Low-density lipoprotein receptor activity in Epstein-Barr virus-transformed lymphocytes from heterozygotes for the D374Y mutation in the PCSK9 gene.
OBJECTIVE
Missense mutations in the proprotein convertase subtilisin/kexin type 9 (PCSK9) gene have been found to cause autosomal dominant hypercholesterolemia. The objective of this study was to investigate possible mechanisms by which mutation D374Y in the PCSK9 gene causes hypercholesterolemia.


METHODS
Binding and internalization of low-density lipoprotein LDL in Epstein-Barr virus (EBV)-transformed lymphocytes from D374Y heterozygotes were examined. The autocatalytic activity of the D374Y mutant was studied in transiently transfected HEK293 cells.


RESULTS
As determined by Western blot analysis of transiently transfected HEK293 cells, the autocatalytic activity of the D374Y mutant was approximately 95% of the wild-type. Levels of PCSK9 mRNA in EBV-transformed lymphocytes from D374Y heterozygotes and normal controls were similar and less than 1/1000 of the level in HepG2 cells. The amount of cell surface LDL receptors (LDLRs) in EBV-transformed lymphocytes from five D374Y heterozygotes was non-significantly increased by 17% compared with the amount in normal controls. LDLR-dependent binding and internalization of LDL in EBV-transformed lymphocytes from D374Y heterozygotes were non-significantly reduced by 11% and 12%, respectively, compared to the corresponding values in normal controls.


CONCLUSIONS
LDLR-mediated endocytosis of LDL is not reduced in EBV-transformed lymphocytes from D374Y heterozygotes. Because of the extremely low levels of PCSK9 mRNA in EBV-transformed lymphocytes, it is possible that the LDLR-dependent endocytosis of LDL could be more severely affected in hepatocytes from D374Y heterozygotes than in EBV-transformed lymphocytes.
Holla
Ø L
ØL
Medical Genetics Laboratory, Department of Medical Genetics, Rikshospitalet University Hospital, Oslo, Norway.
Cameron
J
J
Berge
K E
KE
Kulseth
M A
MA
Ranheim
T
T
Leren
T P
TP
D002461:Cell Line, Transformed; D004705:Endocytosis; D004854:Herpesvirus 4, Human; D006579:Heterozygote; D006801:Humans; D006938:Hyperlipoproteinemia Type II / Q000235:genetics*; D008077:Lipoproteins, LDL / Q000378:metabolism; D008214:Lymphocytes / Q000378:metabolism; D020125:Mutation, Missense / Q000235:genetics*; D000071449:Proprotein Convertase 9; D043484:Proprotein Convertases; D011973:Receptors, LDL / Q000235:genetics* / Q000378:metabolism; D012697:Serine Endopeptidases / Q000235:genetics* / Q000378:metabolism; D018709:Statistics, Nonparametric
D008077:Lipoproteins, LDL; D011973:Receptors, LDL; C472125:PCSK9 protein, human; D000071449:Proprotein Convertase 9; D043484:Proprotein Convertases; D012697:Serine Endopeptidases