The WASp-like protein Scar regulates macropinocytosis, phagocytosis and endosomal membrane flow in Dictyostelium

Sear, a member of the WASp protein family, was discovered in Dictyostelium discoideum during a genetic screen for second-site mutations that suppresse d a developmental defect. Disruption of the scar gene reduced the levels of cellular F-actin by 50%. To investigate the role of Scar in endocytosis, p hagocytosis and endocytic membrane trafficking, processes that depend on ac tin polymerization, we have analyzed a Dicyostelium cell line that is genet ically null for Scar. Rates of fluid phase macropinocytosis and phagocytosi s are significantly reduced in the scar(-) cell-line. In addition, exocytos is of fluid phase is delayed in these cells and movement of fluid phase fro m lysosomes to post-lysosomes is also delayed. Inhibition of actin polymeri zation with cytochalasin A resulted in similar phenotypes, suggesting that Scar-mediated polymerization of the actin cytoskeleton was important in the regulation of these processes. Supporting this conclusion, fluorescence mi croscopy revealed that some endo-lysosomes were ringed with F-actin in cont rol cells but no F-actin was detected associated with endo-lysosomes in Sca r null cells. Disruption of the two genes encoding the actin monomer seques tering protein profilin in wild-type cells causes defects in the rate of pi nocytosis and fluid phase efflux. Consistent with a predicted physical inte raction between Scar and profilin, disrupting the scar gene in the profilin null background results in greater decreases in the rate of fluid phase in ternalization and fluid phase release compared to either mutant alone. Take n together, these data support a model in which Scar and profilin functiona lly interact to regulate internalization of fluid and particles and later s teps in the endosomal pathway, probably through regulation of actin cytoske leton polymerization.