Functional analysis of new 3′ untranslated regions genetic variants in genes associated with genetic hypercholesterolemias

BACKGROUND: Familial hypercholesterolemia (FH) is the best-described autosomal dominant genetic hypercholesterolemia (GH). Mutations in candidate genes can explain a high proportion of FH cases, but for many, no causative mutations are detected (designed non-FG-GH), suggesting the existence of additional genetic variants associated with the disease. OBJECTIVE: We aimed to identify new single-nucleotide variants (SNVs) located at the 3' untranslated regions (3'UTRs) of the low-density lipoprotein receptor, low-density lipoprotein receptor-related protein-associated protein 1, ATP-binding cassette sub-family G member 5, and sterol regulatory element-binding protein 2 genes in non-FH-GH individuals and investigated whether the association of these SNVs with non-FH-GH could be explained by changes in the affinity of regulatory microRNAs (miRNA) targeting the sequences modified by the SNVs. METHODS: The study includes probands with non-FH-GH attending 2 lipid clinics in Spain. We performed functional analyses of selected variants using a luciferase reporter system. Through in silico target-prediction tools, we identified miRNAs, which binding to the 3'UTR could be affected by the presence of specific SNVs. We used analogs and inhibitors of these miRNAs to test this possibility. RESULTS: We identified 11 new SNVs showing significant association with non-FH-GH. We show that the presence of 4 of these SNVs leads to significant changes in the transcriptional levels of the reporter gene. Through mechanistic analysis, we identified 2 miRNAs (miR-27a and miR-133-3p) targeting the 3'UTR of sterol regulatory element-binding protein 2 and an additional miRNA (miR-92a) targeting the 3'UTR of low-density lipoprotein receptor-related protein-associated protein 1. CONCLUSION: Our findings reveal novel regulatory links between certain miRNAs and key genes regulating cholesterol homeostasis. They also highlight the potential of miRNAs as therapeutic targets for the treatment of FH. (C) 2017 National Lipid Association. All rights reserved.