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Selective roles for tumor necrosis factor alpha-converting enzyme/ADAM17 in the shedding of the epidermal growth factor receptor ligand family: the juxtamembrane stalk determines cleavage efficiency.
- Source :
-
The Journal of biological chemistry [J Biol Chem] 2004 Jun 04; Vol. 279 (23), pp. 24179-88. Date of Electronic Publication: 2004 Apr 05. - Publication Year :
- 2004
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Abstract
- Epidermal growth factor (EGF) family ligands are derived by proteolytic cleavage of the ectodomains of integral membrane precursors. Previously, we established that tumor necrosis factor alpha-converting enzyme (TACE/ADAM17) is a physiologic transforming growth factor-alpha (TGF-alpha) sheddase, and we also demonstrated enhanced shedding of amphiregulin (AR) and heparin-binding (HB)-EGF upon restoration of TACE activity in TACE-deficient EC-2 fibroblasts. Here we extended these results by showing that purified soluble TACE cleaved single sites in the juxtamembrane stalks of mouse pro-HB-EGF and pro-AR ectodomains in vitro. For pro-HB-EGF, this site matched the C terminus of the purified human growth factor, and we speculate that the AR cleavage site is also physiologically relevant. In contrast, ADAM9 and -10, both implicated in HB-EGF shedding, failed to cleave the ectodomain or cleaved at a nonphysiologic site, respectively. Cotransfection of TACE in EC-2 cells enhanced phorbol myristate acetate-induced but not constitutive shedding of epiregulin and had no effect on betacellulin (BTC) processing. Additionally, soluble TACE did not cleave the juxtamembrane stalks of either pro-BTC or pro-epiregulin ectodomains in vitro. Substitution of the shorter pro-BTC juxtamembrane stalk or truncation of the pro-TGF-alpha stalk to match the pro-BTC length reduced TGF-alpha shedding from transfected cells to background levels, whereas substitution of the pro-BTC P2-P2' sequence reduced TGF-alpha shedding less dramatically. Conversely, substitution of the pro-TGF-alpha stalk or lengthening of the pro-BTC stalk, especially when combined with substitution of the pro-TGF-alpha P2-P2' sequence, markedly increased BTC shedding. These results indicate that efficient TACE cleavage is determined by a combination of stalk length and scissile bond sequence.
- Subjects :
- ADAM Proteins
ADAM10 Protein
ADAM17 Protein
Amino Acid Sequence
Amphiregulin
Amyloid Precursor Protein Secretases
Animals
Betacellulin
Binding Sites
Blotting, Western
Cell Membrane metabolism
DNA, Complementary metabolism
Disintegrins metabolism
EGF Family of Proteins
Enzyme-Linked Immunosorbent Assay
Epidermal Growth Factor chemistry
Epidermal Growth Factor metabolism
Epiregulin
Epitopes
Fibroblasts metabolism
Glycoproteins metabolism
Heparin chemistry
Intercellular Signaling Peptides and Proteins chemistry
Intercellular Signaling Peptides and Proteins metabolism
Ligands
Membrane Proteins metabolism
Metalloendopeptidases metabolism
Mice
Molecular Sequence Data
Precipitin Tests
Protein Binding
Protein Structure, Tertiary
RNA Interference
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Time Factors
Transfection
ErbB Receptors metabolism
Metalloendopeptidases physiology
Subjects
Details
- Language :
- English
- ISSN :
- 0021-9258
- Volume :
- 279
- Issue :
- 23
- Database :
- MEDLINE
- Journal :
- The Journal of biological chemistry
- Publication Type :
- Academic Journal
- Accession number :
- 15066986
- Full Text :
- https://doi.org/10.1074/jbc.M312141200