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Identification and functional analysis of missense mutations in the lecithin cholesterol acyltransferase gene in a Chilean patient with hypoalphalipoproteinemia.

ABSTRACT: BACKGROUND:Lecithin-cholesterol acyltransferase (LCAT) is a plasma enzyme that esterifies cholesterol in high- and low-density lipoproteins (HDL and LDL). Mutations in LCAT gene causes familial LCAT deficiency, which is characterized by very low plasma HDL-cholesterol levels (Hypoalphalipoproteinemia), corneal opacity and anemia, among other lipid-related traits. Our aim is to evaluate clinical/biochemical features of a Chilean family with a proband showing clinical signs of familial LCAT deficiency, as well as to identify and assess the functional effects of LCAT mutations. METHODS:An adult female proband with hypoalphalipoproteinemia, corneal opacity and mild anemia, as well as her first-degree relatives, were recruited for clinical, biochemical, genetic, in-silico and in-vitro LCAT analysis. Sequencing of exons and intron-exon boundaries was performed to identify mutations. Site-directed mutagenesis was carried out to generate plasmids containing cDNA with wild type or mutant sequences. Such expression vectors were transfected to HEK-239?T cells to asses the effect of LCAT variants in expression, synthesis, secretion and enzyme activity. In-silico prediction analysis and molecular modeling was also used to evaluate the effect of LCAT variants. RESULTS:LCAT sequencing identified rare p.V333?M and p.M404?V missense mutations in compound heterozygous state in the proband, as well the common synonymous p.L363?L variant. LCAT protein was detected in proband's plasma, but with undetectable enzyme activity compared to control relatives. HEK-293?T transfected cells with vector expression plasmids containing either p.M404?V or p.V333?M cDNA showed detectable LCAT protein expression both in supernatants and lysates from cultured cells, but with much lower enzyme activity compared to cells transfected with the wild-type sequence. Bioinformatic analyses also supported a causal role of such rare variations in LCAT lack of function. Additionally, the proband carried the minor allele of the synonymous p.L363?L variant. However, this variant is unlikely to affect the clinical phenotype of the proband given its relatively high frequency in the Chilean population (4%) and its small putative effect on plasma HDL-cholesterol levels. CONCLUSION:Genetic, biochemical, in vitro and in silico analyses indicate that the rare mutations p.M404?V and p.V333?M in LCAT gene lead to suppression of LCAT enzyme activity and cause clinical features of familial LCAT deficiency.

SUBMITTER: Tobar HE

PROVIDER: S-EPMC6549291 | biostudies-literature | 2019 Jun

REPOSITORIES: biostudies-literature

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Identification and functional analysis of missense mutations in the lecithin cholesterol acyltransferase gene in a Chilean patient with hypoalphalipoproteinemia.

Tobar Hugo E HE Cataldo Luis R LR González Trinidad T Rodríguez Ricardo R Serrano Valentina V Arteaga Antonio A Álvarez-Mercado Ana A Lagos Carlos F CF Vicuña Lucas L Miranda José P JP Pereira Ana A Bravo Carolina C Aguilera Concepción M CM Eyheramendy Susana S Uauy Ricardo R Martínez Álvaro Á Gil Ángel Á Francone Omar O Rigotti Attilio A Santos José L JL

Lipids in health and disease 20190605 1

<h4>Background</h4>Lecithin-cholesterol acyltransferase (LCAT) is a plasma enzyme that esterifies cholesterol in high- and low-density lipoproteins (HDL and LDL). Mutations in LCAT gene causes familial LCAT deficiency, which is characterized by very low plasma HDL-cholesterol levels (Hypoalphalipoproteinemia), corneal opacity and anemia, among other lipid-related traits. Our aim is to evaluate clinical/biochemical features of a Chilean family with a proband showing clinical signs of familial LCA ...[more]

PMID: 31164121

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Project description:Lecithin:cholesterol acyltransferase (LCAT) has been shown to play a role in the depletion of lipid oxidation products, but this has so far not been studied in humans. In this study, we investigated processes and parameters relevant to lipid oxidation in carriers of functional LCAT mutations.In 4 carriers of 2 mutant LCAT alleles, 63 heterozygotes, and 63 family controls, we measured activities of LCAT, paraoxonase 1, and platelet-activating factor-acetylhydrolase; levels of lysophosphatidylcholine molecular species, arachidonic and linoleic acids, and their oxidized derivatives; immunodetectable oxidized phospholipids on apolipoprotein (apo) B-containing and apo(a)-containing lipoproteins; IgM and IgG autoantibodies to malondialdehyde-low-density lipoprotein and IgG and IgM apoB-immune complexes; and the antioxidant capacity of high-density lipoprotein (HDL). In individuals with LCAT mutations, plasma LCAT activity, HDL cholesterol, apoA-I, arachidonic acid, and its oxidized derivatives, oxidized phospholipids on apo(a)-containing lipoproteins, HDL-associated platelet-activating factor-acetylhydrolase activity, and the antioxidative capacity of HDL were gene-dose-dependently decreased. Oxidized phospholipids on apoB-containing lipoproteins was increased in heterozygotes (17%; P<0.001) but not in carriers of 2 defective LCAT alleles.Carriers of LCAT mutations present with significant reductions in LCAT activity, HDL cholesterol, apoA-I, platelet-activating factor-acetylhydrolase activity, and antioxidative potential of HDL, but this is not associated with parameters of increased lipid peroxidation; we did not observe significant changes in the oxidation products of arachidonic acid and linoleic acid, immunoreactive oxidized phospholipids on apo(a)-containing lipoproteins, and IgM and IgG autoantibodies against malondialdehyde-low-density lipoprotein. These data indicate that plasma LCAT activity, HDL-associated platelet-activating factor-acetylhydrolase activity, and HDL cholesterol may not influence the levels of plasma lipid oxidation products.

Dataset Information

Identification and functional analysis of missense mutations in the lecithin cholesterol acyltransferase gene in a Chilean patient with hypoalphalipoproteinemia.

Publications

Identification and functional analysis of missense mutations in the lecithin cholesterol acyltransferase gene in a Chilean patient with hypoalphalipoproteinemia.

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