Normolipidemia and hypercholesterolemia in persons heterozygous for the same 1592+5G -> A splice site mutation in the low-density lipoprotein receptor gene

In the present study, we have characterized a unique splice donor G to A su bstitution in the moderately conserved + 5 position in intron 10 of the low -density lipoprotein (LDL) receptor gene. In two Danish families, carriers of the 1592 + 5G --> A mutation display a clinical phenotype ranging from h ealthy normocholesterolemic persons to classical heterozygous familial hype rcholesterolemia (FH) patients. Reverse transcription-polymerase chain reac tion (RT-PCR) of RNA from Epstein-Barr virus (EBV)-transformed lymphoblasts obtained from members of both families demonstrated abnormal splicing gene rating two aberrant mRNAs due to either alternative splicing and skipping o f exon 10 or activation of a cryptic splice site in intron 10 inserting 66 intronic base pairs. These abnormally spliced mRNAs were predicted to encod e two abnormal receptor proteins containing an in-frame deletion of 75 amin o acids and an insertion of 22 novel amino acids, respectively. Results obt ained by immunofluorescence staining, flow cytometry, and confocal microsco py of transfected Chang and COS-7 cells expressing normal and mutant LDL re ceptors were compatible with nearly complete retention of the mutant protei ns in the endoplasmic reticulum. Quantitative measurements of LDL, receptor mRNAs from EBV-transformed lymphoblasts, however, did not reveal any signi ficant differences in variant mRNA contents between mutation carriers in th e families that could be related to degree of hypercholesterolemia.