Identification and characterization of novel RSK substrates

Ras依赖性细胞外信号调节激酶（ERK）级联反应响应于广泛的细胞外刺激以调节多种生物学功能，包括细胞增殖、分化、迁移和存活。该途径在胚胎发生过程中起着特别重要的作用，其中ERK信号传导调节生长因子下游的各种发育过程，包括原肠胚形成、神经模式形成、肢体发育和胎盘形成。因此，在胚胎发育和成年期，这种信号通路的适当调节是至关重要的，因为这种级联中的激活/失活突变会导致各种发育障碍和疾病。 Ras/ERK信号级联的下游是丝氨酸/苏氨酸激酶（RSK 1 -4）的p90核糖体S6激酶（RSK）家族。这些酶被认为是参与细胞生长和增殖的多功能ERK效应子，但关于它们的具体靶点和作用机制知之甚少。多年来，我们的实验室已经开发出小鼠模型，专门解决Ras/ERK通路下游RSK亚型的作用。在这个建议中，我们打算使用定量蛋白质组学方法来确定新的磷酸化底物的RSK亚型。这种方法已经导致了一种新的RSK底物的鉴定，我们打算详细研究并确定RSK在其调节中的作用。 该项目提供了一个很好的机会，培养研究生在生物科学的研究，包括细胞生物学，小鼠遗传学，分子生物学，生物化学，显微镜和蛋白质组学。从长远来看，我们正在开发的技术将有助于解决RSK信号的其他方面，这些结果将成为我们未来几年研究的基础。

The Ras-dependent extracellular signal-regulated kinase (ERK) cascade responds to a wide array of extracellular stimuli to regulate diverse biological functions, including cell proliferation, differentiation, migration and survival. This pathway plays particularly important roles during embryogenesis, where ERK signalling regulates various developmental processes downstream of growth factors, including gastrulation, neural patterning, limb development and placentation. Proper regulation of this signalling pathway is thus crucial during embryogenesis and adulthood, as activating/inactivating mutations within this cascade lead to various developmental disorders and diseases. Downstream of the Ras/ERK signalling cascade lies the p90 ribosomal S6 kinase (RSK) family of serine/threonine kinases (RSK1-4). These enzymes are thought to be multifunctional ERK effectors involved in cell growth and proliferation, but relatively little is known regarding their specific targets and mechanisms of action. Through the years, our lab has developed mouse models to specifically address the role of the RSK isoforms downstream of the Ras/ERK pathway. In this proposal, we intend to use a quantitative proteomics method to identify novel phosphorylation substrates of the RSK isoforms. This approach has already resulted in the identification of a novel RSK substrate, which we intend to study in detail and determine the role of RSK in its regulation. This project provides a great opportunity to train graduate students for research in the biological sciences, including cell biology, mouse genetics, molecular biology, biochemistry, microscopy and proteomics. In the long-term, the technology that we are developing will be useful to address other aspects of RSK signalling and these results will be the basis of our research for years to come.