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

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.

微调突触传递的效率对于学习和记忆至关重要，而突触传递的破坏则对学习和记忆至关重要。 与多种病症相关，包括自闭症谱系障碍、抑郁症和焦虑症。 因此，识别调节突触强度的机制是神经科学的一个中心目标。 可塑性经常由 NMDA 型谷氨酸受体的激活触发，了解其调节机制 最近的研究表明突触强度和 NMDA 受体的关系尤其重要。 突触内蛋白质的纳米级组织可以直接影响激活。 突触蛋白，包括囊泡释放机制和突触后支架和受体，显示出 具有高蛋白质密度局部区域的异质组织，称为纳米域（ND）。 ND 可以在突触上排列形成“纳米柱”，我们的实验室已经证明这是 我们实验室的新工作已经建立了动作电位诱发的囊泡融合和最大受体激活。 突触后细胞粘附分子 (CAM) LRRTM2 在定位 AMPA 受体中的新作用 虽然 CAM 在突触形成和发育中具有明确的作用，但最近的这些研究 研究结果强调了 CAM 可能在成熟细胞中协调突触纳米结构和功能的可能性。 然而，突触前组织和囊泡融合位点的沟通机制 突触后密度内的蛋白质如何使排列发生仍然未知。 研究突触前 CAM PTPσ 协调纳米柱对齐是否重要。 突触形成，存在于成熟突触中，并与前突触和突触形成间接相互作用 位于纳米柱内的突触后机制 PTPσ 的损失会影响突触前和突触后。 生理学，尤其是 NMDA 受体介导的反应，之前研究 PTPσ 的尝试依赖于 NMDA 受体介导的反应。 然而，其解释却很复杂。 我建议通过以下方式阐明 PTPσ 在突触形成中的初始作用。 通过外源蛋白酶的切割，急剧破坏其裂口相互作用，这种高度特异性和急性的。 该方法将允许我操纵 PTPσ 的裂隙相互作用来隔离它们的功能，而不会影响 它在突触形成中的早期作用在整个项目中，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来测试 PTPσ 裂隙相互作用在 这项工作将提供维持纳米柱排列和调节 NMDA 受体介导的传输。 关于突触前 CAM 关键家族在发育后的作用的新颖见解，并将测试 协调突触纳米结构和 NMDA 受体功能的新候选机制。 在该研究金下获得的培训将提供方法论和专业方面的深入和多样化的培训 我的发展将使我在生物医学大学担任学术研究员的职业生涯中取得优异的成绩。