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.

小脑浦肯野神经元中肌球蛋白10的功能 浦肯野神经元（Purkinje neuron，PN）是小脑的主神经元，因为它接收小脑皮质的所有输入，是皮质的唯一输出，并且对于协调，平衡和学习精确的运动任务至关重要。我们先前表明，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（瓦格纳等人，Nat. Cell Biol.2011），并且肌球蛋白18 A将鸟嘌呤核苷酸交换因子-Pix靶向PN棘以促进棘成熟（亚历山大等人，FASEB J 2021）。在这里，我们描述了正在进行的努力，用于定义肌球蛋白10（Myo 10）在PN中的功能，这是唯一的中枢神经系统神经元中拥有非常高的水平，这丝状伪足肌球蛋白，并在没有事先神经支配的情况下进行丝状伪足-脊柱转换。在整个器官水平，六周龄Myo 10敲除（KO）小鼠（Heimsath等人，Sci. Rep. 2017）表现出小脑发育不全和畸形和/或小脑叶缺失。此外，这些小鼠小脑切片的钙结合蛋白染色揭示了PN索马排列和分子层内PN树突状乔木方向的缺陷。重要的是，来自成熟的6个月大的Myo 10 KO小鼠的小脑切片的钙结合蛋白染色揭示了PN形态的主要缺陷，包括树突状分支减少和棘密度大大降低。与这些观察结果一致，Myo 10 KO小鼠表现出小脑功能的显著缺陷（例如维持平衡）。最后，在培养的PN中表达的GFP标记的Myo 10显著地定位于形成神经突的前缘处的丝状伪足样延伸的尖端，并且这些细胞中的Myo 10的miRNA介导的敲低导致棘成熟和细胞极性的缺陷（减少的树突状分支，增加的轴突数量）。总之，这些结果表明，Myo 10是正常小脑发育，小脑功能和PN结构和功能所必需的，它们为未来旨在确定MyTH 4/FERM肌球蛋白促进这些过程的分子机制的努力铺平了道路。