Origin, originality, functions, subversions and molecular signalling of macropinocytosis

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".