Roles of motor proteins in cerebellar Purkinje neuron biology

运动蛋白在小脑浦肯野神经元生物学中的作用

• 批准号: 10699720
• 负责人: JOHN A HAMMER
• 金额: $ 54.43万
• 依托单位: NATIONAL HEART, LUNG, AND BLOOD INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10699720
• 关键词: Axon; Biological Models; Biology; Cell Polarity; Cells; Cerebellar Cortex; Cerebellum; Congenital cerebellar hypoplasia; Defect; Development; Equilibrium; Exhibits; Filopodia; Future; Goals; Guanine Nucleotide Exchange Factors; Image; Knockout Mice; Learning; Lobe; MYO5A gene; Mediating; MicroRNAs; Molecular; Morphology; Motor; Mus; Myosin ATPase; Neurites; Neurons; Organ; Output; Process; Proteins; Role; Slice; Stains; Structure; Synaptic plasticity; Vertebral column; calbindin; density; design; in vivo; knock-down; knockout animal; motor learning; nerve supply; neuronal cell body

Myosin 10 function in cerebellar Purkinje neurons The Purkinje neuron (PN) is the master neuron of the cerebellum, as it receives all inputs into the cerebellar cortex, is the sole output from the cortex, and is essential for coordination, balance and learning precise motor tasks. We showed previously that myosin Va transports tubules of ER into PN spines to promote synaptic plasticity and motor learning (Wagner et al, Nat. Cell Biol. 2011), and that myosin 18A targets the guanine nucleotide exchange factor -Pix to PN spines to promote spine maturation (Alexander et al, FASEB J 2021). Here we describe ongoing efforts using to define the function of myosin 10 (Myo10) in PNs, which are unique among CNS neurons in possessing very high levels of this filopodial myosin, and in undergoing filopodia-to-spine conversion without prior innervation. At the whole organ level, six week-old Myo10 knockout (KO) mice (Heimsath et al, Sci. Rep. 2017) exhibit cerebellar hypoplasia and misshapen and/or missing cerebellar lobes. Additionally, Calbindin staining of cerebellar slices from these mice reveals defects in the alignment of PN soma and in the orientation of PN dendritic arbors within the molecule layer. Importantly, Calbindin staining of cerebellar slices from mature, six month-old Myo10 KO mice reveals major defects in PN morphology, including reduced dendritic arborization and greatly reduced spine density. Consistent with these observations, Myo10 KO mice exhibit significant defects in cerebellar function (e.g. maintaining balance). Finally, GFP-tagged Myo10 expressed in cultured PNs localizes dramatically at the tips of filopodia-like extensions at the leading edge of forming neurites, and the miRNA-mediated knockdown of Myo10 in these cells results in defects in spine maturation and cell polarity (reduced dendritic arborization, increased number of axons). Together, these results argue that Myo10 is required for normal cerebellar development, cerebellar function, and PN structure and function, and they pave the way for future efforts designed to identify the molecular mechanisms by which this MyTH4/FERM myosin promotes these processes.

肌球蛋白 10 在小脑浦肯野神经元中的功能 浦肯野神经元 (PN) 是小脑的主神经元，因为它接收小脑皮层的所有输入，是皮层的唯一输出，对于协调、平衡和学习精确的运动任务至关重要。我们之前表明，肌球蛋白 Va 将 ER 小管转运到 PN 棘中，以促进突触可塑性和运动学习 (Wagner 等人，Nat. Cell Biol. 2011)，并且肌球蛋白 18A 将鸟嘌呤核苷酸交换因子 -Pix 靶向 PN 棘，以促进棘成熟 (Alexander 等人，FASEB J 2021)。在这里，我们描述了用于定义 PN 中肌球蛋白 10 (Myo10) 功能的持续努力，PN 中的肌球蛋白 10 (Myo10) 在 CNS 神经元中是独一无二的，因为它拥有非常高水平的丝状伪足肌球蛋白，并且在没有事先神经支配的情况下经历丝状伪足到脊柱的转换。在整个器官水平上，六周大的 Myo10 敲除 (KO) 小鼠（Heimsath 等人，Sci. Rep. 2017）表现出小脑发育不全以及小脑叶畸形和/或缺失。此外，这些小鼠小脑切片的卡尔宾丁染色显示 PN 胞体排列和分子层内 PN 树突乔木方向的缺陷。重要的是，对成熟、六个月大的 Myo10 KO 小鼠的小脑切片进行卡尔宾丁染色，揭示了 PN 形态的主要缺陷，包括树突分枝减少和脊柱密度大大降低。与这些观察结果一致，Myo10 KO 小鼠表现出小脑功能的显着缺陷（例如维持平衡）。最后，培养的 PN 中表达的 GFP 标记的 Myo10 显着定位于形成神经突前缘的丝状伪足样延伸的尖端，并且这些细胞中 miRNA 介导的 Myo10 敲低导致脊柱成熟和细胞极性缺陷（树突分枝减少，轴突数量增加）。总之，这些结果表明，Myo10 是正常小脑发育、小脑功能以及 PN 结构和功能所必需的，并且它们为未来旨在确定 MyTH4/FERM 肌球蛋白促进这些过程的分子机制的努力铺平了道路。