α1-antitrypsin nonsense mutation associated with a retained truncated protein and reduced mRNA
α1-antitrypsin nonsense mutation associated with a retained truncated protein and reduced mRNA
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DOI：
10.1006/mgme.1998.2680
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发表时间：
1998-04-01
影响因子：
3.8
通讯作者：
Brantly, M
中科院分区：
文献类型：
--
作者：
Lee, J;Novoradovskaya, N;Brantly, M
α（1）-抗胰蛋白酶（α 1AT）在肺中对中性粒细胞弹性蛋白酶介导的蛋白水解提供主要保护。α 1AT缺陷等位基因的遗传与肺气肿和肝病的风险增加相关。（α 1AT无效等位基因导致血清α 1AT完全缺失，并代表与α 1AT缺乏相关的等位基因连续体中的最终结果。导致血清α 1AT缺失的分子机制包括剪接异常、α 1AT编码外显子缺失和过早终止密码子。我们确定了一个意大利人与哮喘，肺气肿，血清α 1AT水平非常低。DNA测序证实了Mprocida缺陷等位基因和一个新的无效等位基因QOtrastevere（c654 G --> A，W194 Z），一个内含子2（IVS 2）剪接受体位点附近的无义突变。为了确定QOtrastevere的分子基础，特别是为了评估这种无义突变是否通过改变剪接干扰mRNA加工，我们使用了一种永久转染QOtrastevere或正常M α 1AT（有或无IVS 2）的中国仓鼠卵巢细胞系。北方印迹分析表明，正常的M结构，有或没有IVS 2，表达α 1 AT mRNA的大小相似。无义突变与α 1AT mRNA中度降低相关，无论是否存在IVS 2。无论剪接的机会如何，α 1 AT mRNA的减少支持了一个预防性易位模型，而不是核扫描模型作为α 1 AT mRNA减少的原因。免疫沉淀后的脉冲追踪研究表明，内质网保留的31 kDa QOtrastevere（α 1 AT）迅速降解。虽然mRNA含量适度降低，但截短形式的保留和快速细胞内降解是不存在分泌型α 1AT的主要机制。(C)北京：科学出版社.
alpha(1)-Antitrypsin (alpha 1AT) provides the major protection in the lung against neutrophil elastase-mediated proteolysis. Inheritance of alpha 1AT deficiency alleles is associated with an increased risk of emphysema and liver disease. (alpha 1AT null alleles cause the total absence of serum alpha 1AT and represent the ultimate in a continuum of alleles associated with alpha 1AT deficiency. The molecular mechanisms responsible for absence of serum alpha 1AT include splicing abnormalities, deletion of alpha 1AT coding exons, and premature stop codons. We identified an Italian individual with asthma, emphysema, and a very low level of serum alpha 1AT. DNA sequencing demonstrated the Mprocida deficiency allele and a novel null allele, QOtrastevere (c654 G --> A, W194Z), a nonsense mutation near the intron 2 (IVS2) splice acceptor site. To determine the molecular basis of QOtrastevere and specifically to evaluate whether this nonsense mutation interfered with mRNA processing by altered splicing, we used a Chinese hamster ovary cell line permanently transfected with QOtrastevere or normal M alpha 1AT with and without IVS2. Northern blot analysis demonstrated that the normal M construct, with or without IVS2, expressed alpha 1AT mRNA of a similar size. The nonsense mutation was associated with moderately reduced alpha 1AT mRNA regardless of the presence or absence of IVS2. Reduction in alpha 1AT mRNA regardless of the opportunity for splicing supports a translational-translocation model as the cause of reduced (alpha 1AT mRNA rather than the nuclear scanning model. Pulse-chase studies followed by immunoprecipitation demonstrated an endoplasmic reticulum-retained 31 kDa QOtrastevere (alpha 1AT, which was rapidly degraded. Although mRNA content was moderately reduced, retention and rapid intracellular degradation of the truncated form are the major mechanisms for the absence of secreted alpha 1AT. (C) 1998 Academic Press.