Mechanism of phagocytosis in dictyostelium discoideum: phagocytosis is mediated by different recognition sites as disclosed by mutants with altered phagocytotic properties

Abstract

The recognition step in the phagocytotic process of the unicellular amoeba dictyostelium discoideum was examined by analysis of mutants defective in phagocytosis, Reliable and simple assays were developed to measure endocytotic uptake. For pinocytosis, FITC-dextran was found to be a suitable fluid-phase marker; FITC-bacteria, latex beads, and erythrocytes were used as phagocytotic substrates. Ingested material was isolated in one step by centrifuging through highly viscous poly(ethyleneglycol) solutions and was analyzed optically. A selection procedure for isolating mutants defective in phagocytosis was devised using tungsten beads as particulate prey. Nonphagocytosing cells were isolated on the basis of their lower density. Three mutant strains were found exhibiting a clear-cut phenotype directly related to the phagocytotic event. In contrast to the situation in wild-type cells, uptake of E. coli B/r by mutant cells is specifically and competitively inhibited by glucose. Mutant amoeba phagocytose latex beads normally but not protein-coated latex, nonglucosylated bacteria, or erythrocytes. Cohesive properties of mutant cells are altered: they do not form EDTA-sensitive aggregates, and adhesiveness to glass or plastic surfaces is greatly reduced. Based upon these findings, a model for recognition in phagocytosis is proposed: (a) A lectin-type receptor specifically mediates binding of particles containing terminal glucose (E. coli B/r). (b) A second class of "nonspecific" receptors mediate binding of a variety of particles by hydrophobic interaction. Nonspecific binding is affected by mutation in such a way that only strongly hydrophobic (latex) but not more hydrophilic particles (e.g., protein-coated latex, bacteria, erythrocytes) can be phagocytosed by mutant amoebae.

Lipopolysaccharide enhances bactericidal activity in Dictyostelium discoideum cells

“…After 45 min at 22°C, cells were harvested by layering 1 × 10 6 cells over a cushion of polyethylene glycol 3500 and centrifuging to separate D. discoideum cells from non-internalized bacterial particles (Vogel et al, 1980). The levels of phagocytosis of the fluorescent bacterial particles were measured using flow cytometry.…”

Pulsed-laser creation and characterization of giant plasma membrane vesicles from cells

“…Before irradiation, dextran did not interact with any particular cellular component, but was slowly taken into the cytoplasm by pinocytosis [37]. Before irradiation, cells incorporated some dextran, but the concentration of dextran in the surrounding media was greater than the concentration in the cytoplasm, as determined by relative fluorescence intensity.…”

The cytohesin paralog Sec7 of Dictyostelium discoideum is required for phagocytosis and cell motility

“…Such a differential behavior might be explained by earlier findings showing that D. discoideum can differentiate particles according to their surface properties and geometry. Furthermore, analysis of the actin organization around the phagosome revealed a distinct and dynamic spatiotemporal pattern [22,28]. A role of Sec7 in phagocytosis is also supported by the potential binding partners as several of them have been implicated in this process and found associated with the phagosome [29].…”