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

描述（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力学。最近，在TGF-¿信号系统上进行了大规模的蛋白质-蛋白质相互作用（PPI）筛选，揭示了一个由数百种相互作用蛋白质组成的复杂网络，其中许多是新的。一个关键的问题是由TGF-¿信号控制的生物过程是否受到这些新发现的蛋白质的调节。TGF-信号控制着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PPI筛选可能是新的发育调节剂的丰富来源，但新的候选药物必须在胚胎背景下进行评估。研究者建议使用非洲爪蟾胚胎来筛选新的TGF-¿信号候选物的发育活性。他的假设是，胚胎功能分析将在TGF-¿PPI筛选中出现的新相互作用蛋白中发现新的发育调节因子。该方法是通过过表达和基因敲低来测试这些蛋白质在非洲爪蟾胚胎中的活性。目的1将评估TGF-¿系统上已发表和正在进行的PPI筛选的结果，以选择新的胚胎试验候选者。在爪蟾胚胎中候选表达的证据将通过EST数据库搜索或cDNA克隆进行初步评估。利用EST数据库、cDNA克隆和热带爪蟾基因组信息收集在神经期或更早时间表达的基因序列信息。Aim 2将在具有抑制mRNA翻译或mRNA前剪接的oligos的胚胎中进行候选基因敲除。这些测试将揭示新的发育调节因子，并扩大我们对TGF-¿信号和胚胎发生的理解。在更广泛的背景下，即将发现的新调控因子将告知各个领域可能通过异常表达或功能影响出生缺陷和疾病的新基因，特别是TGF-¿信号。

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.