Neurotrophin-Induced Regulation of the microRNA Processing Factor Lin28a

描述（由申请人提供）：突触可塑性，被认为是学习和记忆的细胞相关物，涉及响应神经元活动的突触的选择性加强和减弱。基因表达的变化结合新的蛋白质合成长期以来一直被认为是突触连接持久变化的必要条件。然而，令人惊讶的是，最近的研究表明，转录水平的调控不一定与翻译水平的调控相关，因此细胞中mRNA的谱与蛋白质组仅适度相关。脑源性神经营养因子（BDNF）是突触连接长期变化的重要调节因子。它在巩固学习和记忆中起重要作用，并且在广泛的神经和认知障碍中被错误调节，包括情绪障碍、焦虑症和精神分裂症以及神经退行性疾病。有趣的是，BDNF通过调节microRNA生物合成途径的组分来增强转录mRNA的一个非常特异的子集（约4%）的表达。这种调节的一个重要方面是诱导一种名为Lin28a的蛋白质，它是一种多能性因子，可抑制特定microRNA的生物合成，以前被认为不存在于成熟细胞类型，如神经元。该研究的第一个目标是了解BDNF增强Lin28a蛋白从而调节基因表达的分子机制和信号通路。了解BDNF信号通路中涉及的这些机制有可能揭示可能成为未来治疗靶点的调控点。本节中的实验将需要分子、生物化学和遗传学方法来分析培养的小鼠海马神经元中BDNF信号传导结果。此外，拟议研究的第二个目标是在体外和体内神经系统中增强和抑制Lin28a蛋白的活性。这些操作后的细胞形态和行为读数分析将直接表明BDNF介导的Lin28a蛋白诱导如何影响神经元结构以及认知功能。这些实验有可能直接提示认知和神经系统疾病的治疗药物。本节中的方法将包括免疫组织化学，固定细胞成像和分析，以及野生型和遗传操作小鼠中基于学习任务的学习任务的评估。总的来说，拟议的实验将增强我们目前对正常学习和记忆的分子机制的了解，这些机制可能在大脑疾病和紊乱中失调。

DESCRIPTION (provided by applicant): Synaptic plasticity, regarded as the cellular correlate of learning and memory, involves a selective strengthening and weakening of synapses in response to neuronal activity. Changes in gene expression combined with novel protein synthesis have long been recognized as necessary for lasting changes in synaptic connections. However, surprising recent studies have shown that regulation at the level of transcription is not necessarily correlated with regulation at the level of translation, and thus that the profile of mRNA in a cell is only moderately correlated with the proteome. Brain-derived neurotrophic factor (BDNF) is a crucial regulator of long-lasting changes in synaptic connectivity. It plays an important role in consolidation of learning and memory, and is misregulated in a wide range of neurologic and cognitive disorders including mood disorders, anxiety disorders, and Schizophrenia as well as neurodegenerative disease. Interestingly, BDNF enhances the expression of an extraordinarily specific subset (about 4%) of transcribed mRNAs by regulating components of the microRNA biogenesis pathway. One important aspect of this regulation is the induction of a protein called Lin28a, which is a pluripotency factor that inhibits biogenesis of specific microRNAs and was thought previously to be absent from mature cell types such as neurons. The first goal of the proposed research is to understand the molecular mechanisms and signaling pathways through which BDNF enhances Lin28a protein, thus regulating gene expression. Understanding such mechanisms involved in the BDNF signaling pathway has the potential to uncover regulatory points that could be the target of future therapeutics. Experiments in this section will require molecular, biochemical, and genetic approaches to analyze BDNF signaling outcomes in cultured mouse hippocampal neurons. Additionally, the second goal of the proposed research is to enhance and repress activity of Lin28a protein in in vitro and in vivo neuronal systems. Analysis of cell morphological and behavioral readouts following these manipulations will directly indicate how BDNF-mediated induction of Lin28a protein may impact neuronal structure as well as cognitive function. These experiments have the potential to directly suggest therapeutic agents for cognitive and neurologic disorders. Methods in this section will include immunohistochemistry, fixed cell imaging and analysis, and assessment of hippocampal-based learning tasks in wild type and genetically manipulated mice. Overall, the proposed experiments will enhance our current knowledge of the molecular mechanisms that underlie learning and memory normally and that may be dysregulated in diseases and disorders of the brain.