BACKGROUND: Large deletions and duplications within the low-density lipoprotein receptor (LDLR) gene make up approximately 10% of LDLR pathogenic variants found in Czech patients with familial hypercholesterolemia. The goal of this study was to test the hypothesis that all probands with each rearrangement share identical breakpoints inherited from a common ancestor and to determine the role of Alu repetitive elements in the generation of these rearrangements. METHODS: The breakpoint sequence was determined by PCR amplification and Sanger sequencing. To confirm the breakpoint position, an NGS analysis was performed. Haplotype analysis of common LDLR variants was performed using PCR and Sanger sequencing. RESULTS: The breakpoints of 8 rearrangements within the LDLR gene were analysed, including the four most common LDLR rearrangements in the Czech population (number of probands ranging from 8 to 28), and four less common rearrangements (1-4 probands). Probands with a specific rearrangement shared identical breakpoint positions and haplotypes associated with the rearrangement, suggesting a shared origin from a common ancestor. All breakpoints except for one were located inside an Alu element. In 6 out of 8 breakpoints, there was high homology (≥ 70%) between the two Alu repeats in which the break occurred. CONCLUSIONS: The most common rearrangements of the LDLR gene in the Czech population likely arose from one mutational event. Alu elements likely played a role in the generation of the majority of rearrangements inside the LDLR gene.
- MeSH
- genová přestavba MeSH
- hyperlipoproteinemie typ II * genetika epidemiologie MeSH
- lidé MeSH
- mutace MeSH
- receptory LDL genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika MeSH
Background: Pathogenic variants in the low density lipoprotein receptor gene are associated with familial hypercholesterolemia. Some of these variants can result in incorrect folding of the LDLR protein, which is then accumulated inside the cell and cannot fulfill its function to internalize LDL particles. We analyzed the functional impact of 10 LDLR variants localized in the beta-propeller of epidermal growth factor precursor homology domain. The experimental part of the work was complemented by a structural analysis on the basis of 3D LDLR protein structure. Methods: T-Rex Chinese hamster ovary cells transfected with the human LDLR gene were used for live cell imaging microscopy, flow cytometry, and qRT-PCR analysis. Results: Our results showed that the analyzed LDLR protein variants can be divided into three groups. (1) The variants buried inside the 3D protein structure expressing proteins accumulated in the endoplasmic reticulum (ER) with no or reduced plasma membrane localization and LDL particle internalization, and associated with an increased gene expression of ER-resident chaperones. (2) The variants localized on the surface of 3D protein structure with slightly reduced LDLR plasma membrane localization and LDL particle internalization, and associated with no increased mRNA level of ER-resident chaperones. (3) The variants localized on the surface of the 3D protein structure but expressing proteins with cell responses similar to the group 1. Conclusion: All analyzed LDLR variants have been evaluated as pathogenic but with different effects on protein localization and function, and expression of genes associated with ER stress.
- Publikační typ
- časopisecké články MeSH
- Publikační typ
- abstrakt z konference MeSH
Autosomal dominant hypercholesterolemia (ADH), more known as familial hypercholesterolemia (FH), is a lipid metabolism disorder characterized by an elevation in low-density lipoprotein cholesterol (LDL-C) and increased risk for cardiovascular disease. In this study, we assessed a spectrum of mutations causing ADH in 3914 unrelated Czech patients with clinical diagnosis of hypercholesterolemia. Samples have been collected within the framework of the MedPed project running in the Czech Republic since 1998. So far we have found 432 patients (11.0 %) with the APOB gene mutation p.(Arg3527Gln) and 864 patients (22.1 %) with the LDLR gene mutation. In 864 probands carrying the LDLR gene mutation, 182 unique allelic variants were detected. We have identified 14 patients homozygous for mutations in the LDLR or APOB genes. We performed function analyses of p.(Leu15Pro) and p.(Gly20Arg) sequence variations.
- MeSH
- apolipoprotein B-100 genetika MeSH
- genetická variace genetika MeSH
- genetické pozadí * MeSH
- hyperlipoproteinemie typ II krev epidemiologie genetika MeSH
- LDL-cholesterol krev genetika MeSH
- lidé MeSH
- receptory LDL genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
- Geografické názvy
- Česká republika MeSH
- Publikační typ
- abstrakt z konference MeSH
- MeSH
- anticholesteremika aplikace a dávkování farmakokinetika farmakologie MeSH
- ektroforéza na škrobovém gelu metody MeSH
- hyperlipoproteinemie typ II * diagnóza farmakoterapie MeSH
- izoenzymy krev MeSH
- kreatinkinasa krev MeSH
- lidé MeSH
- statiny aplikace a dávkování farmakokinetika farmakologie škodlivé účinky MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- abstrakt z konference MeSH
- Publikační typ
- abstrakt z konference MeSH
BACKGROUND: Familial hypercholesterolemia (FH), a major risk for coronary heart disease, is predominantly associated with mutations in the genes encoding the low-density lipoprotein receptor (LDLR) and its ligand apolipoprotein B (APOB). RESULTS: In this study, we characterize the spectrum of mutations causing FH in 2239 Czech probands suspected to have FH. In this set, we found 265 patients (11.8%) with the APOB mutation p.(Arg3527Gln) and 535 patients (23.9%) with a LDLR mutation. In 535 probands carrying the LDLR mutation, 127 unique allelic variants were detected: 70.1% of these variants were DNA substitutions, 16.5% small DNA rearrangements, and 13.4% large DNA rearrangements. Fifty five variants were novel, not described in other FH populations. For lipid profile analyses, FH probands were divided into groups [patients with the LDLR mutation (LDLR+), with the APOB mutation (APOB+), and without a detected mutation (LDLR-/APOB-)], and each group into subgroups according to gender. The statistical analysis of lipid profiles was performed in 1722 probands adjusted for age in which biochemical data were obtained without FH treatment (480 LDLR+ patients, 222 APOB+ patients, and 1020 LDLR-/APOB- patients). Significant gradients in i) total cholesterol (LDLR+ patients > APOB+ patients = LDLR-/APOB- patients) ii) LDL cholesterol (LDLR+ patients > APOB+ patients = LDLR-/APOB- patients in men and LDLR+patients > APOB+ patients >LDLR-/APOB- patients in women), iii) triglycerides (LDLR-/APOB- patients > LDLR+ patients > APOB+ patients), and iv) HDL cholesterol (APOB+ patients > LDLR-/APOB- patients = LDLR+ patients) were shown. CONCLUSION: Our study presents a large set of Czech patients with FH diagnosis in which DNA diagnostics was performed and which allowed statistical analysis of clinical and biochemical data.
- MeSH
- apolipoproteiny B genetika MeSH
- biologické markery krev MeSH
- dítě MeSH
- dospělí MeSH
- fenotyp MeSH
- frekvence genu MeSH
- genetická predispozice k nemoci MeSH
- genetické asociační studie MeSH
- hodnocení rizik MeSH
- hyperlipoproteinemie typ II krev diagnóza epidemiologie genetika MeSH
- lidé středního věku MeSH
- lidé MeSH
- lineární modely MeSH
- lipidy krev MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mutace * MeSH
- mutační analýza DNA MeSH
- receptory LDL genetika MeSH
- rizikové faktory MeSH
- rozdělení chí kvadrát MeSH
- Check Tag
- dítě MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Česká republika MeSH
