Two di-leucine-based motifs account for the different subcellular localizations of the human endothelin-converting enzyme (ECE-1) isoforms

Endothelin-converting enzyme (ECE-1) is a type II integral membrane protein which plays a key role in the biosynthetic pathway of the vasoconstricting endothelins, Three ECE-1 isoforms, differing by their N-terminal cytoplasm ic tails, are generated from a single gene. When expressed in CHO cells, th ey display comparable enzymatic activity but whereas ECE-1a is strongly exp ressed at the cell surface, ECE-1b is exclusively intracellular and ECE-1c presents an intermediate distribution. In the present study these different localizations were further described at the ultrastructural level, by elec tron microscope immunocytochemistry, To characterize the motifs responsible for the intracellular localization of ECE-1b we constructed chimeric prote ins and point mutants. Two dileucine-based motifs, contained in the N-termi nal part of ECE-1b, were thus identified. One of these motifs (LV), display ed by both ECE-1b and ECE-1c, accounts for the reduced surface expression o f ECE-1c as compared to ECE-1a. Mutation of both motifs (LL and LV) induces a very strong appearance of ECE-1b at the cell surface indicating that the ir presence in the N-terminal extremity of ECE-1b is critical for its exclu sively intracellular localization.