Regulation of Actin Dynamics at Functional Subdomains within Dendritic Spines

描述（由申请人提供）：树突棘容纳大脑中兴奋性突触的突触后成分。突触具有固有的可塑性，树突棘是维持和修饰突触强度的重要场所，被认为是行为学习和发育可塑性的基础。越来越多的证据表明，脊柱内包含相互联系的功能子域。其中最值得注意的是突触后密度，定位和调节突触后神经递质受体数量的分子机器。此外，棘包含位于棘膜中的内吞区，其基本上远离突触。许多神经系统疾病的突触功能障碍与这些功能亚区发生的过程受损有关。重要的是，许多这些过程需要持续的肌动蛋白聚合。肌动蛋白细胞骨架在棘内的动态调节是必要的棘形态可塑性，以及突触的组成和功能的维护，并在突触的神经递质受体的插入，稳定和内吞。肌动蛋白聚合和稳定性的变化伴随着长时程增强和其他形式的突触可塑性的诱导。然而，很少有人知道的网站内的刺，肌动蛋白重排是专门需要的突触强度的改变被启动和维持。我建议，正在进行的和独立的调节肌动蛋白聚合发生在脊柱亚域，如突触后密度和内吞区。我提出的实验将测试这一假设，通过测量肌动蛋白聚合和运动在个人，活树突棘使用共聚焦，超分辨率和单粒子技术，并利用荧光蛋白标记的分子标记突触后密度（PSD）和内吞区。使用这些检测，我将测试一个广泛提出的机制，PSD与肌动蛋白细胞骨架通过蛋白coreplatin相互作用，并研究如何组织肌动蛋白在LTP诱导过程中改变脊柱内。这些结果将提供有关单个树突棘内细胞骨架组织和调节的基本新信息，这对于理解疾病中棘肌动蛋白失调的参与至关重要。

DESCRIPTION (provided by applicant): Dendritic spines house the postsynaptic components of excitatory synapses in the brain. Synapses are inherently plastic, and dendritic spines represent an important locus for maintenance and modification of synaptic strength which is thought to underlie behavioral learning and developmental plasticity. Increasing evidence exists that spines contain within them interlinked sets of functional subdomains. Most notable of these is the postsynaptic density, the molecular machine which positions and regulates the number of postsynaptic neurotransmitter receptors. In addition, spines contain an endocytic zone positioned in the spine membrane substantially away from the synapse. The synaptic dysfunction underlying many neurological disorders is associated with impairment of processes occurring at these functional subdomains. Importantly, a number of these processes require ongoing actin polymerization. Dynamic regulation of the actin cytoskeleton within spines is necessary for spine morphological plasticity as well as maintenance of synaptic composition and function, and underlies the insertion, stabilization, and endocytosis of neurotransmitter receptors at the synapse. Changes in actin polymerization and stability within spines accompany and are required for the induction of long term potentiation and other forms of synaptic plasticity. However, little is known about the sites within spines at which actin rearrangement is specifically required for alterations in synaptic strength to be initiated and maintained. I propose that ongoing and independent regulation of actin polymerization occurs at spine subdomains such as the postsynaptic density and endocytic zone. My proposed experiments will test this hypothesis by measuring actin polymerization and movement within individual, live dendritic spines using confocal, super-resolution, and single-particle techniques, and utilizing fluorescent protein-tagged molecules to mark the postsynaptic density (PSD) and endocytic zones. Using these assays, I will then test a widely proposed mechanism that the PSD interacts with the actin cytoskeleton via the protein cortactin, and examine how the organization of actin is altered within spines during induction of LTP. These results will provide fundamental new information about cytoskeletal organization and regulation within individual dendritic spines, critical for understanding the involvement of spine actin dysregulation in disease.