The N-terminal segment of endothelin-converting enzyme (ECE)-1b contains a di-leucine motif that can redirect neprilysin to an intracellular compartment in Madin-Darby canine kidney (MDCK) cells.
Endothelin-converting enzyme (ECE)-1 is a membrane-bound metallopeptidase of the neprilysin (NEP) family. ECE-1 is responsible for the conversion of inactive big-endothelins into active endothelins. Three different isoforms of human ECE-1 (ECE-1a, ECE-1b and ECE-1c) have been identified. They differ in their N-terminal cytosolic regions, have distinct tissue distribution and intracellular localization. ECE-1a and ECE-1c are both located at the cell surface whereas ECE-1b is targeted to an intracellular compartment. To better understand the nature of the signal responsible for the targeting of ECE-1b to the intracellular compartment, we have constructed several ECE/NEP chimaeric proteins and expressed them by transfection into Madin-Darby canine kidney (MDCK) cells. This allowed us to identify a nine amino acid segment in the cytosolic tail of ECE-1b that is sufficient to relocate NEP from the cell surface to an intracellular compartment. Site-directed mutagenesis on these chimaeras led to the identification of two leucine residues as part of the intracellular retention signal.
Cailler
F
F
Department of Biochemistry, Faculty of Medicine, University of Montreal, CP 6128, Succursale Centre Ville, Montreal, Quebec, H3C 3J7, Canada.
Zappulla
J P
JP
Boileau
G
G
Crine
P
P
D000595:Amino Acid Sequence; D000818:Animals; D016282:Aspartic Acid Endopeptidases / Q000235:genetics / Q000378:metabolism*; D002451:Cell Compartmentation; D002478:Cells, Cultured; D004285:Dogs; D000072396:Endothelin-Converting Enzymes; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D007668:Kidney / Q000166:cytology; D007930:Leucine; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D008666:Metalloendopeptidases / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D015260:Neprilysin / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D007527:Isoenzymes; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D016282:Aspartic Acid Endopeptidases; D008666:Metalloendopeptidases; D015260:Neprilysin; C000606923:ECE1 protein, human; D000072396:Endothelin-Converting Enzymes; D007930:Leucine