Synthetic defects revealed by uncoupling ESCRT-0 from ESCRT-I. (A) Strains (PLY3734, PLY3779, PLY3777, and PLY3781) of the indicated genotype were assessed for sorting of GFP-Cps1, Ste3-GFP, Ste3-GFP–Ub, and Sna3-GFP. All alleles were stably integrated into the genome as nonepitope alleles under the control of their endogenous promoter. The GFP fluorescence images and corresponding DIC images are shown. (B) The same strains as in A were analyzed for sorting of the lipid marker NBD-PC. (C) Model for function of UBDs in the ESCRT machinery. Model 1 depicts the UBDs of ESCRT-0, -I, and -II acting together to present multiple binding sites for cargo. This model predicts that other UBDs are present in the ESCRT complexes. Model 2 shows that although UBDs of ESCRT-0 bind Ub cargo, the UBDs of ESCRT-I and -II may interact with Ub in other proteins and may regulate the assembly or activity of the MVB sorting apparatus. (D) Schematic of the various UBDs within the ESCRTs and the connections that tie the ESCRTs together in a supercomplex. Bars, 5 µm.