Bamji-Mirza Michelle, Li Yan, Najem Dema, Liu Qing Yan, Walker Douglas, Lue Lih-Fen, Stupak Jacek, Chan Kenneth, Li Jianjun, Ghani Mahdi, Yang Ze, Rogaeva Ekaterina, Zhang Wandong
Department of Pathology and Laboratory Medicine, Faculty of Medicine, University of Ottawa, Ottawa, ON, Canada.
Human Health Therapeutics Portfolio, Life Sciences Division, National Research Council Canada, Ottawa, ON, Canada.
J Alzheimers Dis. 2016 Jun 13;53(3):875-92. doi: 10.3233/JAD-150965.
Alzheimer's disease (AD) is characterized by extracellular deposits of amyloid-β (Aβ) in the brain. ABCA7 is highly expressed in the brain and a susceptibility gene for late-onset AD (LOAD). The minor alleles at two ABCA7 single-nucleotide polymorphisms (SNPs), rs3764650 (T>G; intron13) and rs3752246 at a predicted myristoylation site (C>G; exon33; p.Gly1527Ala), are significantly associated with LOAD risk; however, the mechanism of this association is unknown. Functional consequences of both SNPs were examined in HEK293 and CHO cells stably expressing AβPPSwe. Luciferase reporter assays in HEK293 cells suggested that intron13 carrying rs3764650 major T-allele (int13-T) possessed promoter-enhancing capabilities. Co-transfection experiments with hABCA7 and int13-T resulted in significantly increased ABCA7 protein level relative to that with int13-G. Expression of hABCA7 carrying rs3752246 risk allele led to increases in secreted Aβ40 and Aβ42 and β-secretase activity in CHO- and HEK-AβPPSwe cells. Hydroxymyristic acid treatment of cells expressing hABCA7 carrying the rs3752246 major G allele resulted in increased β-secretase activity and levels of Aβ, suggesting that lack of myristoylation contributes to the observed cell-phenotypes. Molecular weight determination, by gel-electrophoresis and mass spectrometry, of hABCA7 peptides spanning position 1527 showed loss of post-translational modification in the risk-allele peptide. These results suggest that decreased expression, or impaired function, of ABCA7 may contribute to AD pathology.
阿尔茨海默病（AD）的特征是大脑中存在β淀粉样蛋白（Aβ）的细胞外沉积物。ABCA7在大脑中高度表达，是晚发性AD（LOAD）的一个易感基因。ABCA7的两个单核苷酸多态性（SNP），即rs3764650（T>G；内含子13）和预测的肉豆蔻酰化位点处的rs3752246（C>G；外显子33；p.Gly1527Ala）的次要等位基因，与LOAD风险显著相关；然而，这种关联的机制尚不清楚。在稳定表达AβPPSwe的HEK293和CHO细胞中检测了这两个SNP的功能后果。HEK293细胞中的荧光素酶报告基因检测表明，携带rs3764650主要T等位基因（int13-T）的内含子13具有启动子增强能力。与hABCA7和int13-T的共转染实验导致ABCA7蛋白水平相对于int13-G显著增加。携带rs3752246风险等位基因的hABCA7的表达导致CHO和HEK-AβPPSwe细胞中分泌的Aβ40和Aβ42以及β-分泌酶活性增加。用羟基肉豆蔻酸处理表达携带rs3752246主要G等位基因的hABCA7的细胞，导致β-分泌酶活性和Aβ水平增加，这表明肉豆蔻酰化的缺失导致了观察到的细胞表型。通过凝胶电泳和质谱法测定跨越第1527位的hABCA7肽的分子量，结果显示风险等位基因肽中翻译后修饰缺失。这些结果表明，ABCA7表达降低或功能受损可能导致AD病理。
