Functional Genomic Studies of Neuronal Differentiation

描述（由申请人提供）：脊椎动物神经系统内神经元的精确分化是由转录因子相互作用的复杂遗传程序以及环境线索对这些相互作用的调节所指定的。尽管许多特定的转录因子和信号通路在神经元分化中的作用已被表征，但我们对所涉及的详细机制的理解仍远未完成。本提案利用几种哺乳动物基因组的基因组序列信息，脊椎动物神经系统发育在整个进化过程中高度保守的基本观察结果，以及功能基因组学的最新发展，提出了神经元分化的高通量功能基因组分析。在这个提议中要测试的假设是，在神经元分化过程中诱导的基因周围的进化保守序列对神经元诱导的编排很重要。在第一个具体目标中，将诱导P19胚胎癌细胞进行神经元分化，并进行微阵列杂交研究，以鉴定受转录调控的基因。在第二个特定目标中，将使用高通量微阵列转染方法分析P19细胞中进化保守的小鼠基因组非基因序列（cng）的转录调控，这些序列最近被提出代表顺式调控序列簇。在第三个特定目标中，蛋白激酶和转录因子在这些顺式作用序列的调控中的作用将被表征。这些研究的最终结果将是详细了解神经诱导基因的重要顺式调控序列以及特定信号转导途径在调节其转录激活中的作用。这些研究的结果将对未来治疗神经退行性疾病（包括阿尔茨海默病和帕金森病）的治疗方法具有重要意义。

DESCRIPTION (provided by applicant): The precise differentiation of neurons within the vertebrate nervous system is specified by a complex genetic program of transcription factor interactions, as well as modulation of these interactions by environmental cues. Although the roles of many specific transcription factors and signaling pathways have been characterized in neuronal differentiation, our understanding of the detailed mechanisms involved is still far from complete. This proposal employs genomic sequence information of several mammalian genomes, the fundamental observation that vertebrate nervous system development is highly conserved throughout evolution, and recent developments in functional genomics to propose a high-throughput functional genomic analysis of neuronal differentiation. The hypothesis to be tested in this proposal is that evolutionary conserved sequences surrounding genes induced during neuronal differentiation are important to the orchestration of neuronal induction. In the first specific aim, P19 embryonic carcinoma cells will be induced to undergo neuronal differentiation and microarray hybridization studies will be carried out to identify genes that are transcriptionally regulated. In the second specific aim, evolutionarily conserved non-genic sequences (CNGs) of the mouse genome, that have recently been proposed to represent clusters of cis- regulatory sequences, will be analyzed for transcriptional regulation using a high-throughput microarray transfection method in P19 cells. In the third specific aim, the role of protein kinases and transcription factors in the regulation of these cis-acting sequences will be characterized. The end result of these studies will be a detailed understanding of important cis-regulatory sequences of neuronally induced genes and the role of specific signal transduction pathways in the modulation of their transcriptional activation. The results of these studies will be important for future therapeutic approaches to treatment of neurodegenerative diseases including Alzheimer's and Parkinson's diseases.