Mechanisms involved in GPI-anchored-protein signaling dimer formation

各种细胞表面蛋白通过糖基磷脂酰肌醇(GPI)锚定在细胞膜上。虽然这些蛋白质作为信号分子发挥作用，但参与信号传递的机制在很大程度上是未知的。在本研究中，我们以GPI锚定的补体调节蛋白CD59为模型，通过化学交联剂对表面蛋白进行交联化，确定了CD59结合蛋白，揭示了该蛋白本身就是CD59分子。我们还建立了以C9多肽为配体直接测定CD59信号的方法。通过与C9多肽的连接，检测到细胞内钙离子的特异性增加。这些发现使得对CD59信号的直接评估成为可能，而不是通过抗体的交联性激活。GPI锚定的蛋白分布在被称为“筏”的膜域上。小窝蛋白是一种木筏(小窝)上的主要蛋白质，在各种类型的信号传递中起着关键分子的作用。为了了解小窝蛋白是否参与CD59信号转导，我们测试了小窝蛋白在血液和CD59信号转导的血细胞系中的表达。令我们惊讶的是，他们中没有一个人表达了洞察力。只有处于激活状态的成人T细胞白血病(ATL)细胞亚系表达小窝蛋白。我们现在正在利用这些阳性细胞来阐明小窝蛋白在CD59信号转导中的作用。

Various cell surface proteins are anchored to cell membranes via a glycosyl-phosphatidylinositol (GPI) anchor. Although these proteins function as signaling molecules, mechanisms involved in the signaling are largely unknown. In this study we focused on a GPI-anchored complement regulatory protein, CD59, as a model for elucidating the mechanisms involved in the GPI-anchored protein mediating signaling.We identified a CD59-associating protein by cross-linking surface proteins by chemical cross-linker and revealed that the protein is CD59 molecule itself. We also developed the method for direct measurement of CD59 signaling by means of a C9 peptide as a ligand. By ligation with the C9 peptide, specific increase in intracellular Ca2+ was detected. These findings enable the direct assessment of CD59 signaling rather than by activation with Ab cross-linking.GPI-anchored proteins are distributed on membrane domains designated "rafts". Caveolin, a main protein on one type of rafts (caveolae), functions as a key molecule in various types of signaling. To know whether caveolin is involved in CD59 signaling, we tested caveolin expression in blood and blood cell lines in which CD59 signaling is identified. To our surprise none of them expressed caveolin. Only sub line of adult T cell leukemia (ATL) cell lines in a ctivated states expressed caveolin. We are now on the way to elucidate the function of caveolin in CD59 signaling using these positive cells.