Department of Traditional Chinese Medicine, Shengli Clinical Medical College of Fujian Medical University, Fujian Provincial Hospital, Fuzhou, China.
Department of Traditional Chinese Medicine, Fujian Provincial Hospital, Fuzhou, China.
PLoS One. 2024 Oct 11;19(10):e0310547. doi: 10.1371/journal.pone.0310547. eCollection 2024.
Familial hypercholesterolemia (FH) is a common monogenic autosomal dominant disorder, primarily mainly caused by pathogenic mutations in the low-density lipoprotein receptor (LDLR) gene. Through phenotypic-genetic linkage analysis, two LDLR pathogenic mutations were identified in FH families: c.G1027A (p.Gly343Ser) and c.G1879A (p.Ala627Thr).
Whole exome sequencing was conducted on the proband with familial hypercholesterolemia to identify the target gene and screen for potential pathogenic mutations. The suspicious responsible mutation sites in 14 family members were analyzed using Sanger sequencing to assess genotype-phenotype correlations. Mutant and wild type plasmids were constructed and transfected into HEK293T cells to evaluate LDLR mRNA and protein expression. In parallel, bioinformatics tools were employed to predict structural and functional changes in the mutant LDLR.
Immunofluorescence analysis revealed no significant difference in the intracellular localization of the p.Gly343Ser mutation, whereas protein expression of the p.Ala627Thr mutation was decreased and predominantly localized in the cytoplasm. Western blotting has showed that protein expression levels of the mutant variants were markedly declined in both cell lysates and supernatants. Enzyme linked immunosorbent assay has demonstrated that LDLR protein levels in the supernatant of cell culture medium was not significant different from those of the wild-type group. However, LDLR protein levels in the cell lysate of both the Gly343Ser and Ala627Thr variants groups were significantly lower than those in the wild-type group. Bioinformatic predictions further suggested that these mutations may affect post-translational modifications of the protein, providing additional insight into the mechanisms underlying the observed reduction in protein expression.
In this study, we identified two heterozygous pathogenic variants in the LDLR gene, c.G1027A (p.Gly343Ser) and c.G1879A (p.Ala627Thr), in a family with familial hypercholesterolemia. We also conducted preliminary investigations into the mechanisms by which these mutations contribute to disease pathology.
家族性高胆固醇血症（FH）是一种常见的单基因常染色体显性遗传病，主要由低密度脂蛋白受体（LDLR）基因突变引起。通过表型-遗传连锁分析，在 FH 家系中发现了 LDLR 两种致病性突变：c.G1027A（p.Gly343Ser）和 c.G1879A（p.Ala627Thr）。
对家族性高胆固醇血症先证者进行全外显子组测序，以鉴定靶基因并筛选潜在的致病性突变。对 14 名家系成员的可疑致病变异位点进行 Sanger 测序分析，以评估基因型-表型相关性。构建突变型和野生型质粒并转染 HEK293T 细胞，以评估 LDLR mRNA 和蛋白表达。同时，采用生物信息学工具预测突变 LDLR 的结构和功能变化。
免疫荧光分析显示，p.Gly343Ser 突变的细胞内定位无明显差异，而 p.Ala627Thr 突变的蛋白表达减少且主要定位于细胞质。Western blot 显示突变型变体的蛋白表达水平在细胞裂解物和上清液中均显著降低。酶联免疫吸附试验显示，细胞培养上清液中 LDLR 蛋白水平与野生型组无显著差异。然而，Gly343Ser 和 Ala627Thr 变体组的细胞裂解物中 LDLR 蛋白水平明显低于野生型组。生物信息学预测进一步表明，这些突变可能影响蛋白的翻译后修饰，为观察到的蛋白表达减少的机制提供了更多的见解。
本研究在一个家族性高胆固醇血症家系中发现了 LDLR 基因的两个杂合致病性突变 c.G1027A（p.Gly343Ser）和 c.G1879A（p.Ala627Thr）。我们还对这些突变导致疾病发病机制的机制进行了初步研究。
