Myotubularin PI 3-Phosphatases as Regulators of Peripheral Nerve Myelination

肌管蛋白 PI 3-磷酸酶作为周围神经髓鞘形成的调节剂

• 批准号: 9252598
• 负责人: FRED L ROBINSON
• 金额: $ 33.69万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-15 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9252598
• 关键词: 1-Phosphatidylinositol 4-Kinase; Affect; Back; Binding; Biochemical; Cell Survival; Cell division; Cell membrane; Charcot-Marie-Tooth Disease; Coculture Techniques; Code; Complex; Defect; Deposition; Disease; Endocytosis; Eukaryota; Family; Generations; Genetically Engineered Mouse; Goals; Guanine Nucleotide Exchange Factors; Guanosine Triphosphate Phosphohydrolases; High Pressure Liquid Chromatography; Homeostasis; Hydrolysis; In Vitro; Inherited; Knockout Mice; Lead; Limb structure; Lipids; Location; MADD gene; Mediating; Membrane; Membrane Protein Traffic; Membrane Proteins; Metabolism; Modeling; Morphology; Mutation; Myelin; Myelin Sheath; Organelles; Output; Pathway interactions; Patients; Peripheral Nerves; Peripheral Nervous System Diseases; Pharmacology; Phosphatidylinositols; Phospholipase; Phosphoric Monoester Hydrolases; Phosphotransferases; Protein Isoforms; Proteins; Recruitment Activity; Recycling; Regulation; Research; Role; Scaffolding Protein; Schwann Cells; Sensory; Signal Transduction; Signaling Molecule; Tertiary Protein Structure; Testing; Therapeutic; Transmembrane Transport; Type 4b Charcot Marie Tooth Disease; United States; axonal degeneration; base; cell growth; cell motility; disease-causing mutation; effective therapy; human disease; knockout gene; loss of function mutation; member; muscle degeneration; myelination; myotubularin; nervous system disorder; novel; phosphatidylinositol 3,5-diphosphate; phosphatidylinositol 3-phosphate; phosphoinositide 3-phosphate; public health relevance; rab GTP-Binding Proteins; scaffold; trafficking

DESCRIPTION (provided by applicant): The overall goal of the proposed research is to determine the mechanisms by which disturbances in phosphoinositide (PI) regulation lead to abnormal membrane trafficking and cellular signaling in myelinating Schwann cells. We are studying this question in the context of a specific form of demyelinating Charcot-Marie-Tooth peripheral neuropathy (type 4B~ CMT4B), which is characterized by abnormal myelination and severe axonal degeneration. CMT is one of the most common inherited neurological disorders, affecting about 1 in 2500 worldwide. This condition leads to progressive degeneration of the muscles of the extremities and loss of sensory function. CMT4B is caused by loss of function mutations in either myotubularin-related protein 2 (MTMR2) or MTMR13, which belong to a large family of phosphatases that act as key regulators of PI signaling. MTMR2 is a PI 3-phosphatase that specifically dephosphorylates phosphatidylinositol 3-phosphate (PI3P) and phosphatidylinositol 3,5-bisphosphate (PI[3,5]P2). PI3P and PI(3,5)P2 regulate membrane traffic within the endosomal/lysosomal pathway. Therefore, it is theorized that CMT4B arises from defects in membrane transport in Schwann cells. MTMR13 is a catalytically inactive "pseudophosphatase" that associates directly with MTMR2. MTMR13 appears to function as a scaffold protein that regulates MTMR2. The first aim of this proposal is to define the phosphoinositide kinase-phosphatase network that controls PI3P and PI(3,5)P2 regulation in Schwann cells. The impact of the loss of PI kinases and phosphatases on phosphoinositide levels will be evaluated using HPLC-based phosphoinositide profiling. In parallel, the impact of the loss of PI kinases and phosphatases on myelination will be assessed using in vitro myelinating cultures and morphological examination of peripheral nerves of knockout mice. The second aim of the study is to define the function of the Mtmr13 pseudophosphatase in Schwann cell membrane traffic by determining how Mtmr13's activation of Rab GTPases regulates myelination, and by assessing the role of the Mtmr2-Mtmr13 complex in the regulation of endocytosis. These goals will be accomplished using biochemical studies to identify interacting Rab GTPases, as well as in vitro myelinating co-cultures to assess the relevance of specific interactions to myelination. The final aim of this proposal is to determine which domains of Mtmr13 control its specific functions. Collectively, these studies will allow us to define the specific, critical roles of phosphoinositides in myelinating Schwann cells. These studies also may well form the basis of a rational approach to the treatment of CMT4B by pharmacological targeting of the PI3P-PI(3,5)P2 pathway.

描述（由申请人提供）：拟议研究的总体目标是确定磷酸肌醇（PI）调节紊乱导致髓鞘化雪旺细胞中异常膜运输和细胞信号传导的机制。 我们正在研究一种特殊形式的Charcot-Marie-Tooth脱髓鞘周围神经病（4 B ~ CMT 4 B型）的背景下，这是一个问题，其特征是异常髓鞘形成和严重的轴突变性。 CMT是最常见的遗传性神经系统疾病之一，在全球范围内影响约1/2500。 这种情况导致四肢肌肉的进行性退化和感觉功能的丧失。 CMT 4 B是由肌微管蛋白相关蛋白2（MTMR 2）或MTMR 13中的功能缺失突变引起的，所述MTMR 2或MTMR 13属于作为PI信号传导的关键调节剂的磷酸酶大家族。 MTMR 2是PI 3-磷酸酶，其特异性地使磷脂酰肌醇3-磷酸（PI 3 P）和磷脂酰肌醇3，5-二磷酸（PI[3，5]P2）去磷酸化。 PI 3 P和PI（3，5）P2调节内体/溶酶体途径内的膜运输。 因此，理论上CMT 4 B是由许旺细胞中的膜转运缺陷引起的。 MTMR 13是一种无催化活性的“假磷酸酶”，与MTMR 2直接相关。 MTMR 13似乎作为调节MTMR 2的支架蛋白发挥作用。 该建议的第一个目的是定义在雪旺细胞中控制PI 3 P和PI（3，5）P2调节的磷酸肌醇激酶-磷酸酶网络。 将使用基于HPLC的磷酸肌醇谱分析评价PI激酶和磷酸酶缺失对磷酸肌醇水平的影响。 同时，将使用体外髓鞘形成培养物和敲除小鼠外周神经的形态学检查来评估PI激酶和磷酸酶的损失对髓鞘形成的影响。 该研究的第二个目的是通过确定Mtmr 13对Rab GTP酶的激活如何调节髓鞘形成，以及通过评估Mtmr 2-Mtmr 13复合物在胞吞调节中的作用，来确定Mtmr 13假磷酸酶在许旺细胞膜交通中的功能。 这些目标将通过生物化学研究来实现，以确定相互作用的Rab GTP酶，以及体外髓鞘形成共培养物，以评估特定相互作用与髓鞘形成的相关性。 本提案的最终目的是确定Mtmr 13的哪些域控制其特定功能。 总的来说，这些研究将使我们能够确定特定的，关键作用的磷酸肌醇髓鞘雪旺细胞。 这些研究也可以很好地形成通过药理学靶向PI 3 P-PI（3，5）P2途径治疗CMT 4 B的合理方法的基础。