FIGURE 1:

K63Ub chains are required for efficient sorting to yeast MVBs. SUB cells genetically modified to express only plasmid-encoded wt or single-KR-mutant Ub were transformed with a plasmid encoding Ear1-mCherry (mCh) under the control of the GPD1 promoter. (A) Ear1-mCherry ubiquitylation profile. Total protein extracts separated by SDS–PAGE were immunoblotted with an anti-DsRed antibody. The arrow and arrowhead indicate the monoubiquitylated and polyubiquitylated forms of Ear1-mCherry, respectively. The anti-Pgk1 blot serves as a loading control. (B) Ear1-mCherry sorting to the MVB pathway was assessed by fluorescence microscopy.