A Neural Development mRNA Decay Network

描述（由申请人提供）：发育过程中的基因表达通常只在转录水平上进行研究。然而，一个额外的和必要的水平的控制发生在转录后的mRNA衰变。在神经系统发育过程中，mRNA衰变的调节尤为重要，神经元的结构需要远离其合成位点的mRNA选择性稳定，细胞多样性的产生需要调节增殖和分化的mRNA的程序性衰变。有缺陷的mRNA衰变与许多神经出生缺陷有关，包括脆性x综合征（最常见的遗传性精神损伤形式）和脊髓性肌萎缩症（婴儿死亡的主要遗传原因）。胚胎发育过程中mRNA衰变的研究先前由于缺乏允许在体内，细胞类型特异性测量转录物稳定性的方法而受到阻碍。我们已经开发了一种技术（基于tu标记方法），克服了这一技术挑战，并允许在完整的果蝇胚胎中对mRNA衰变进行神经特异性的全基因组测量。该技术为系统级方法提供了基础，使我们能够构建神经发育mRNA衰变网络。该网络将包含以下信息：在胚胎神经系统中表达的所有mRNA的衰减率，靶向这些mRNA衰减的顺式元件，与这些顺式元件结合的反式rna结合蛋白和mirna，以及mRNA - rna结合蛋白相互作用的空间动力学。这些信息将通过系统级方法获得，该方法结合了野生型和RBP突变胚胎的mRNA衰减率的全基因组分析，候选线性和结构顺式元件的计算鉴定，顺式的分子遗传学定义

DESCRIPTION (provided by applicant): Gene expression during development is often exclusively studied at the level of transcription. However, an additional and essential level of control occurs via post-transcriptional mRNA decay. Regulation of mRNA decay is particularly important during nervous system development, where the structure of neurons requires selective stabilization of mRNAs far from their site of synthesis and the generation of cellular diversity requires the programmed decay of mRNAs that regulate proliferation and differentiation. Defective mRNA decay has been implicated in many neurological birth defects, including fragile X-syndrome (the most common form of hereditary mental impairment) and spinal muscular atrophy (a leading genetic cause of infant mortality). The study of mRNA decay during embryonic development has previously been hindered by the lack of methods allowing in vivo, cell type-specific measurements of transcript stability. We have developed a technique (based on TU-tagging methods) that overcomes this technical challenge and allows neural-specific, genome-wide measurements of mRNA decay in intact Drosophila embryos. This technique provides the foundation for a systems-level approach that will allow us to construct a Neural Development mRNA Decay Network. This network will contain the following information: the decay rates of all mRNAs expressed in the embryonic nervous system, the cis-elements that target these mRNAs for decay, the trans-acting RNA-binding proteins and miRNAs that bind these cis-elements, and the spatial dynamics of mRNA - RNA-binding protein interactions. This information will be obtained using a systems-level approach that combines genome-wide analysis of mRNA decay rates in wildtype and RBP mutant embryos, computational identification of candidate linear and structural cis-elements, molecular genetic definition of cis element effects on transcript stability in vivo, biochemical analysis of RBP-mRNA interactions in vitro and in vivo, and imaging of fluorescent mRNA - RBP interactions within neurons. This work will generate a comprehensive and predictive network map of neural mRNA decay dynamics, thus filling a significant gap in current models of gene expression during neural development.