SSA: Phosphoregulation of argonaute function in synaptic plasticity and memory

SSA：突触可塑性和记忆中阿尔古特功能的磷酸调节

• 批准号: 2588291
• 金额: --
• 依托单位: University of Bristol
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2021
• 资助国家: 英国
• 起止时间: 2021 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2588291
• 关键词: SSA; Phosphoregulation; argonaute; function; synaptic

MicroRNAs (miRNAs) are fundamentally important for fine-tuning protein synthesis by associating with Argonaute (Ago) proteins in the RNA-induced silencing complex (RISC) to repress the translation of target mRNAs. Long-term synaptic plasticity underlies learning and memory by modifying neural circuitry, a major component of which is morphological changes of dendritic spines, which contain the postsynaptic machinery. Regulated miRNA activity plays a key role in this process by modulating the translation of numerous synaptic proteins, and miRNA dysregulation is implicated in neurological disorders that involve synaptic dysfunction, such as Alzheimer's disease. Stimulation of postsynaptic NMDARs and consequent calcium-dependent intracellular signalling is the major upstream component of synaptic plasticity in the brain. Our recent in vitro work identified phosphorylation of Ago2 at Ser387 as an essential mechanism for regulating specific RISC protein-protein interactions and consequent miRNA-dependent gene silencing to control NMDAR-dependent dendritic spine plasticity (Rajgor et al., EMBO J., 2018). In this project, we aim to investigate these mechanisms in vivo; we hypothesise that specific types of synaptic plasticity, and hence memory processes, require Ser387 phosphorylation-dependent changes in RISC protein-protein interactions and miRNA activity. To examine functional roles of Ago2 phosphorylation in synaptic plasticity and memory, this project will compare S387A (phospho-null) Ago2 knock-in mice with wild-type litter-mates in a battery of behavioural tests and brain slice synaptic electrophysiology assays. These will be chosen to assess a diverse range of learning and memory processes, which involve different types of synaptic plasticity. E.g. object recognition memory requires cortical long-term depression (LTD), or spatial memory involves hippocampal long-term potentiation (LTP). To define which miRNAs are involved, we will carry out Ago2 immunoprecipitations from lysates prepared from relevant brain regions, to analyse bound miRNAs by microarrays and RISC assembly by Western blotting. In addition, a proteomics screen will be carried out to analyse RISC protein-protein interactions in an unbiased manner. A role for RISC regulation by Ago2 phosphorylation in learning and memory is as yet unexplored, therefore this project will define novel concepts in miRNA-dependent mechanisms of brain function. It will also provide an excellent training in biochemistry, electrophysiology and behavioural neuroscience. We are looking for an enthusiastic and innovative student with a degree in neuroscience, biological or medical science. The project will be supervised by Prof. Jonathan Hanley (biochemistry), Prof Clea Warburton (behavioural neuroscience) and Dr. Jon Brown (synaptic physiology). For further information, please contact Jonathan Hanley ([e-mail]).

microRNA（miRNAs）通过与RNA诱导沉默复合物（RISC）中的Argonaute（Ago）蛋白结合来抑制靶mRNA的翻译，对于微调蛋白质合成至关重要。长期突触可塑性是学习和记忆的基础，通过修改神经回路，其中一个主要组成部分是树突棘的形态变化，其中包含突触后机制。调节的miRNA活性通过调节许多突触蛋白的翻译在该过程中起关键作用，并且miRNA失调与涉及突触功能障碍的神经系统疾病如阿尔茨海默病有关。突触后NMDAR的刺激和随后的钙依赖性细胞内信号传导是脑中突触可塑性的主要上游组分。我们最近的体外工作鉴定了Ago 2在Ser 387处的磷酸化是调节特定RISC蛋白-蛋白相互作用和随后的miRNA依赖性基因沉默以控制NMDAR依赖性树突棘可塑性的必要机制（Rajgor et al.，EMBO J.，2018年）。在这个项目中，我们的目标是在体内研究这些机制;我们假设特定类型的突触可塑性，因此记忆过程，需要Ser 387磷酸化依赖的RISC蛋白质-蛋白质相互作用和miRNA活性的变化。为了研究Ago 2磷酸化在突触可塑性和记忆中的功能作用，该项目将在一系列行为测试和脑切片突触电生理学测定中比较S387 A（磷酸化无效）Ago 2敲入小鼠与野生型同窝小鼠。这些将被选择来评估各种各样的学习和记忆过程，其中涉及不同类型的突触可塑性。例如，物体识别记忆需要皮质长时程抑制（LTD），或空间记忆涉及海马长时程增强（LTP）。为了确定哪些miRNAs参与，我们将从相关脑区制备的裂解物中进行Ago 2免疫沉淀，通过微阵列分析结合的miRNAs，并通过Western印迹分析RISC组装。此外，还将进行蛋白质组学筛选，以公正的方式分析RISC蛋白质-蛋白质相互作用。在学习和记忆中Ago 2磷酸化对RISC调节的作用尚未被探索，因此该项目将定义脑功能的miRNA依赖机制的新概念。它还将提供生物化学、电生理学和行为神经科学方面的出色培训。我们正在寻找一个热情和创新的学生在神经科学，生物或医学科学学位。该项目将由Jonathan汉利教授（生物化学）、Clea Warburton教授（行为神经科学）和Jon Brown博士（突触生理学）监督。欲了解更多信息，请联系乔纳森汉利（jon.汉利@ bristol.ac.uk）。