Amyere Mustapha, Mettlen Marcel, Van Der Smissen Patrick, Platek Anna, Payrastre Bernard, Veithen Alex, Courtoy Pierre J
Cell biology Unit, UCL and ICP, Brussels, Belgium.
Int J Med Microbiol. 2002 Feb;291(6-7):487-94. doi: 10.1078/1438-4221-00157.
Macropinocytosis refers to the formation of primary large endocytic vesicles of irregular size and shape, generated by actin-driven evaginations of the plasma membrane, whereby cells avidly incorporate extracellular fluid. Macropinosomes resemble "empty" phagosomes and show no difference with the "spacious phagosomes" triggered by the enteropathogenic bacteria Salmonella and Shigella. Macropinosomes may fuse with lysosomes or regurgitate their content back to the extracellular space. In multiple cell types, macropinocytosis is a transient response to growth factors. When amoebas are cultured under axenic conditions, macropinocytosis is induced so as to fulfil nutritional requirements. In immature dendritic cells, macropinocytosis allows for extensive sampling of soluble antigens; after a few days of maturation, this activity vanishes as processed peptides are being presented. Macropinosomes are also formed at the leading edge of motile leukocytes or neurons. In all these examples, macropinocytosis appears tightly regulated. Transformation of fibroblasts by Src or Ras also results in constitutive formation of macropinosomes at "ruffling" zones, that could be related to accelerated cell motility. Like phagocytosis, macropinocytosis depends on signalling to the actin cytoskeleton. We have explored this signalling in transformed cells. v-Src and K-Ras activate PI3K and PLC, as demonstrated by in situ production of the corresponding lipid products. Pharmacological inhibitors of PI3K and PLC and stable transfection leading to a dominant-negative PI3-kinase construct in transformed fibroblasts abolish macropinocytosis, demonstrating that both enzyme activities are essential. Conversely, stable transfection leading to a dominant-positive P13K in non-transformed fibroblasts is sufficient to induce macropinocytosis. Combination of experiments allows to conclude that P13K and PLC act in sequential order. In non-polarized cells expressing a thermosensitive v-Src mutant, v-Src kinase activation accelerates fluid-phase endocytosis. In polarized MDCK cells, this stimulation occurs selectively at the apical domain and the response is selectively abrogated by pharmacological inhibitors of P13K and PLC. Thus, two paradigmatic oncogenes cause constitutive macropinocytosis. For v-Src, this response is polarized at the apical membrane. It is suggested that, in enterocytes that do not normally phagocytose, the P13K-PLC signalling pathway leading to selective induction of macropinocytosis at the luminal surface has been subverted by enteropathogenic bacteria to penetrate via "spacious phagosomes".
巨吞饮作用是指由肌动蛋白驱动质膜外翻形成大小和形状不规则的初级大型内吞囊泡，借此细胞大量摄取细胞外液。巨吞饮小泡类似于“空的”吞噬小体，与肠道致病菌沙门氏菌和志贺氏菌引发的“宽敞吞噬小体”并无差异。巨吞饮小泡可与溶酶体融合，或将其内容物反吐回细胞外空间。在多种细胞类型中，巨吞饮作用是对生长因子的一种短暂反应。当变形虫在无菌条件下培养时，会诱导巨吞饮作用以满足营养需求。在未成熟的树突状细胞中，巨吞饮作用允许广泛摄取可溶性抗原；在成熟几天后，随着加工后的肽段呈递，这种活动消失。巨吞饮小泡也在运动性白细胞或神经元的前沿形成。在所有这些例子中，巨吞饮作用似乎受到严格调控。Src或Ras介导的成纤维细胞转化也会导致在“边缘波动”区域组成性形成巨吞饮小泡，这可能与细胞运动加速有关。与吞噬作用一样，巨吞饮作用依赖于向肌动蛋白细胞骨架发出信号。我们在转化细胞中探索了这种信号传导。v-Src和K-Ras激活PI3K和PLC，相应脂质产物的原位产生证明了这一点。PI3K和PLC的药理抑制剂以及在转化的成纤维细胞中稳定转染导致显性负性PI3激酶构建体可消除巨吞饮作用，表明这两种酶活性都是必不可少的。相反，在未转化的成纤维细胞中稳定转染导致显性正性P13K足以诱导巨吞饮作用。综合实验可以得出结论，P13K和PLC按顺序起作用。在表达温度敏感型v-Src突变体的非极化细胞中，v-Src激酶激活会加速液相内吞作用。在极化的MDCK细胞中，这种刺激选择性地发生在顶端区域，并且该反应被PI3K和PLC的药理抑制剂选择性地消除。因此，两种典型的癌基因会导致组成性巨吞饮作用。对于v-Src，这种反应在顶端膜处是极化的。有人提出，在正常情况下不进行吞噬的肠细胞中，导致在管腔表面选择性诱导巨吞饮作用的PI3K-PLC信号通路已被肠道致病菌颠覆，使其能够通过“宽敞吞噬小体”穿透。
