Role of PLSCR in Cell Response to Growth Factors

描述(申请人提供)：磷脂加扰酶1(PLSCR1)是一种多棕榈酰化的界面质膜蛋白，最初在红细胞、血小板、白细胞和其他血细胞中被发现，被认为在细胞激活、损伤或凋亡时介导膜磷脂的跨双层运动。已发现PLSCR1与多种生长因子受体相关，包括表皮生长因子受体(EGF-R)。PLSCR1的表达受EGF和相关生长因子的转录诱导，并被参与EGF-R和相关受体启动的信号转导通路的酪氨酸激酶磷酸化。我们最近观察到新合成的PLSCR1可以定位到细胞核，我们发现每当多肽不能棕榈酰化时，就会发生PLSCR1的核输入。相反，棕榈酰化的PLSCR1是质膜脂“筏”的组成部分，富含胆固醇和鞘磷脂的膜微域被认为参与了受体信号平台的组装和激活的EGF-R及其相关受体从质膜的内向转运。我们还观察到，在PLSCR1-/-小鼠中，中性粒细胞的产生和成熟对选定的生长因子的反应是受损的，而这些动物的成纤维细胞和上皮细胞对EGF和相关生长因子的反应减弱，特别是对EGF依赖的c-Src激活的影响。在本项目中，我们旨在阐明PLSCR1在EGF-R和相关生长因子受体的信号通路中的作用。我们的具体目标是：Aim1--确定质膜PLSCR1的表达水平是否影响细胞表面的表达、拓扑结构、对配体的敏感性和/或受体启动的EGF和相关生长因子对信号通路的激活。AIM2--确定PLSCR1是否影响EGF-R在膜脂筏上的定位，它与关键的适配蛋白和信号通路的相互作用，或者它的内吞摄取，内体转运，以及受体激活后的降解。AIM3-确定PLSCR1进入细胞核的机制，并确定核PLSCR1在细胞因子刺激的细胞转录反应中可能发挥的作用。我们相信，这项研究将为通过激活的生长因子受体影响信号的调控机制提供新的见解。

DESCRIPTION (provided by applicant): Phospholipid scramblase 1 (PLSCR1) is a multiply palmitoylated, endofacial plasma membrane protein first identified in red cells, platelets, leukocytes, and other blood cells, which is thought to mediate the transbilayer movement of membrane phospholipids in response to cell activation, injury, or apoptosis. PLSCR1 has been found associated with several growth factor receptors, including epidermal growth factor receptor (EGF-R). PLSCR1 expression is transcriptionally induced by EGF and related growth factors, and it is phosphorylated by tyrosine kinases that are involved in the signal transduction pathways initiated through EGF-R and related receptors. We recently observed that newly synthesized PLSCR1 can localize to the nucleus, and we found nuclear import of PLSCR1 to occur whenever the polypeptide fails to palmitoylate. By contrast, palmitoylated PLSCR1 is a component of plasma membrane lipid "rafts", cholesterol- and sphingomyelin-rich membrane microdomains that are believed to be involved in both assembly of receptor signaling platforms and the endocytic trafficking of activated EGF-R and related receptors from the plasma membrane. We also observed that neutrophil production and maturation in response to select growth factors is impaired in PLSCR1-/-mice, whereas fibroblasts and epithelial cells from these animals exhibit attenuated responses to EGF and related growth factors, with particular effect on EGF-dependent activation of c-Src. In this Project, we aim to elucidate the role of PLSCR1 in the signaling pathways of EGF-R and related growth factor receptors. Our Specific Aims are: Aim1 -- To determine whether the level of expression of plasma membrane PLSCR1 influences cell surface expression, topology, sensitivity to ligand, and/or receptor-initiated activation of signaling kinases by EGF and related growth factors. AIM2 -- To determine whether PLSCR1 influences the localization of EGF-R to membrane lipid rafts, its interactions with key adaptor proteins and signaling kinases, or its endocytic uptake, endosomal trafficking, and degradation following receptor activation. AIM3 - To identify the mechanism by which PLSCR1 traffics into the nucleus, and to determine what role nuclear PLSCR1 might play in the cellular transcription response to cytokine stimulation. We believe that this research will provide new insights into the regulatory control mechanisms that affect signaling by activated growth factor receptors.