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Familial hypercholesterolemia: genetic diagnosis











Figure 3 The LDLR promoter regulation. Notes: Low-density lipoprotein receptor gene (LDLR) 5promoter region of 300 bp is represented, numbered the A of the ATG codon as +1. The major regulatory regions are indicated with different colors: FP2 (from 280 to 268); FP1 (238 to 217); repeat (ReP) 1 (196 to 181); ReP 2 (161 to 146); ReP 3 (145 to 128); and TATA box (from 116 to 110 and 107 to 101). interaction between cis-element and trans-element at the proximal promoter drives high levels transcription when sterol level becomes deficient. The sterol regulatory element (SRE) binding protein transcription factor interacts with the SRE-1 of REP 2, whereas SP1 transcription factors interact with ReP 1 and ReP 3 to promoter high levels of LDLR gene transcription in response to low intracellular sterol concentrations. SP1 also involved with constitutive, basal-level expression of the LDLR gene. The TATA boxes recruit and direct the assembly of general transcription factors at the promoter.

The third domain of the LDLr is a region of 58 amino acids rich in threonine and serine residues that is encoded by exon 15. The function of this domain is unknown, but has been observed that this region serve as attachment sites for O-linked carbohydrate chains.1,66 This region shows minimal sequence conservation among six species analyzed and can be deleted without adverse effects on receptor function in cultured fibroblasts.66 It is thought that this domain plays a role in the stabilization of the receptor.1 Actually, 41 allelic variants within exon 15 are registered in LDLR databases.

The transmembrane domain that contains 22 hydrophobic amino acids is coded by exon 16 and the 5end of exon 17. This domain is essential for anchor the LDLr to the cell membrane. The cytoplasmic domain of the LDLr, that compromises 50 amino acid residues, is encoded by the remainder 3region of the exon 17 and the 5end of the exon 18.1 This domain contains two sequence signals for targeting the LDLr to the surface and for localizing the receptor in coated pits.67 This domain is the most conserved region of the LDLr, which is more than 86% identical among six species.1 Only a few allelic variants, 5.9% of total, have been identified within these domains.

Nowadays, over 1,000 mutations in LDLR have been described in FH patients along many populations (see http:// www.ucl.ac.uk/fh, http://www.umd.necker.fr). The naturally occurring LDLR can produce defects in transcription, posttranscription processes, translation, and posttranslation processes.59 FH mutations have been classified into five classes depending on the phenotypic behavior of mutant protein.43

Class 1 mutations are known as “null alleles.” These kinds of mutations are due to LDLR promoter deletion, by frameshift, nonsense, splicing mutations, or rearrangements in a way that messenger RNA (mRNA) is not produced; it generates an abnormal mRNA or normal in size but in a reduced concentration.68

Class 2 mutations are transport defective alleles which encode for proteins that do not have an adequate three- dimensional structure after being synthesized and keep blocked, complete or partially (2A and 2B, respectively) in transport process between endoplasmic reticulum and golgi apparatus. This defect is caused, normally, by missense mutations or small deletions in LDLR avoiding partial or completely the folding protein. These mutations are located within exons that encode ligand-binding domain and EGFP-like domain.69

Class 3 mutations are binding defective alleles which encode for LDLr that are synthesized and transported to cell surface but are not able to bind LDL particles. This is a heterogeneous group, because LDL binding activity goes from 2% to 30% of normal. This defect is due to rearrange- ments in repeat cysteine residues in binding ligand domain or repeat deletions in EGFP-like domain.70

Class 4 mutations are known as internalization-defective alleles which produce proteins that are not able to group into clathrin-coated pits; therefore, LDLr are not internalized (4A: only cytoplasmic domain is affected and 4B: also affected transmembrane domain).66

Finally, recycling-defective alleles are also named as class 5 mutations, such as missense mutations in EGFP-like domain, which encode for LDLr that are not able to release LDL particles in endosomes avoiding receptor return to cell surface.59

The heterogeneity observed in FH patients in relation to plasma LDLc levels and CHD has been suggested due to dif- ferences in the nature of the mutation in the LDLR and several studies have been published in support of this.39,40,71–73 Even HMGCoA reductase inhibitors may depend on the nature of the mutation in the LDLR gene.74,75 Recently, a study carried out by our group, with 436 Spanish FH patients with known LDLR mutations classified as null alleles or defective alleles, has demonstrated that patients with a molecular diagnosis of

The Application of Clinical Genetics 2010:3

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