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

Autor: Tobar HE; Department of Nutrition, Diabetes and Metabolism, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile., Cataldo LR; Department of Nutrition, Diabetes and Metabolism, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile., González T; Department of Nutrition, Diabetes and Metabolism, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile., Rodríguez R; Department of Nutrition, Diabetes and Metabolism, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile., Serrano V; Department of Nutrition, Diabetes and Metabolism, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile., Arteaga A; Department of Nutrition, Diabetes and Metabolism, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile., Álvarez-Mercado A; INYTA, University of Granada, Granada, Spain., Lagos CF; Facultad de Medicina y Ciencia, Universidad San Sebastián, Campus Los Leones, Santiago, Chile., Vicuña L; Departamento de Estadísticas, Facultad de Matemáticas, Pontificia Universidad Católica de Chile, Santiago, Chile., Miranda JP; Department of Nutrition, Diabetes and Metabolism, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile., Pereira A; INTA, Universidad de Chile, Santiago, Chile., Bravo C; Department of Nutrition, Diabetes and Metabolism, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile., Aguilera CM; Department of Biochemistry and Molecular Biology II, Institute of Nutrition and Food Technology 'José Mataix', Center of Biomedical Research, University of Granada, Granada, Spain., Eyheramendy S; Departamento de Estadísticas, Facultad de Matemáticas, Pontificia Universidad Católica de Chile, Santiago, Chile., Uauy R; INTA, Universidad de Chile, Santiago, Chile.; División de Pediatría, Escuela de Medicina, Pontificia Universidad Católica de Chile, Santiago, Chile., Martínez Á; Fundación Ciencia & Vida, Santiago, Chile., Gil Á; INYTA, University of Granada, Granada, Spain., Francone O; Pfizer Global Research and Development, San Diego, USA., Rigotti A; Department of Nutrition, Diabetes and Metabolism, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile.; Centro de Nutrición Molecular y Enfermedades Crónicas, Escuela de Medicina, Pontificia Universidad Católica de Chile, Santiago, Chile., Santos JL; Department of Nutrition, Diabetes and Metabolism, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile. jsantosm@uc.cl.
Jazyk: angličtina
Zdroj: Lipids in health and disease [Lipids Health Dis] 2019 Jun 05; Vol. 18 (1), pp. 132. Date of Electronic Publication: 2019 Jun 05.
DOI: 10.1186/s12944-019-1045-0
Abstrakt: 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.
Databáze: MEDLINE
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