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: 2022
• 资助国家: 加拿大
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=761154
• 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.

神经元具有重组其结构、功能和连接以响应刺激的非凡能力。在众多支持这些神经塑性变化的信号分子和蛋白质效应子中，活性调节细胞骨架相关蛋白 (Arc) 可以被认为是最核心、也是最通用的参与者之一。过去的研究强调了 Arc 在神经元中的多重作用；然而，是什么让这种蛋白质承担一种细胞功能而不是另一种细胞功能，或与适当的分子效应器相互作用以响应特定信号，仍然知之甚少。这项发现资助提案的总体目标是检验这一假设，即 Arc 蛋白在精确位点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，更好地了解这种类型的翻译后修饰 (PTM) 如何影响 Arc 生物学将把我们对大脑可塑性分子基础的一般理解推进到新的领域。在博士后培训期间，我使用多学科方法来寻找 Arc 表达和功能的药理学修饰剂。作为这项调查的一部分，我使用串联质谱和其他补充生化策略收集了 Arc 赖氨酸乙酰化的第一个证据。对我确定为乙酰化候选靶​​标的四个 Arc 赖氨酸残基的初步分析表明，对这些位点中的至少一个 (K24) 进行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着促进 Arc 蛋白的积累（Lalonde 等人，已被 Nature Communications 接受）。总体而言，这一发现表明赖氨酸乙酰化对 Arc 生物学有强烈影响，并支持需要对神经元中这种修饰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）确定先前鉴定的 Arc 赖氨酸残基的单独乙酰化如何在培养的神经元中塑造 Arc 依赖性过程，例如 AMPA 受体运输和树突棘形态发生； 2) 借助蛋白质组学技术确定赖氨酸乙酰化如何改变Arc亚细胞分布及其与其他蛋白质的相互作用； 3) 使用靶向 RNAi 筛选策略揭示神经元细胞中介导 Arc 乙酰化的特定乙酰转移酶和脱乙酰转移酶。这三项互补的研究将共同​​清晰地描述赖氨酸乙酰化对 Arc 神经生物学的影响，并为将来我们对 Arc 乙酰化的研究扩展到大脑发育、行为控制以及学习和记忆机制提供坚实的经验基础。