Post-Translational Control of Neuroplasticity Effector Arc/Arg3.1

神经可塑性效应器 Arc/Arg3.1 的翻译后控制

• 批准号: RGPIN-2018-04616
• 负责人: Lalonde, Jasmin
• 金额: $ 2.26万
• 依托单位: University of Guelph
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2018
• 资助国家: 加拿大
• 起止时间: 2018-01-01 至 2019-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=656432
• 关键词: Post; Translational; Control; Neuroplasticity; Effector

Neurons have the remarkable capacity to reorganize their structure, function, and connections in response to stimuli. Among the many signaling molecules and protein effectors found supporting these neuroplastic changes, the Activity-regulated cytoskeleton-associated protein (Arc) can be considered one of the central, as well as most versatile, players. Past research highlight the multiple roles of Arc in neurons; however, what allows this protein to assume one cellular function over another, or interact with the appropriate molecular effectors in response to a specific signal, remains poorly understood. The overarching goal of this Discovery Grant proposal is to test the hypothesis that lysine acetylation of Arc protein at precise sites produces a strong modulatory effect on its subcellular distribution, functions, and/or affinity with different partners. Ultimately, a better understanding of how this type of post-translational modification (PTM) is influencing Arc biology will advance our general understanding of the molecular basis of brain plasticity into new territories. During my postdoctoral training, I used a multidisciplinary approach to search for pharmacological modifiers of Arc expression and function. As part of this investigation, I collected the first evidence of Arc lysine acetylation using tandem mass spectrometry and other complementary biochemical strategies. An initial analysis of the four Arc lysine residues, which I identified as candidate targets for acetylation, revealed that modification of at least one of these sites (K24) could strikingly promote Arc protein accumulation by limiting its polyubiquitination and subsequent proteasomal degradation (Lalonde et al., accepted by Nature Communications). Overall, this finding suggests a strong influence of lysine acetylation on Arc biology and supports the need for a broader investigation into the functional impact, as well as molecular underpinnings, of this modification in neurons. In order to fulfill that goal, I propose to: 1) determine how the individual acetylation of previously identified Arc lysine residues could shape Arc-dependent processes, like AMPA receptor trafficking and dendritic spine morphogenesis, in cultured neurons; 2) establish with the help of proteomic techniques how lysine acetylation changes Arc subcellular distribution and its interaction with other proteins; and 3) uncover the specific acetyltransferase and deacetyltransferase enzymes mediating Arc acetylation in neuronal cells using a targeted RNAi screening strategy. Together, these three complementary lines of research will offer a clear portrait of the influence of lysine acetylation on the neurobiology of Arc, and provide a solid empirical foundation for extending, in the future, our examination of Arc acetylation towards brain development, behavioral control, and the mechanisms of learning and memory.

神经元有非凡的能力来重组它们的结构、功能和连接，以响应刺激。在支持这些神经可塑性变化的许多信号分子和蛋白质效应器中，活性调节的细胞骨架相关蛋白(Arc)可以被认为是中心的，也是最多功能的参与者之一。过去的研究强调了Arc在神经元中的多种作用；然而，是什么允许这种蛋白质承担一种细胞功能，或者与适当的分子效应器相互作用以响应特定的信号，仍然知之甚少。这项发现拨款提案的首要目标是检验这样一个假设，即Arc蛋白精确位点上的赖氨酸乙酰化对其亚细胞分布、功能和/或与不同伙伴的亲和力产生强烈的调节作用。最终，更好地理解这种类型的翻译后修饰(PTM)是如何影响Arc生物学的，将推动我们对大脑可塑性的分子基础的总体理解进入新的领域。在我的博士后培训期间，我使用了一种多学科的方法来寻找Arc表达和功能的药理修饰因素。作为这项研究的一部分，我使用串联质谱仪和其他补充生化策略收集了第一个关于Arc赖氨酸乙酰化的证据。对四个Arc赖氨酸残基的初步分析表明，至少其中一个位点(K24)的修饰可以通过限制其多泛素化和随后的蛋白酶体降解来显著促进Arc蛋白的积累(Lalonde等人，被《自然通讯》接受)。总体而言，这一发现表明了赖氨酸乙酰化对Arc生物学的强烈影响，并支持有必要对这种修饰对神经元的功能影响以及分子基础进行更广泛的研究。为了实现这一目标，我建议：1)确定先前确定的Arc赖氨酸残基的单个乙酰化如何在培养的神经元中形成Arc依赖的过程，如AMPA受体的运输和树突棘的形态发生；2)借助蛋白质组学技术确定赖氨酸乙酰化如何改变Arc的亚细胞分布及其与其他蛋白质的相互作用；以及3)使用靶向RNAi筛选策略揭示介导神经细胞Arc乙酰化的特定乙酰基转移酶和脱乙酰基转移酶。这三条互补的研究路线将为赖氨酸乙酰化对Arc神经生物学的影响提供一个清晰的画像，并为未来我们对Arc乙酰化的研究扩展到大脑发育、行为控制和学习和记忆机制提供坚实的经验基础。