The role of the C-terminal domain of PCSK9 and SEC24 isoforms in PCSK9 secretion

y Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a secretory protein that promotes low-density lipoprotein receptor (LDLR) degradation and thereby regulating plasma levels of LDL cholesterol. Previous studies have revealed the role of the C-terminal domain (CTD) of PCSK9 in its secretion, however, how CTD regulates PCSK9 secretion is not completely understood. Additionally, SEC24A, the cargo adaptor protein of the coat protein complex II, has been implicated in the secretion of mouse PCSK9. Here, we investigated how CTD and SEC24 regulated PCSK9 secretion in humans. We found that mutant PCSK9(1-528), in which amino acids from 529 to the end (amino acid 692) were deleted, was maturated and secreted from cells as effectively as the wild-type protein. On the other hand, lacking amino acids 454 to 692 in mutant PCSK9(1-453) significantly reduced its maturation and secretion, but to a lesser extent when compared to mutants PCSK9(1-446), PCSK9(1-445) and PCSK9(1-444), that all markedly impaired PCSK9 maturation. However, mutant PCSK9(1-444) virtually eliminated PCSK9 secretion while PCSK9(1-446) and PCSK9(1-445) could still be adequately detected in culture medium. Interestingly, mutation of Pro(445) to other amino acid residues considerably impaired the secretion of mutant PCSK9(1-445) but not the full-length protein. We also found that natural variants in CTD including S462P, S465L, E482G, R495Q and A522T impaired PCSK9 secretion. Further, the knockdown of SEC24A, SEC24B, SEC24C but not SEC24D reduced secretion of the full-length PCSK9 but not mutant PCSK9(1-446). Therefore, SEC24A, SEC24B, and SEC24C facilitate endogenous PCSK9 secretion from cultured human hepatocytes, that are most likely mediated by the CTD of PCSK9. Our studies also indicate that the CTD of PCSK9 may allosterically and independently modulate the stability of the hinge region. Collectively, these data revealed that the CTD of PCSK9 and the hinge region play a critical role in PCSK9 maturation and secretion.