Tumor necrosis factor-alpha convertase (ADAM17) mediates regulated ectodomain shedding of the severe-acute respiratory syndrome-coronavirus (SARS-CoV) receptor, angiotensin-converting enzyme-2 (ACE2).
Angiotensin-converting enzyme-2 (ACE2) is a critical regulator of heart function and a cellular receptor for the causative agent of severe-acute respiratory syndrome (SARS), SARS-CoV (coronavirus). ACE2 is a type I transmembrane protein, with an extracellular N-terminal domain containing the active site and a short intracellular C-terminal tail. A soluble form of ACE2, lacking its cytosolic and transmembrane domains, has been shown to block binding of the SARS-CoV spike protein to its receptor. In this study, we examined the ability of ACE2 to undergo proteolytic shedding and investigated the mechanisms responsible for this shedding event. We demonstrated that ACE2, heterologously expressed in HEK293 cells and endogenously expressed in Huh7 cells, undergoes metalloproteinase-mediated, phorbol ester-inducible ectodomain shedding. By using inhibitors with differing potency toward different members of the ADAM (a disintegrin and metalloproteinase) family of proteases, we identified ADAM17 as a candidate mediator of stimulated ACE2 shedding. Furthermore, ablation of ADAM17 expression using specific small interfering RNA duplexes reduced regulated ACE2 shedding, whereas overexpression of ADAM17 significantly increased shedding. Taken together, these data provided direct evidence for the involvement of ADAM17 in the regulated ectodomain shedding of ACE2. The identification of ADAM17 as the protease responsible for ACE2 shedding may provide new insight into the physiological roles of ACE2.
Lambert
Daniel W
DW
Proteolysis Research Group, School of Biochemistry and Microbiology, University of Leeds, Leeds LS2 9JT, United Kingdom. d.w.lambert@leeds.ac.uk
Yarski
Mike
M
Warner
Fiona J
FJ
Thornhill
Paul
P
Parkin
Edward T
ET
Smith
A Ian
AI
Hooper
Nigel M
NM
Turner
Anthony J
AJ
D051722:ADAM Proteins; D000072198:ADAM17 Protein; D053829:Amyloid Precursor Protein Secretases; D000085962:Angiotensin-Converting Enzyme 2; D016282:Aspartic Acid Endopeptidases / Q000378:metabolism; D001665:Binding Sites; D002268:Carboxypeptidases / Q000378:metabolism*; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D019483:Disintegrins / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases; D006801:Humans; D015151:Immunoblotting; D008565:Membrane Proteins / Q000378:metabolism; D008666:Metalloendopeptidases / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D010447:Peptide Hydrolases / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D007703:Peptidyl-Dipeptidase A; D010703:Phorbol Esters; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D020033:Protein Isoforms / Q000737:chemistry; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry; D012330:RNA, Double-Stranded / Q000737:chemistry; D034741:RNA, Small Interfering / Q000378:metabolism; D045473:Severe acute respiratory syndrome-related coronavirus / Q000378:metabolism*; D013997:Time Factors; D014162:Transfection
D018076:DNA, Complementary; D019483:Disintegrins; D008565:Membrane Proteins; D010455:Peptides; D010703:Phorbol Esters; D020033:Protein Isoforms; D012330:RNA, Double-Stranded; D034741:RNA, Small Interfering; D012313:RNA; D053829:Amyloid Precursor Protein Secretases; D002268:Carboxypeptidases; D010450:Endopeptidases; D010447:Peptide Hydrolases; D007703:Peptidyl-Dipeptidase A; C000705307:ACE2 protein, human; D000085962:Angiotensin-Converting Enzyme 2; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human; D051722:ADAM Proteins; C098339:ADAM9 protein, human; D008666:Metalloendopeptidases; D000072198:ADAM17 Protein; C000606462:ADAM17 protein, human