Investigating the functional consequences of SynGAP1 SUMOylation at mammalian synapses

智力残疾影响着全世界数以百万计的人，是一个主要的健康和经济负担。ID是由改变不同脑蛋白功能的突变引起的。其中一种蛋白质是SynGAP1。在人类中，SYNGAP1基因的截断或错义变异与ID、癫痫和自闭症谱系障碍有关。SynGAP1对突触的形成和形态至关重要，是兴奋性神经传递的负调节因子。此外，SUMOylation的翻译后修饰过程正在成为突触发育和功能的关键调节因子。我有关键的试验数据显示SynGAP1是突触的SUMO靶点。这就提出了SUMOylation调节SynGAP1活性和功能，在突触形成、形态和功能中发挥关键作用的假说。为了揭示SynGAP1 SUMOylation对其分子功能的生理影响，我将确定哪些信号级联介导其SUMOylation，以及这如何影响其酶活性、分子相互作用和亚细胞定位。为了确定对突触形态和神经传递（由SynGAP1控制，通常在ID中改变）的影响，我将利用活细胞成像和电生理方法，对表达不能与SUMO结合的SynGAP形式（SynGAP1- k149r）的SynGAP1- /-神经元进行复杂的生化分析。因此，拟议的项目将利用最先进的技术来直接解决调节神经元中SynGAP1功能及其控制的突触事件的分子机制。此外，它将揭示关于相扑在ID发展中的作用的基本见解。这是一个至关重要的新问题，因为对正确的大脑发育和功能至关重要的几种蛋白质是SUMO的目标。因此，该项目也将促进未来通过靶向SUMO来恢复被破坏的突触功能的策略。智力残疾影响着全世界数以百万计的人，是一个主要的健康和经济负担。ID是由改变不同脑蛋白功能的突变引起的。其中一种蛋白质是SynGAP1。在人类中，SYNGAP1基因的截断或错义变异与ID、癫痫和自闭症谱系障碍有关。SynGAP1对突触的形成和形态至关重要，是兴奋性神经传递的负调节因子。此外，SUMOylation的翻译后修饰过程正在成为突触发育和功能的关键调节因子。我有关键的试验数据显示SynGAP1是突触的SUMO靶点。这就提出了SUMOylation调节SynGAP1活性和功能，在突触形成、形态和功能中发挥关键作用的假说。为了揭示SynGAP1 SUMOylation对其分子功能的生理影响，我将确定哪些信号级联介导其SUMOylation，以及这如何影响其酶活性、分子相互作用和亚细胞定位。为了确定对突触形态和神经传递（由SynGAP1控制，通常在ID中改变）的影响，我将利用活细胞成像和电生理方法，对表达不能与SUMO结合的SynGAP形式（SynGAP1- k149r）的SynGAP1- /-神经元进行复杂的生化分析。因此，拟议的项目将利用最先进的技术来直接解决调节神经元中SynGAP1功能及其控制的突触事件的分子机制。此外，它将揭示关于相扑在ID发展中的作用的基本见解。这是一个至关重要的新问题，因为对正确的大脑发育和功能至关重要的几种蛋白质是SUMO的目标。因此，该项目也将有助于未来通过靶向SUMO来恢复被破坏的突触功能的策略。

ntellectual disability (ID) affects millions of individuals worldwide and represents a major health and economic burden. ID results from mutations altering the function of different brain proteins. One of these proteins is SynGAP1. In humans, truncated or missense variants in the SYNGAP1 gene are associated with ID, epilepsy and autism spectrum disorders. SynGAP1 is critical for synapse formation and morphology and is a negative regulator of excitatory neurotransmission. Furthermore, the post-translational modification process of SUMOylation is emerging as a key regulator of synapse development and function. I have key pilot data showing that SynGAP1 is a SUMO target at synapses. This raises the hypothesis that SUMOylation regulates SynGAP1 activity and function, to play a pivotal role in synapse formation, morphology and function. To unveil the physiological consequences of SynGAP1 SUMOylation on its molecular function, I will determine which signaling cascades mediate its SUMOylation, and how this impacts its enzyme activity, molecular interactions and subcellular localisation. To determine effects on synapse morphology and neurotransmission (which are controlled by SynGAP1 and usually altered in ID), I will exploit live-cell imaging and electrophysiological approaches with sophisticated biochemical assays in Syngap1-/- neurons that express a form of SynGAP that cannot bind to SUMO (SynGAP1-K149R). Therefore, the proposed project will exploit state of the art technologies to directly address the molecular mechanisms regulating SynGAP1 function in neurons and the synaptic events it controls. Furthermore, it will reveal essential insights regarding the role of SUMO in the development of ID. This is an emerging issue of critical importance, since several proteins essential for correct brain development and function are SUMO targets. Therefore, this project will also facilitate future strategies to restore disrupted synaptic function by targeting SUMO.ntellectual disability (ID) affects millions of individuals worldwide and represents a major health and economic burden. ID results from mutations altering the function of different brain proteins. One of these proteins is SynGAP1. In humans, truncated or missense variants in the SYNGAP1 gene are associated with ID, epilepsy and autism spectrum disorders. SynGAP1 is critical for synapse formation and morphology and is a negative regulator of excitatory neurotransmission. Furthermore, the post-translational modification process of SUMOylation is emerging as a key regulator of synapse development and function. I have key pilot data showing that SynGAP1 is a SUMO target at synapses. This raises the hypothesis that SUMOylation regulates SynGAP1 activity and function, to play a pivotal role in synapse formation, morphology and function. To unveil the physiological consequences of SynGAP1 SUMOylation on its molecular function, I will determine which signaling cascades mediate its SUMOylation, and how this impacts its enzyme activity, molecular interactions and subcellular localisation. To determine effects on synapse morphology and neurotransmission (which are controlled by SynGAP1 and usually altered in ID), I will exploit live-cell imaging and electrophysiological approaches with sophisticated biochemical assays in Syngap1-/- neurons that express a form of SynGAP that cannot bind to SUMO (SynGAP1-K149R). Therefore, the proposed project will exploit state of the art technologies to directly address the molecular mechanisms regulating SynGAP1 function in neurons and the synaptic events it controls. Furthermore, it will reveal essential insights regarding the role of SUMO in the development of ID. This is an emerging issue of critical importance, since several proteins essential for correct brain development and function are SUMO targets. Therefore, this project will also facilitate future strategies to restore disrupted synaptic synaptic function by targeting SUMO.