Role of Sphingosine Kinase 1 in the p53-dependent DNA Damage Response

描述（由申请人提供）：在任何时候，细胞都需要能够对能够诱导DNA损伤的环境压力源做出反应。各种各样的保护机制确保DNA损伤得到修复，而不是传递给下一代细胞。参与这一过程的关键分子之一是p53蛋白。当p53状态受损时，细胞无法应对环境压力，DNA受损细胞的复制导致癌症，这是美国第二大死亡原因。我们实验室之前的研究结果表明，p53信号对DNA损伤的反应可能与鞘脂有关。鞘脂是细胞生长和死亡的重要调节因子。生物活性脂质神经酰胺和鞘氨醇作为促死亡分子，鞘氨醇-1-磷酸（SIP）已被证明能刺激细胞增殖和血管生成。调节这些前死亡分子和促生长分子之间平衡的主要酶是鞘氨酸激酶1 （SK1）。我们的初步结果表明，在DNA损伤诱导p53时，神经酰胺水平升高，鞘氨酸激酶1水平降低。因此，生物活性鞘脂水平的调节可能是p53 DNA损伤反应的关键组成部分，这些途径的相互作用值得进一步研究。通过了解这些途径的细节，我们将能够确定治疗兴趣的靶点。该项目的目标是确定SKI在介导p53依赖性DNA损伤反应中的作用。该项目将利用野生型和p53敲除小鼠胚胎成纤维细胞来研究依赖于基因毒性应激诱导p53的鞘脂水平的改变。小鼠模型也将用于评估这些信号机制如何与体内癌症发展有关。本研究的目的是：1)确定基因毒性应激和p53诱导对SK1/S1P通路的影响；2)确定细胞中p53依赖性SKI降解的机制和表现；3)在体内评估p53通路与SKI/SIP通路的联系。我们的长期目标是确定鞘脂参与细胞应激反应和有机癌症发展的机制，这个项目代表了我们在实现这一目标方面迈出的一步。

DESCRIPTION (provided by applicant): At any time, cells need to be able to respond to environmental stressors that are capable of inducing DNA damage. Various protective mechanisms are in place to ensure that DNA damage is repaired and not passed on to the next generation of cells. One of the key molecules involved in this process is the protein p53. When p53 status is compromised, the cell is left unable to cope with environmental stress and the replication of cells with damaged DNA results in cancer, the second greatest cause of mortality in the United States. Previous results in our lab suggest that p53 signalling, in response to DNA damage, may involve sphingolipids. Sphingolipids are important regulators of cell growth and death. While the bioactive lipids ceramide and sphingosine act as pro-death molecules, sphingoslne-1-phosphate (SIP) has been shown to stimulate proliferation and angiogenesis. The primary enzyme regulating the balance between these prodeath and pro-growth molecules is sphingosine kinase 1 (SK1). Our preliminary results show increased ceramide levels and decreased sphingosine kinase 1 levels in response to the induction of p53 via DNA damage. Thus, regulation of bioactive sphingolipid levels may be a key component in the p53 DNA damage response and the interaction of these pathways warrants further investigation. By understanding the details of these pathways we will be able to identify targets of therapeutic interest. The goal of this project is to determine the role of SKI in mediating the p53-dependent DNA damage response. This project will utilize wild type and p53 knockout mouse embryonic fibroblasts to investigate alterations in sphingolipid levels that are dependent on the induction of p53 by genotoxic stress. A mouse model will also be used to evaluate how these signaling mechanisms pertain to cancer development in vivo. The aims of this study are 1) determine the effects of genotoxic stress and p53 induction on the SK1/S1P pathway; 2) determine the mechanism and manifestations of p53 dependent SKI degradation in the cell; and 3) evaluate the connection between the p53 pathway and SKI/SIP pathway in vivo. This project represents an incremental step in achieving our long-term goal of determining the mechanism of involvement of sphingolipids in the cellular stress response and in organismal cancer development. PUBLIC HEALTH RELEVANCE: When human cells experience excessive stress that results in damage to their DNA, p53 levels go up to prevent the cell from passing on damaged genetic Information which could lead to cancer. This project seeks to determine how p53 regulates bioactive signalling lipids to achieve this function. This work could eventually lead to the development of more efficient cancer therapeutics with less severe side effects.