Elucidating the dynamic role of PTPsigma in synaptic nano-organization and NMDA receptor function

阐明 PTPsigma 在突触纳米组织和 NMDA 受体功能中的动态作用

• 批准号: 10606077
• 负责人: Emily M. DeMarco
• 金额: $ 4.22万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-01 至 2026-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10606077
• 关键词: AMPA Receptors; Action Potentials; Acute; Affect; Anxiety Disorders; Binding; Brain; Cell Adhesion Molecules; Cells; Chronic; Cognition; Communication; Compensation; Confocal Microscopy; DLG4 gene; DNA; Development; Electrophysiology (science); Engineering; Evoked Potentials; Excitatory Synapse; Experimental Designs; Extracellular Domain; Family; Fellowship; Genetic; Genetic Complementation Test; Glutamate Receptor; Goals; Hippocampus; Image; Knock-out; Lead; Learning; Measures; Mediating; Memory; Mental Depression; Methodology; Microscopy; Modeling; Molecular Biology; N-Methyl-D-Aspartate Receptors; N-Methylaspartate; N-terminal; Nanostructures; Neurons; Neurosciences; Optics; Pathology; Peptide Hydrolases; Physiology; Play; Positioning Attribute; Probability; Process; Protein Family; Protein Tyrosine Phosphatase; Proteins; Receptor Activation; Regulation; Research Personnel; Resistance; Resolution; Role; Scaffolding Protein; Site; Structure; Synapses; Synaptic Cleft; Synaptic Transmission; Techniques; Testing; Training; Universities; Vesicle; Work; autism spectrum disorder; career; density; experimental study; extracellular; improved; insight; knock-down; live cell imaging; member; mutant; nano; nanocolumn; nanoscale; neuropsychiatric disorder; neurotransmitter release; novel; patch clamp; postsynaptic; presynaptic; protein complex; receptor; receptor function; response; scaffold; single molecule; superresolution microscopy; synaptogenesis; transmission process; vesicular release; AMPA Receptors; Action Potentials; Acute; Affect; Anxiety Disorders; Binding; Brain; Cell Adhesion Molecules; Cells; Chronic; Cognition; Communication; Compensation; Confocal Microscopy; DLG4 gene; DNA; Development; Electrophysiology (science); Engineering; Evoked Potentials; Excitatory Synapse; Experimental Designs; Extracellular Domain; Family; Fellowship; Genetic; Genetic Complementation Test; Glutamate Receptor; Goals; Hippocampus; Image; Knock-out; Lead; Learning; Measures; Mediating; Memory; Mental Depression; Methodology; Microscopy; Modeling; Molecular Biology; N-Methyl-D-Aspartate Receptors; N-Methylaspartate; N-terminal; Nanostructures; Neurons; Neurosciences; Optics; Pathology; Peptide Hydrolases; Physiology; Play; Positioning Attribute; Probability; Process; Protein Family; Protein Tyrosine Phosphatase; Proteins; Receptor Activation; Regulation; Research Personnel; Resistance; Resolution; Role; Scaffolding Protein; Site; Structure; Synapses; Synaptic Cleft; Synaptic Transmission; Techniques; Testing; Training; Universities; Vesicle; Work; autism spectrum disorder; career; density; experimental study; extracellular; improved; insight; knock-down; live cell imaging; member; mutant; nano; nanocolumn; nanoscale; neuropsychiatric disorder; neurotransmitter release; novel; patch clamp; postsynaptic; presynaptic; protein complex; receptor; receptor function; response; scaffold; single molecule; superresolution microscopy; synaptogenesis; transmission process; vesicular release

Fine tuning the efficiency of synaptic transmission is essential for learning and memory, while its disruption is associated with diverse pathologies including autism spectrum disorder, depression, and anxiety disorders. Thus, identifying mechanisms that regulate synaptic strength is a central goal in neuroscience. Since such plasticity is frequently triggered by activation of NMDA-type glutamate receptors, understanding the regulation of NMDA receptors is particularly critical. Recent studies indicate synaptic strength and NMDA receptor activation can be directly affected by the nanometer-scale organization of proteins within the synapse. Many synaptic proteins, including vesicle release machinery and postsynaptic scaffolds and receptors, display a heterogeneous organization with local regions of high protein density, known as nanodomains (NDs). These NDs can be aligned across the synapse to form a “nanocolumn”, which our lab has demonstrated is the site of action potential-evoked vesicle fusion and maximal receptor activation. New work from our lab has established a novel role for the postsynaptic cell-adhesion molecule (CAM) LRRTM2 in positioning AMPA receptors within the nanocolumn. While CAMs have well-established roles in synapse formation and development, these recent findings highlight the possibility that CAMs may coordinate synaptic nanostructure and function in the mature synapse. However, the mechanism by which presynaptic organization and vesicle fusion sites are communicated to proteins within the postsynaptic density to enable alignment to occur remains unknown. In this proposal I will investigate whether the presynaptic CAM PTPσ coordinates nanocolumn alignment. PTPσ is important for synapse formation, is present in the mature synapse, and forms indirect interactions with both pre- and postsynaptic machinery located within the nanocolumn. Loss of PTPσ impacts both pre- and postsynaptic physiology, most notably NMDA receptor-mediated responses. Previous attempts to study PTPσ have relied on chronic manipulations, such as knockouts and knockdowns. However, interpretations are complicated by its initial role in synapse formation during development. I propose to elucidate the ongoing functions of PTPσ by acutely disrupting its cleft interactions via cleavage by an exogenous protease. This highly specific and acute approach will allow me to manipulate PTPσ’s cleft interactions to isolate their functions, without compromising its earlier role in synapse formation. Throughout this project, I will use super-resolution microscopy, electrophysiology, molecular biology, and live-cell imaging to test the role of PTPσ cleft interactions in maintaining nanocolumn alignment and regulating NMDA receptor-mediated transmission. This work will provide novel insight regarding the roles of a critical family of presynaptic CAMs following development and will test a new candidate mechanism for the coordination of synaptic nanostructure and NMDA receptor function. The training obtained under this fellowship will provide deep and diverse training in methodologies and professional development that will prepare me excel in my career as an academic researcher at a biomedical university.

微调突触传播的效率对于学习和记忆至关重要，而其中断是 与潜水病理学有关，包括自闭症谱系障碍，抑郁症和焦虑症。 这是确定调节突触强度的机制是神经科学的核心目标。自从这样 塑性通常是由NMDA型谷氨酸受体激活引发的，了解调节 NMDA受体特别关键。最近的研究表明合成强度和NMDA受体 激活可以直接受到突触中蛋白质的纳米尺度组织的影响。许多 突触蛋白，包括囊泡释放机械和突触后脚手架和接收器，显示一个 具有高蛋白质密度的局部区域，称为纳米域（NDS）。这些 ND可以在突触中对齐以形成“纳米柱”，我们的实验室证明了这是 动作潜在诱发的蔬菜融合和最大受体激活。我们实验室的新工作已经建立 突触后细胞粘附分子（CAM）LRRTM2在定位AMPA接收器中的新作用 纳米柱。尽管凸轮在突触形成和发展中具有完善的角色，但这些最近 发现突出了凸轮可以协调突触纳米结构并在成熟中功能的可能性 突触。但是，传达突触前组织和囊泡融合点的机制 对于突触后密度内的蛋白质以使对齐能够进行对齐仍然未知。在这个建议中，我将 研究突触前CAMPTPσ是否坐标纳米颜色对齐。 PTPσ对 突触形成，存在于成熟的突触中，并形成与前和前的间接相互作用 突触后机械位于纳米船柱内。 PTPσ的损失会影响突触前和突触后 生理学，最著名的是NMDA受体介导的反应。以前研究PTPσ的尝试已在 长期操纵，例如淘汰赛和敲除。但是，解释很复杂 在开发过程中的突触形成中的最初作用。我建议通过 急性通过外源蛋白酶的切割来破坏其裂缝相互作用。这个高度特异性和敏锐 方法将使我能够操纵PTPσ的裂口相互作用以隔离其功能，而不会妥协 它在突触形成中的早期作用。通过这个项目，我将使用超分辨率显微镜， 电生理学，分子生物学和活细胞成像，以测试PTPσ裂口相互作用在 维持纳米颜色比对和调节NMDA受体介导的传播。这项工作将提供 关于开发后突触前凸轮的关键家族的作用的新颖见解，并将测试 突触纳米结构和NMDA受体功能协调的新候选机制。 根据该奖学金获得的培训将为方法和专业人士提供深厚的潜水员培训 在我在生物医学大学担任学术研究员职业生涯中，我的发展使我的发展使我脱颖而出。