CRCNS: PKMzeta-Dependent Protein Synthesis Maintains Synaptic Plasticity

描述（由申请人提供）：我们都有可以追溯到青年时代的记忆；我们记得四岁时住的房子；我们记得一位最喜欢的老师。储存这些记忆的机制被认为是特定神经元回路中突触连接的长期可塑性。然而，这种假定的记忆的细胞基础依赖于通常比记忆寿命短得多的蛋白质。这正是长期记忆和突触可塑性的一个基本问题：如何在不断降解的蛋白质的基础上储存人类一生的记忆？最近，研究表明，脑特异性PKC亚型蛋白激酶M （PKM）在维持突触的后期长期增强（L-LTP）和长期记忆中起着独特的作用。然而，这个关键的观察结果并不能解释PKM如何？？可以克服蛋白质周转和扩散的自然降解作用。这个提议的中心假设是PKM?？，通过控制其自身的合成，可以形成一个双稳定的系统，这可以解释突触特定的长期可塑性和记忆的维持。在此，我们提出在生物物理模型中以数学方式表达这一假设，并对该模型进行分析，从而提出可测试的实验预测。然后我们将直接在PKM上测试这些预测。-介导的持续性突触增强，使用专为测试该理论而设计的新技术。智力优势：PKM的发现？？是突触可塑性和长期记忆维持的必要和充分条件，从根本上改变了学习和记忆领域，但真正实现长期可塑性和记忆持久的机制还需要进一步了解。本建议采用理论与实验相结合的方法来解决这些问题。这种双稳定性依赖于翻译调节的理论不仅对神经科学而且对一般生物学都是新颖的。我们的合作是唯一有资格开展所提出的工作，因为Shouval实验室在与实验组合作建模突触可塑性方面有丰富的经验，而Sacktor实验室是PKM科学的先驱。并且对所建议的技术有丰富的经验。实验技术包括两种测试预测所必需的新方法。首先，我们建议测试该模型对L-LTP中蛋白质翻译的预测，而不是使用可能存在毒性和间接作用问题的一般蛋白质合成抑制剂，而是使用反义寡脱氧核苷酸直接指向PKM的翻译起始位点。mRNA特异性阻断PKM?？综合感应和维护。第二，因为PKM？-介导的增强既高度稳定又快速可逆，我们将使用为L-LTP维持研究优化的快流海马切片室来验证该模型的关键预测。翻译依赖双稳定性的随机模拟在计算生物学中也是新颖的。更广泛的影响：作为第一个证明经验依赖的分子机制，PKM?？具有重要的临床意义，并且在过去的一年中已被证明有助于多种神经和精神疾病的生物学研究，包括创伤后应激障碍，中枢神经性疼痛和药物滥用。为了帮助快速增长的PKM兴趣？？在许多实验室中，我们将使我们的模型对更大的社区开放，允许其他科学家测试、修改并将他们的发现合并到模型中，从而加快科学发现的步伐。因为NSF的一个重要目标是整合研究和教育，我们将培养多样化的学生。我们的实验室已经在培训本科生，Shouval实验室每年夏天通过REU项目（PI S. Cox, Rice）和UT系统资助项目（PI H. Shouval）招收本科生，并全年招收当地本科生，Sacktor实验室在指导当地弱势高中生方面有着悠久的历史（例如，通过英特尔项目）。这两个实验室都致力于向公众推广；例如，一篇关于PKM的文章？？《记忆》上了《纽约时报》的头版我们渴望将这种拓展扩展到生物科学理论与实验之间的相互作用领域。

DESCRIPTION (provided by applicant): We all have memories that date back to our youth; we remember the house we lived in at age 4; we remember a favorite schoolteacher. The mechanism for storing these memories is believed to be the long-term plasticity of synaptic connections within specific neuronal circuits. However, this putative cellular basis of memory relies on proteins that typically have lifetimes far shorter than the memory. Here exactly lies a fundamental problem of long-term memory and synaptic plasticity: How can memories be stored for a human lifetime on the basis of proteins that are continuously degrading? Recently, it was shown that the brain-specific PKC isoform, protein kinase M??(PKM?), plays a unique role in maintaining both late long-term potentiation (L-LTP) of synapses and long-term memory. This crucial observation, however, does not explain how PKM??can overcome the natural degrading effect of protein turnover and diffusion. The central hypothesis of this proposal is that PKM??, through its control of its own synthesis, can form a bi-stable system, which can account for the maintenance of synapse specific long-term plasticity and memory. Here we propose to mathematically formulate this hypothesis within a biophysical model, and to analyze this model so as to propose testable experimental predictions. We then will directly test these predictions on PKM?-mediated persistent synaptic potentiation, using novel techniques tailored for testing the theory. Intellectual Merit: The finding that PKM??is both necessary and sufficient for the maintenance of synaptic plasticity and long-term memory has fundamentally changed the field of learning and memory, but much needs to be learned about the mechanisms that can actually accomplish the persistence of long-term plasticity and memory. This proposal addresses these questions using a combined theoretical and experimental approach. Such a theory in which bi-stability depends on regulation of translation is novel not only for neuroscience but also for biology in general. Our collaboration is uniquely qualified to carry out the proposed work because the Shouval lab has ample experience in modeling synaptic plasticity in collaboration with experimental groups, and the Sacktor lab has pioneered the science of PKM??and has ample experience with the proposed techniques. The experimental techniques include two new methodologies necessary for testing the predictions. First, we propose to test the model's predictions on protein translation in L-LTP, not by general protein synthesis inhibitors that may have issues of toxicity and indirect effects, but by use of antisense oligodeoxynucleotides directed to the translation start site of PKM??mRNA to specifically block PKM??synthesis in induction and maintenance. Second, because PKM?-mediated potentiation is both highly stable and yet rapidly reversible, we will use a fast-flow hippocampal slice chamber optimized for the study of the maintenance of L-LTP to test key predictions of the model. The proposed stochastic simulations of translation-dependent bi-stability are also novel in computational biology. Broader Impact: As the first demonstrated molecular mechanism of experience-dependent, long-term information storage in the brain, PKM??has significant clinical implications, and within the last year has been shown to contribute to in the biology of a variety of neurological and psychiatric diseases, including post-traumatic stress disorder, central neuropathic pain, and drug abuse. In order to assist the rapidly growing interest in PKM??in many labs, we will make our model accessible to the larger community, allowing for other scientists to test, modify, and incorporate their findings into the model, thus accelerating the pace of scientific discovery. Because an important goal for NSF is to integrate research and education, we will train a diverse pool of students. Our labs already train undergraduates, the Shouval lab takes undergraduates each summer through an REU program (PI S. Cox, Rice), and a UT system grant (PI H. Shouval), and local undergraduates throughout the year, and the Sacktor lab has had a long history of mentoring local disadvantaged high school students (e.g., through the Intel program). Both labs are dedicated to public outreach; for example, an article on PKM??and memory was on the front page of The New York Times. We are eager to extend this type of outreach to the domain of the interaction between theory and experiment in biological sciences.