The majority of clathrin punctae on the plasma membrane (A) and the contractile vacuole (B–D) contain AP2, AP180, and epsin, but not Hip1r. (A and B) Wild-type cells expressing GFP-AP180 (green) and RFP-clathrin light chain (red) were immunostained for AP2α (blue). (C) Epsin localizes on GFP-AP180-labeled contractile vacuoles. Wild-type cells expressing GFP-AP180 (green) and epsin-RFP (red) were immunostained with a contractile vacuole marker, Rh50 (blue). (D) Hip1r does not localize on AP180-labeled contractile vacuoles. Wild-type cells expressing GFP-AP180 (green) were immunostained for Hip1r (red). Contractile vacuoles are indicated by arrows. Scale bar, 10 μm.