Clathrin heavy chain is required for spore cell but not stalk cell differentiation in Dictyostelium discoideum

Abstract

Previous studies of a clathrin-minus Dictyostelium cell line revealed important roles for clathrin heavy chain (clathrin) in endocytosis, secretion of lysosomal hydrolases and osmoregulation. In this paper, we examine the contribution of clathrin-mediated membrane traffic to development in Dictyostelium discoideum. Clathrin-minus cells were delayed in early development. When exposed to starvation conditions, clathrin-minus cells streamed and aggregated more slowly than wild-type cells. Although clathrin-minus cells displayed only 40% the level of extracellular cyclic AMP binding normally found in wild-type cells, they responded chemotactically to extracellular cyclic AMP. Clathrin-minus cells down-regulated cyclic AMP receptors, but only to half the extent of wild-type cells. We found that the extent of development of clathrin-minus cells was variable and influenced by environmental conditions. Although the mutant cells always progressed beyond the tipped mound stage, the final structure varied from a finger-like projection to a short, irregular fruiting body. Microscopic examination of these terminal structures revealed the presence of intact stalks but a complete absence of spores. Clathrin-minus cells expressed prestalk (ecmA and ecmB) and prespore (psA and cotB) genes normally, but were blocked in expression of the sporulation gene spiA. Using clathrin-minus cells that had been transformed with various promoter-lacZ reporter constructs, we saw only partial sorting of clathrin-minus prestalk and prespore cells. Even when mixed with wild-type cells, clathrin-minus cells failed to sort correctly and never constructed functional spores. These results suggest three roles for clathrin during Dictyostelium development. First, clathrin increases the efficiency of early development. Second, clathrin enables proper and efficient patterning of prestalk and prespore cells during culmination. Third, clathrin is essential for differentiation of mature spore cells.