Common Proprotein Convertase Subtilisin/Kexin Type 9 (PCSK9) Epitopes Mediate Multiple Routes for Internalization and Function

Abstract

Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a soluble protein that directs membrane-bound receptors to lysosomes for degradation. In the most studied example of this, PCSK9 binding leads to the degradation of low density lipoprotein receptor (LDLR), significantly affecting circulating LDL-C levels. The mechanism mediating this degradation, however, is not completely understood. We show here that LDLR facilitates PCSK9 interactions with amyloid precursor like protein 2 (APLP2) at neutral pH leading to PCSK9 internalization, although direct binding between PCSK9 and LDLR is not required. Moreover, binding to APLP2 or LDLR is independently sufficient for PCSK9 endocytosis in hepatocytes, while LDL can compete with APLP2 for PCSK9 binding to indirectly mediate PCSK9 endocytosis. Finally, we show that APLP2 and LDLR are also required for the degradation of another PCSK9 target, APOER2, necessitating a general role for LDLR and APLP2 in PCSK9 function. Together, these findings provide evidence that PCSK9 has at least two endocytic epitopes that are utilized by a variety of internalization mechanisms and clarifies how PCSK9 may direct proteins to lysosomes.

Proteolytic cleavage of antigen extends the durability of an anti-PCSK9 monoclonal antibody

“…Importantly, mAbs LY and AM in complex with PCSK9 bound to APLP2 in an ELISA-based assay and the addition of furin reduced the binding of LY but not AM (K. M. Schroeder, unpublished observations). Our preliminary fi ndings support the hypothesis proposed by DeVay and coworkers ( 34,35 ) that APLP2 mediates the CL of anti-PCSK9 antibodies that are bound to PCSK9. We are pursuing the hypothesis that LY avoids the APLP2-dependent, antigen-mediated CL pathway by permitting the proteolytic cleavage of PCSK9.…”

“…The fi nding of DeVay and coworkers ( 34,35 ) of most relevance to our work was that an anti-PCSK9 antibody was transported with PCSK9 to lysosomes by the APLP2-mediated pathway. Our preliminary studies confi rmed that PCSK9 binds via its C-terminal domain to APLP2 at acidic pH (K. M. Schroeder, unpublished observations).…”

“…We can propose a mechanism which links PCSK9 accumulation to the more rapid CL of the antibodies based on the studies in DeVay and coworkers ( 34,35 ). These authors demonstrated that an anti-PCSK9 antibody was delivered to lysosomes for degradation by a specifi c pathway involved in PCSK9 CL.…”

“…C: mAbs that block the furin cleavage of PCSK9 are especially susceptible to PCSK9-mediated CL. The PCSK9 CL pathway could be mediated by APLP2, which binds to the C-terminal domain of PCSK9 even in the presence of the catalytic domain mAbs ( 34,35 ). D: Following the proteolytic cleavage of PCSK9, LY avoids antigenmediated CL because the domains required for this CL pathway are found in the 52 kDa truncated form of PCSK9.…”

Amyloid Precursor-like Protein 2 and Sortilin Do Not Regulate the PCSK9 Convertase-mediated Low Density Lipoprotein Receptor Degradation but Interact with Each Other

“…This suggested that PCSK9 can divert LDLR to lysosomes from the luminal side of the membrane via its Cys-His-rich domain interaction with APLP2 (31). Recent evidence also showed that the LDLR forms a complex with APLP2 at the cell surface (32). This finding was not unprecedented, as APLP2 is involved in the transport of transmembrane proteins such as MHC class I molecules to lysosomes (33,34).…”