The extracellular N terminus of the endothelin B (ETB) receptor is cleaved by a metalloprotease in an agonist-dependent process

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Item Type:	Article
Title:	The extracellular N terminus of the endothelin B (ETB) receptor is cleaved by a metalloprotease in an agonist-dependent process
Creators Name:	Grantcharova, E., Furkert, J., Reusch, H.P., Krell, H.W., Papsdorf, G., Beyermann, M., Schulein, R., Rosenthal, W. and Oksche, A.
Abstract:	The extracellular N terminus of the endothelin B (ET(B)) receptor is susceptible to limited proteolysis (cleavage at R64 downward arrow S65), but the regulation and the functional consequences of the proteolysis remain elusive. We analyzed the ET(B) receptor or an ET(B)-GFP fusion protein stably or transiently expressed in HEK293 cells. After incubation of cells at 4 degrees C, only the full-length ET(B) receptor was detected at the cell surface. However, when cells were incubated at 37 degrees C, N-terminal cleavage was observed, provided endothelin 1 was present during the incubation. Cleavage was not inhibited by internalization inhibitors (sucrose, phenylarsine oxide). However, in cells incubated with both internalization inhibitors and metalloprotease inhibitors (batimastat, inhibitor of TNFalpha-convertase) or metal chelators (EDTA, phenanthroline), the cleavage was blocked, indicating that metalloproteases cleave the agonist-occupied ET(B) receptor at the cell surface. Functional analysis of a mutant ET(B) receptor lacking the first 64 amino acids ([Delta2-64]ET(B) receptor) revealed normal functional properties, but a 15-fold reduced cell surface expression. The results suggest a role of the N-terminal proteolysis in the regulation of cell surface expression of the ET(B) receptor. This is the first example of a multispanning membrane protein, which is cleaved by a metalloprotease, but retains its functional activity and overall structure.
Keywords:	Amino Acid Sequence, Cell Line, Cell Membrane, Cyclic AMP, Drug Dose-Response Relationship, Polyacrylamide Gel Electrophoresis, Green Fluorescent Proteins, Immunoblotting, Inositol Phosphates, Kinetics, Ligands, Luminescent Proteins, Metalloendopeptidases, Molecular Sequence Data, Mutation, Peptides, Protein Binding, Tertiary Protein Structure, Endothelin B Receptor, Endothelin Receptors, Recombinant Fusion Proteins, Temperature, Time Factors, Transfection
Source:	Journal of Biological Chemistry
ISSN:	0021-9258
Publisher:	American Society for Biochemistry and Molecular Biology
Volume:	277
Number:	46
Page Range:	43933-43941
Date:	15 November 2002
Official Publication:	https://doi.org/10.1074/jbc.M208407200
PubMed:	View item in PubMed

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