Molecular machinery and signal transduction of phagocytosis and macropinocytosis

巨噬细胞吞噬作用和巨噬细胞吞噬作用分别是细胞内摄取细胞外颗粒和液体的表面运动，在先天免疫和抗原提呈中发挥重要作用。这些细胞表面运动是由驱动F-肌动蛋白重组和膜运输的分子机制介导的，这些机制由复杂的信号转导途径巧妙地控制。本研究的主要目的是解决巨噬细胞吞噬和吞噬过程中肌醇磷脂信号与分子机制之间的关系。在本研究中，我们利用生物成像和图像分析的方法，揭示了大胞质体膜上PI(4，5)P2和PI(3，4，5)P3的水平与肌动蛋白细胞骨架的变化密切相关。PI(4，5)P2在形成巨饮细胞杯的膜褶皱中的浓度比其他区域增加了一倍以上。PI(4，5)P2在接近大针体前达到最大值，在大针体内化后迅速下降。而PI(3，4，5)P3则在大磷脂小体内化前后的大磷脂小体形成部位局部产生。我们的发现表明，I类PI3激酶局部控制的肌醇磷脂水平对于调节肌动蛋白结合蛋白的功能是重要的，肌动蛋白结合蛋白影响细胞表面膜结构的变化。此外，我们还证明了PI(3)P是第三类PI3激酶的主要产物，对于形成的大胞体的融合是必不可少的。

Phagocytosis and macropinocytosis are endocytic cell surface movements for taking extracellular particles such and fluid, respectively, and play important roles in innate immunity and antigen presentation. These cell surface movements are mediated by molecular machineries driving F-actin reorganization and membrane traffic, which are ingeniously controlled by complicated signal transduction pathways. The main aim of this research project is to resolve the relationship between molecular machineries and phosphoinositide signal during phagocytosis and macropinocytosis.In this study using bioimaging and image analysis, we revealed that levels of PI(4,5)P2 and PI(3,4,5)P3 in the membrane of forming macropinosomes were closely linked to the changes in actin cytoskeleton. The concentrations of PI(4,5)P2 in the membrane ruffles forming macropinocytic cups increased more than double that in other areas. The PI(4,5)P2 levels reached its maximum just before closing into macropinosomes, and rapidly fell after macropinosome internalization. In contrast, the PI(3,4,5)P3 was locally produced at the site of macropionosome formation around the time of macropinosome internalization. Our findings suggest that locally controlled levels of phosphoinositides by class I PI3kinase are important for regulating the function of actin-binding proteins which affect changes in the cell surface membrane architecture. Furthermore, we have shown that PI(3)P which is the main product of class III PI3kinase is essential for the fusion of formed macropinosomes.