Dissecting the Role of ATP1A1 in Unconventional Secretion of Fibroblast Growth Factor 2

剖析 ATP1A1 在成纤维细胞生长因子 2 非常规分泌中的作用

• 批准号: 290053622
• 负责人: Professor Dr. Walter Nickel
• 金额: --
• 依托单位: Biochemie-Zentrum (BZH)
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2015
• 资助国家: 德国
• 起止时间: 2014-12-31 至 2019-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/290053622?language=en
• 关键词: Dissecting; Role; ATP1A1; Unconventional; Secretion

Multiple, mechanistically distinct pathways mediate protein secretion from mammalian cells. The classical secretory pathway involves signal peptide dependent protein translocation into the lumen of the endoplasmic reticulum followed by vesicular transport via the Golgi to the plasma membrane. Upon membrane fusion of secretory vesicles with the plasma membrane, cargo proteins destined for the extracelluar space are dispatched on cell surfaces. For long, the presence of a signal peptide has been considered obligatory for the ability of a protein to exit cells. However, many examples of extracellular proteins lacking signal peptides have been identified. This process has been termed unconventional protein secretion with Fibroblast Growth Factor 2 (FGF2) being a prominent example. A detailed understanding of the mechanism of FGF2 secretion from cells is of extraordinary relevance for biomedical research. This is because FGF2 is a key mediator of tumor-induced angiogenesis. In addition, FGF2 is a key survival factor of tumor cells preventing apoptosis by an autocrine secretion-signaling loop that confers resistance of tumor cells against anti-cancer drugs. We have previously demonstrated that FGF2 is secreted from tumor cells by direct translocation across plasma membranes. This process depends on PI(4,5)P2 dependent membrane recruitment, a phosphoinositide enriched in the inner leaflet of plasma membranes. Membrane recruited FGF2 oligomerizes and forms lipidic pores in the plasma membrane, a process that is regulated by Tec kinase mediated tyrosine phosphorylation of FGF2. Membrane pores formed by FGF2 oligomers form dynamic structures that are believed to allow for membrane translocation of FGF2 monomers. This process depends on cell surface heparan sulfate proteoglycans required to trap FGF2 on cell surfaces resulting in directional transport of FGF2 into the extracellular space. In a recent study, we have identified and validated a new component of the secretory machinery of FGF2, the integral membrane protein ATP1A1. ATP1A1 is a plasma membrane resident protein whose cytoplasmic domain forms a direct contact with FGF2. This interaction is required for FGF2 secretion from cells. Using biochemical and structural methods as well as cell-based experiments this research proposal aims at dissecting the molecular mechanism by which ATP1A1 acts as a component of the machinery mediating FGF2 secretion from tumor cells.

多种机制上不同的途径介导哺乳动物细胞的蛋白质分泌。经典的分泌途径涉及信号肽依赖性蛋白质易位到内质网腔中，然后通过高尔基体囊泡运输到质膜。当分泌囊泡与质膜的膜融合时，去往细胞外空间的货物蛋白被分派到细胞表面上。长期以来，信号肽的存在被认为是蛋白质离开细胞的能力所必需的。然而，已经鉴定了许多缺乏信号肽的细胞外蛋白的实例。该过程被称为非常规蛋白质分泌，成纤维细胞生长因子2（FGF2）是一个突出的例子。对细胞分泌FGF2的机制的详细了解对于生物医学研究具有非凡的意义。这是因为FGF2是肿瘤诱导的血管生成的关键介质。此外，FGF2是肿瘤细胞的关键存活因子，其通过自分泌分泌信号传导环防止细胞凋亡，所述自分泌信号传导环赋予肿瘤细胞对抗癌药物的抗性。我们先前已经证明，FGF2是通过直接易位穿过质膜从肿瘤细胞分泌的。该过程依赖于PI（4，5）P2依赖性膜募集，其是质膜内小叶中富集的磷酸肌醇。膜募集的FGF2寡聚化并在质膜中形成多孔孔，这是一个由Tec激酶介导的FGF2酪氨酸磷酸化调节的过程。由FGF2寡聚体形成的膜孔形成动态结构，其被认为允许FGF2单体的膜移位。该过程取决于将FGF 2捕获在细胞表面上所需的细胞表面硫酸乙酰肝素蛋白聚糖，导致FGF 2定向转运到细胞外空间中。在最近的一项研究中，我们已经确定并验证了一个新的组成部分的分泌机制的FGF2，整合膜蛋白ATP 1A1。ATP 1A1是一种质膜驻留蛋白，其胞质结构域与FGF 2形成直接接触。这种相互作用是细胞分泌FGF2所必需的。使用生物化学和结构的方法，以及基于细胞的实验，这项研究的目的是解剖的分子机制，ATP 1A1作为一个组成部分的机械介导的FGF2分泌肿瘤细胞。