FIGURE 4.

The gain-of-function mutant PCSK9-D374Y, but not WT PCSK9, decreases cell surface LDLR on adjacent cells. a, immunocytochemistry of cell surface LDLR (blue) in HepG2 transiently overexpressing a bicistronic vector (pIRES) encoding WT PCSK9 and EGFP (green) is shown. b–d, at 24 h after transfection, cells overexpressing a nonsecreted catalytically inactive PCSK9 (H226A), WT, or the gain-of-function mutant (D374Y) were sorted for EGFP and LDLR. Graphs were derived from FACS analyses (supplemental Fig. S1), and values were normalized to those of the H226A-expressing cells. Secreted human PCSK9 levels were measured using an ELISA (b). Error bars represent three separate experiments. Scale bar, 20 μm.