(A) Subcellular localizations of Vld1-nG and Vph1-mCh in WT, pep12∆, vps27∆, and apl6∆ cells. (B) Vld1 contains a conserved acidic di-leucine motif at its C-terminus. (C) Subcellular localizations of vld1∆6AA-nG (last 6 amino acids of Vld1 deleted) in WT cell and pep12∆ cells. (D) A single E233 to A mutation caused the trafficking defects of vld1E233A-nG in both WT and pep12∆ cells. (E) Subcellular localizations of Gld1-nG and Vph1-mCh in WT, vps27∆, and pep12∆ cells. (F) Subcellular localizations of Gld1-nG in WT and vps35∆ cells. (G) A model summarizing the key findings of this study. White dashed lines indicate the periphery of yeast cells. Scale bar: 2 µm.