Processing of pro-atrial natriuretic peptide by corin in cardiac myocytes.
Corin is a type II transmembrane serine protease abundantly expressed in the heart. In a previous study using transfected 293 cells, we showed that corin converted pro-atrial natriuretic peptide (pro-ANP) to atrial natriuretic peptide (ANP), suggesting that corin is likely the pro-ANP convertase. Because other serine proteases such as thrombin and kallikrein had previously also been shown to cleave pro-ANP in vitro, it remained to demonstrate that corin is indeed the endogenous pro-ANP convertase in cardiomyocytes. In this study, we examined pro-ANP processing in a murine cardiac muscle cell line, HL-5. Northern analysis showed that corin mRNA was present in HL-5 cells. In HL-5 cells transfected with a plasmid expressing pro-ANP, recombinant pro-ANP was converted to mature ANP as determined by Western analysis, indicating the presence of the endogenous pro-ANP convertase in these cells. The processed recombinant ANP was shown to be active in an enzyme-linked immunosorbent assay-based cGMP assay in baby hamster kidney cells. The processing of recombinant pro-ANP in HL-5 cells was highly sequence-specific, because mutation R98A, but not mutations R101A and R102A, in pro-ANP prevented the conversion of pro-ANP to ANP. Expression of recombinant wild-type corin enhanced the processing of pro-ANP in HL-5 cells. In contrast, overexpression of active site mutant corin S985A or transfection of oligonucleotide small interfering RNA duplexes directed against the mouse corin gene completely inhibited the processing of recombinant pro-ANP in HL-5 cells. These results indicate that corin is the physiological pro-ANP convertase in cardiac myocytes.
Wu
Faye
F
Department of Cardiovascular Research, Berlex Biosciences, Richmond, California 94804, USA.
Yan
Wei
W
Pan
Junliang
J
Morser
John
J
Wu
Qingyu
Q
D000818:Animals; D009320:Atrial Natriuretic Factor / Q000378:metabolism*; D015152:Blotting, Northern; D015153:Blotting, Western; D002460:Cell Line; D006224:Cricetinae; D006152:Cyclic GMP / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D051379:Mice; D009154:Mutation; D009206:Myocardium / Q000166:cytology* / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D011498:Protein Precursors / Q000378:metabolism*; D012313:RNA / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000378:metabolism*; D014018:Tissue Distribution; D014162:Transfection
D011498:Protein Precursors; D011994:Recombinant Proteins; D012313:RNA; D009320:Atrial Natriuretic Factor; C119812:CORIN protein, human; C485617:Corin protein, mouse; D012697:Serine Endopeptidases; D006152:Cyclic GMP