Plasma membrane CD81 complexes with PCSK9 and LDLR and its levels are reduced by PCSK9 [Lipids]

July 20th, 2015 by Le, Q.–T., Blanchet, M., Seidah, N. G., Labonte, P.

Proprotein convertase subtilisin/kexin type 9 (PCSK9) is an important factor in plasma cholesterol regulation through modulation of low-density lipoprotein receptor (LDLR) levels. Naturally occurring mutation can lead to hyper- or hypo-cholesterolemia in human. Recently, we reported that PCSK9 was also able to modulate CD81 in Huh7 cells. In the present study, several gain-of-function (GOF) and loss-of-function (LOF) as well as engineered mutants of PCSK9 were compared for their ability to modulate the cell surface expression of LDLR and CD81. While PCSK9 GOF D374Y enhances the degradation both receptors, D374H and D129N seem to only reduce LDLR levels. In contrast, mutations in the C-terminal hinge-CHRD segment primarily affect the PCSK9-induced CD81 degradation. Furthermore, when C-terminally fused to an ACE2-transmembrane anchor, the secretory N-terminal catalytic or hinge-CHRD domains of PCSK9 were able to reduce CD81 and LDLR levels. These data confirm that PCSK9 reduces CD81 levels via an intracellular pathway, as reported for LDLR. Using immunocytochemistry, a proximity ligation assay and coimmunoprecipitation, we found that the cell surface level of PCSK9 was enhanced upon overexpression of CD81, and that both PCSK9 and LDLR interact with this tetraspanin protein. Interestingly, using CHO-A7 cells lacking LDLR expression, we revealed that LDLR was not required for the degradation of CD81 by PCSK9, but its presence strengthened the PCSK9 effect.