Macrocyst Development in Dictyostelium Discoideum

The early events of macrocyst development in Dictyostelium discoideum have been investigated using a new culturing system. When cells of opposite mating-types, HM1 and NC4, are cultured together at the appropriate temperature in the dark, giant cells appear, ingest the surrounding amoebae, and develop into macrocysts. Although these giant cells have been assumed to be the products of the fusion of opposite mating-type cells, no experimental evidence to prove this assumption has been obtained using such mixed-culture systems. In order to avoid the complexities involved in mixed-culturing, we have developed a new system involving the separate culture, and later mixing, of opposite mating-type cells. This has enabled us to obtain direct evidence that giant cells are produced by fusion between opposite mating-type cells. Cell fusion occurs immediately after mixing and is completed within 30min. As a number of cells fuse simultaneously, giant cells produced by this method are very large multinucleated cells, and not binucleated zygptes. Using this system we also discovered the following facts related to giant cell formation: (1) cells can acquire their fusion competence without the presence of cells of the opposite mating-type; (2) only HM1 cells require darkness to acquire their fusion competence; (3) the restrictive temperature, 25°C, inhibits the induction of fusion competence in HM1 cells, but not in NC4 cells.