Embryonic Functional Screen of a TGF?? Protein-Protein Interaction Network

描述(由申请人提供)：系统生物学领域开始定义生物调控过程的复杂动态。最近，对转化生长因子-β信号系统进行了大规模的蛋白质-蛋白质相互作用(PPI)筛选，揭示了数百个相互作用的蛋白质的复杂网络，其中许多是新奇的。一个关键的问题是，由转化生长因子信号控制的生物过程是否受这些新发现的蛋白质的调控。转化生长因子-β信号调控许多重要的生物过程，尤其是细胞分化和胚胎发育。因此，大规模的PPI筛选可能是新的发育调节剂的丰富来源，但新的候选者必须在萌芽状态下进行评估。研究人员建议使用非洲爪哇胚胎来筛选发育活动的新的转化生长因子-β信号候选。他的假设是，胚胎功能分析将在从转化生长因子-PPI筛查中出现的新的相互作用蛋白中发现新的发育调节因子。方法是通过过度表达和基因敲除来测试这些蛋白质在非洲爪哇胚胎中的活性。AIM 1将评估已公布的和正在进行的转化生长因子系统PPI筛查的结果，以选择新的胚胎测试候选者。在非洲爪哇胚胎中候选表达的证据将通过EST数据库搜索或通过cDNA克隆进行初步评估。将使用EST数据库、cDNA克隆和热带非洲爪哇基因组信息来收集在神经细胞或更早时间表达的基因的序列信息。AIM 2将在带有吗啡寡核苷酸的胚胎中进行候选基因敲除，这些寡聚核苷酸可以阻止mRNA翻译或前mRNA剪接。这些测试将揭示新的发育调节因素，并扩大我们对转化生长因子-β信号转导和胚胎发生的理解。在更广泛的背景下，即将发现的新调控因子将向各个领域通报可能通过异常表达或功能，特别是在转化生长因子-β信号转导中影响出生缺陷和疾病的新基因。 与公共健康相关：生长因子信号，如那些由转化生长因子的亲属提供的信号，控制着许多重要的生物学过程，如细胞分化、胚胎发育、伤口愈合和组织/干细胞更新。本申请中提出的测试将揭示在脊椎动物胚胎发育过程中新的转化生长因子-β信号调节因子的潜在功能。这些实验将指向可能影响人类发育过程的新基因，如果它们的活动或表达异常，可能会导致出生缺陷和疾病。

DESCRIPTION (Provided by Applicant): The field of Systems Biology is beginning to define the complex dynamics of biological regulatory processes. Recently, large-scale protein-protein interaction (PPI) screens have been performed on the TGF-¿ signaling system, revealing a complicated network of hundreds of interacting proteins, many novel. A key question is whether biological processes governed by TGF-¿ signals are regulated by these newfound proteins. TGF-¿ signals govern many important biological processes, but particularly cell differentiation and embryonic development. Therefore, large-scale PPI screens are potentially a rich source of new developmental regulators, but new candidates must be evaluated in an embryonic context. The investigator proposes to use Xenopus embryos to screen new TGF-¿ signaling candidates for developmental activity. His hypothesis is that embryonic functional assays will uncover novel developmental regulators among new interacting proteins emerging from TGF-¿ PPI screens. The approach is to test these proteins for activity in Xenopus embryos by overexpression and gene knockdown. Aim 1 will evaluate the results of published and ongoing PPI screens on the TGF-¿ system to select novel candidates for embryonic tests. Evidence of candidate expression in Xenopus embryos will be initially assessed by EST database searches, or by cDNA cloning. Sequence information will be gathered for genes expressed at neurula or earlier times using EST databases, cDNA cloning, and Xenopus tropicalis genomic information. Aim 2 will perform candidate gene knockdown in embryos with morpholino oligos that block mRNA translation or pre-mRNA splicing. These tests will reveal new developmental regulators and expand our understanding of TGF-¿ signaling and embryogenesis. In broader context, new regulators to be uncovered will inform various fields of new genes that could affect birth defects and diseases through abnormal expression or function, particularly in TGF-¿ signaling. PUBLIC HEALTH RELEVANCE: Growth factor signals, such as those provided by relatives of TGF-¿, govern many important biological processes, such as cell differentiation, embryonic development, wound healing, and tissue/stem cell renewal. The tests proposed in this application will reveal potential functions of new regulators of TGF-¿ signaling during vertebrate embryogenesis. The experiments will point to new genes that might affect human developmental processes, and which might cause birth defects and disease if their activity or expression is abnormal.