Investigation of experience-dependent post transcriptional regulation of Drosophi

描述(申请人提供)：虽然大量的经历会产生短暂记忆，但其中只有一部分会形成稳定的长期记忆。生物体的经验和内部状态的价值在决定从短期记忆到长期记忆的转变过程中发挥着重要作用。然而，外界经验和生物体内部状态是如何在分子水平上相互作用以影响记忆存储的，这在很大程度上是人们所知的 未知。新蛋白质的合成是形成稳定的长期记忆所必需的。CPEB是一个RNA结合蛋白家族，调节神经元中活性依赖的蛋白质合成。在海兔中，特定的CPEB家族成员ApCPEB对于持续24小时以上的活性依赖性突触强度增加是必需的。在果蝇中，CPEB家族成员Orb2需要将记忆稳定在12小时以上。在小鼠和人类中，CPEB家族成员CPEB3与情景记忆的形成有关。这些观察结果表明，CPEB家族成员在记忆的持久性中起着进化保守的作用。果蝇Orb2基因从两个不同的转录起始点产生两个不同的初级转录本，编码Orb2A和Orb2B蛋白。选择性地使Orb2A蛋白失活的突变专门破坏长期记忆的稳定。我们发现，Orb2AmRNA有两种形式：一种是不编码Orb2A蛋白的内含子非剪接形式(Orb2A-NP)，另一种是编码全长Orb2A蛋白的剪接形式(Orb2A-PC)。令人惊讶的是，Orb2A-NP mRNA是成年大脑中普遍存在的异构体，暴露于形成果蝇关联性长期记忆的线索中，例如饥饿的果蝇暴露在蔗糖和气味中，会增加编码拼接的Orb2A-PC转录本的蛋白质水平。哺乳动物剪接调节因子Nova的果蝇同源基因能结合Orb2A转录本，并调控Orb2A-PC mRNA的形成。在这个建议中，我们打算剖析行为线索和依赖NovA的分子机制，这些分子机制调节蛋白质编码Orb2A转录本的水平。我们还打算解决何时以及多长时间需要Orb2蛋白来保持记忆。我们希望，了解稳定记忆的经验和分子之间的相互作用，将有助于理解成瘾和创伤后综合征，在这些症状中，单一的经验可以形成极其稳定的记忆。此外，我们还希望深入了解受调控的信使核糖核酸在长时间记忆中的作用。

DESCRIPTION (provided by applicant): Although a large number of experiences produce transient memory only some of them form stable long-term memory. The value of the experience and internal state of the organism plays an important role in determining this transition from short-term to long-term memory. However, how external experience and the internal state of the organism interact at a molecular level to influence memory storage is largely unknown. The synthesis of new proteins is required to form stable long-term memory. CPEB, a family of RNA-binding proteins, regulate activity-dependent protein synthesis in the neuron. In Aplysia, a specific CPEB family member, ApCPEB, is required for the persistence of activity-dependent increase in synaptic strength beyond 24 hours. In Drosophila, a CPEB family member Orb2 is required to stabilize memory beyond 12 hours. In mice and human a CPEB family member CPEB3 has been linked to episodic memory formation. These observations suggest that CPEB family members play evolutionarily conserved role in the persistence of memory. The Drosophila Orb2 gene produces two distinct primary transcripts from two different transcription start sites, encoding the Orb2A and Orb2B proteins. Mutations that selectively inactivate Orb2A protein specifically destabilize long-term memory. We have found that the Orb2A mRNA has two forms; an intron containing unspliced form that does not code for Orb2A protein (Orb2A-np) and a spliced form that codes for full length Orb2A protein (Orb2A-pc). Surprisingly, the Orb2A-np mRNA is the prevalent isoform in the adult brain and exposure to cues that form associative long-term memory in flies, such as exposure of hungry flies to sucrose and odor increases the level of protein coding spliced Orb2A-pc transcript. The Drosophila orthologue of mammalian splicing regulator Nova binds Orb2A transcript and regulates the formation of Orb2A-pc mRNA. In this proposal we intend to dissect the behavioral cues and Nova-dependent molecular mechanism that regulate the level of protein coding Orb2A transcript. We also intend to address when and how long Orb2 proteins are required for persistence of memory. We hope that an understanding of the interaction of experiences and molecules that stabilize memory would be useful to understand addictions and post traumatic syndromes, in which a single experience forms extremely stable memory. Also, we hope to gain insight into the role of regulated mRNA processing in long-lasting memory.