磷酸化和正相关关系 协同神经营养因子结合及其相互关系 P75增强trk受体激活的能力将是 检查过了。P75的体外诱变，随后在非 神经元和神经元细胞系，将被用来定义重要的 P75函数中涉及的结构元素--特别关注 细胞质结构域中可能的调节磷酸化位点，以及 可能需要的跨膜结构域中的半胱氨基残基 用于产生功能性受体二聚体。P75基因在细胞周期中的作用 含神经营养因子的内吞囊泡的细胞内转运 Trk受体将在非神经细胞和 培养中的神经元。电子的免疫组织化学技术 将进行微观层面的研究，以表征 P75和trk受体在胞内膜池中的分布。共焦 荧光显微镜，使用对pH敏感的染料，将进行 雏鸡轴突内内体酸化时间的测定 培养中的胚胎交感神经和感觉轴突。这些信息， 结合检测神经营养因子的pH依赖性的结果 与受体的结合，将揭示功能是否活跃 神经营养因子/受体复合体可以完整地从神经中运输 神经元胞体的末端，以传递逆行营养 信号。这些研究将描述基本的过程 神经营养因子塑造发育的机制 大脑和周围神经系统，以及通过什么再生 鼓励受伤的神经系统。

A specific model for the functional interaction of 75 kD (p75) and tyrosine kinase (trk) neurotrophin receptors will be tested. The model proposes that the functional form of p75 is dimeric, that dimerization is regulated by phosphorylation, that neurotrophins act upon p75 and trk receptors independently, that activation of p75 by neurotrophins follows the Monod model for concerted conformational transitions, and that the active conformation of p75 preferentially binds trk dimers, facilitating trk receptor dimerization and activation. Also, the possibility that p75 facilitates retrograde axonal signaling by regulating retrograde axonal transport of active neurotrophin/trk complexes will be examined. Binding studies and biochemical studies of p75 receptors expressed in various non-neuronal and neuronal cell lines will be performed to examine the relationship of receptor dimerization, phosphorylation, and positively cooperative neurotrophin binding, and relationship of these properties to the ability of p75 to enhance trk receptor activation will be examined. In vitro mutagenesis of p75, followed by expression in non- neuronal and neuronal cell lines, will be employed to define important structural elements involved in p75 function - particularly focusing on putative regulatory phosphorylation sites in the cytoplasmic domain, and a cysteinyl residue within the transmembrane domain which may be required for generation of functional receptor dimers. The effects of p75 on intracellular trafficking of endocytic vesicles bearing neurotrophins and trk receptors will be examined, in non-neuronal cells, and in axons of neurons in culture. Immunohistochemical techniques at the electron microscopic level will be performed to characterize the distribution of p75 and trk receptors among intracellular membrane pools. Confocal fluorescence microscopy, with pH-sensitive dyes, will be performed to determine the timing of acidification of endosomes within axons of chick embryo sympathetic and sensory axons in culture. This information, coupled with the results of examining the pH-dependence of neurotrophin binding to receptors, will reveal whether functionally active neurotrophin/receptor complexes may be transported intact from the nerve terminus to the neuronal cell body in order to convey retrograde trophic signals. These studies will characterize processes which are fundamental to the mechanisms by which neurotrophic factors shape the developing brain and peripheral nervous system, and by which regeneration of the injured nervous system is encouraged.