Functional analysis of corin protein domains required for PCSK6-mediated activation.
Atrial natriuretic peptide (ANP) is a cardiac hormone essential for normal blood pressure and cardiac function. Corin is a transmembrane serine protease that activates ANP. Recently, we identified proprotein convertase subtilisin/kexin-6 (PCSK6), also called PACE4, as the long-sought corin activator. Both corin and PCSK6 are expressed in cardiomyocytes, but corin activation occurs only on the cell surface. It remains unknown if cell membrane association is needed for PCSK6 to activate corin. Here we expressed corin deletion mutants in HEK293 cells to analyze the domain structures required for PCSK6-mediated activation. Our results show that soluble corin lacking the transmembrane domain was activated by PCSK6 in the conditioned medium but not intracellularly. Recombinant PCSK6 also activated the soluble corin under cell-free conditions. Moreover, PCSK6-mediated corin activation was not enhanced by cell membrane fractions. These results indicate that cell membrane association is unnecessary for PCSK6 to activate corin. Experiments with monensin that blocks PCSK6 secretion and immunostaining indicated that the soluble corin and PCSK6 were secreted via different intracellular pathways, which may explain the lack of corin activation inside the cell. We also found that the protein domains in the corin pro-peptide region were dispensable for PCSK6-mediated activation and that addition of heparan sulfate and chondroitin sulfate or treatment with heparinase or chondroitinase did not alter corin activation by PCSK6 in HEK293 cells. Together, our results provide important insights into the molecular and cellular mechanisms underlying PCSK6-mediated corin activation that is critical for cardiovascular homeostasis.
['Chen|Shenghan|S|', 'Wang|Hao|H|', 'Li|Heng|H|', 'Zhang|Yue|Y|', 'Wu|Qingyu|Q|']
D019943:Amino Acid Substitution D000818:Animals D020134:Catalytic Domain D002460:Cell Line D002462:Cell Membrane D002474:Cell-Free System D017077:Culture Media, Conditioned D004789:Enzyme Activation D017353:Gene Deletion D006801:Humans D007476:Ionophores D051379:Mice D009154:Mutation D032383:Myocytes, Cardiac D010446:Peptide Fragments D043484:Proprotein Convertases D054730:Protein Interaction Domains and Motifs D011498:Protein Precursors D011993:Recombinant Fusion Proteins D055571:Secretory Pathway D012697:Serine Endopeptidases D012995:Solubility
Corin PCSK6 Transmembrane domain Type II transmembrane serine protease Zymogen activation