Characterization of microRNAs expressed in the developing songbird brain

描述（由申请人提供）：在发育中的鸣禽大脑中表达的microRNA的表征项目概述人类儿童的语言发育和鸣禽的歌曲学习之间有许多相似之处。因此鸣禽为研究控制神经回路发育和发声学习的分子和细胞机制提供了一个独特的模型系统。MicroRNA是一类非蛋白质编码RNA分子，其在广泛的生物事件中，特别是在发育期间，在转录后调节基因表达。我们计划研究miRNAs在斑胸草雀神经发育和可塑性中的调节功能，其中鸣唱行为和潜在的神经回路已经被广泛建立。在这个为期两年的探索性项目中，我们将专注于系统的全基因组表征斑胸草雀大脑中的miRNA表达，跨越鸟类的整个生命周期。该项目有两个具体目标。在Specific Aim I中，将从从早期胚胎阶段到老年的不同发育阶段的斑胸草雀脑中制备小RNA文库;将使用超高通量Solexa测序技术对文库进行测序;将使用生物信息学分析对miRNA候选序列进行注释。在Specific Aim II中，将对选定的一组miRNA候选物进行北方印迹杂交和原位杂交，以验证它们的表达，并将它们的表达映射到回路发育期间的鸣唱控制核。优先考虑参与神经发育，可塑性和发育障碍的miRNAs，以及预测靶基因参与这些过程的miRNAs。该初始实验将生成必要的信息（例如，时间和空间的miRNA表达在发展中的歌曲系统），从而建立了我们的长期研究的阶段，在神经发育/可塑性和发育障碍的作用的miRNA。鸣禽大脑中系统的全基因组miRNA谱可能会扩展miRNA库，并且它们在发育过程中的表达模式可能会为其功能提供见解。除了我们自己的工作之外，实验鉴定的miRNA序列可能用于斑胸草雀基因组序列的注释。此外，鸣禽和其他物种中的miRNA的比较分析可能会增加我们对miRNA基因家族进化的总体理解。公共卫生相关性：microRNA调节基因表达并在脑发育中发挥重要作用;因此，了解miRNA功能的分子机制对于了解正常脑发育过程和发育障碍至关重要。miRNA研究的重要性已经被以下事实所证明：目前已经有几种基于RNAi的药物正在临床试验中进行测试，用于从癌症到眼疾的疾病。我们的实验结果，其中在神经回路发育和发声学习的背景下研究miRNA，可能有助于理解并最终治愈人类儿童的发育障碍（例如，自闭症、诵读困难等）。

DESCRIPTION (provided by applicant): Characterization of microRNAs expressed in the developing songbird brain Project Summary Many parallels are shared between language development in human children and song learning in songbirds. Thus songbirds provide a unique model system to study the molecular and cellular mechanisms controlling neural circuit development and vocal learning. MicroRNAs are a class of non-protein coding RNA molecules that regulate gene expression post transcriptionally in a wide array of biological events, especially during development. We plan to study the regulatory functions of miRNAs in neural development and plasticity in zebra finches, where both the song behavior and the underlying neural circuits have been extensively established. In this two-year exploratory project, we will focus on a systematic genome-wide characterization of miRNA expression in the zebra finch brain across a bird's entire life span. This project has two specific aims. In Specific Aim I, small RNA libraries from zebra finch brains at different developmental stages ranging from the early embryonic stage to old age will be made; the libraries will be sequenced using the ultra-high-throughput Solexa sequencing technology; miRNA candidate sequences will be annotated with Bioinformatic analysis. In Specific Aim II, Northern blot hybridization and in situ hybridization will be performed for a selected group of miRNA candidates to validate their expression and to map their expression to the song control nuclei during circuit development. Priorities will be given to miRNAs that have been implicated in neural development, plasticity, and developmental disorders, as well as miRNAs whose predicted target genes have been implicated in these processes. This initial experiment will generate necessary information (e.g., the temporal and spatial miRNA expression in the developing song system), thus setting up the stage for our longer-range research on the roles of miRNAs in neural development/plasticity and in developmental disorders. A systematic genome-wide miRNA profiling in the songbird brain may extend the miRNA repertoire and their expression patterns during development may provide insights to their functions. In addition to our own work, the experimentally identified miRNA sequences may be used in the annotation of the zebra finch genome sequence. Also, comparative analysis of miRNAs in songbirds and other species may add to our understanding of the evolution of miRNA gene family in general. PUBLIC HEALTH RELEVANCE: MicroRNAs regulate gene expression and play important roles in brain development; therefore, understanding the molecular mechanisms of miRNA function is essential for understanding normal brain developmental processes and developmental disorders. The importance of miRNA research has been exemplified by the fact that currently there are already several RNAi-based drugs being tested in clinical trials for illnesses ranging from cancer to eye disease. Results from our experiment, where miRNAs are studied in the context of neural circuit development and vocal learning, may contribute to understanding and eventually curing developmental disorders in human children (e.g., autism, dyslexia, etc).