The CFTR Corrector, VX-809 (Lumacaftor), Rescues ABCA4 Trafficking Mutants: a Potential Treatment for Stargardt Disease.
The CFTR Corrector, VX-809 (Lumacaftor), Rescues ABCA4 Trafficking Mutants: a Potential Treatment for Stargardt Disease.
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DOI：
10.33594/000000146
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发表时间：
2019-01-01
期刊：
影响因子：
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通讯作者：
Cebotaru, Liudmila
中科院分区：
文献类型：
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作者：
Liu, Qiangni;Sabirzhanova, Inna;Cebotaru, Liudmila
背景/目的：ABCA 4基因突变导致Stargardt黄斑变性，最终导致法律的失明。我们研究了两种常见的突变体，A1038 V（NBD 1）和G1961 E（NBD 2），目的是探索它们如何与细胞的质量控制机制相互作用。这项研究的目的是确定如何这些突变体可以rescued.METHODS：我们表达野生型和突变ABCA 4在HEK 293细胞中，并研究了运输和加工的突变的影响和纠正拯救他们的能力。结果：G1961 E对攻击体抑制剂tubacin和溶酶体抑制剂bafilomycin A敏感。这两种突变体都会导致热休克蛋白Hsp 27的减少。将表达突变体的HEK 293细胞与VX-809（一种FDA批准的用于治疗囊性纤维化的药物）一起孵育，使A1038 V和G1961 E的水平增加2至3倍。重要的是，VX-809增加了质膜上两种突变体的水平，表明运输已经恢复。将额外的Hsp 27转染到细胞中也增加了两种突变体的稳态水平。然而，在VX-809的组合中，Hsp 27的加入引起了蛋白质表达的急剧增加，特别是在G1961突变体中，增加了约5倍。结论：我们的研究结果提供了一个新的机制，为拯救ABCA 4贩运突变体的基础上恢复Hsp 27。我们的研究结果为Stargardt病的治疗提供了一条途径。
BACKGROUND/AIMS: Mutations in ABCA4 cause Stargardt macular degeneration, which invariably ends in legal blindness. We studied two common mutants, A1038V (in NBD1) and G1961E (in NBD2), with the purpose of exploring how they interact with the cell's quality control mechanism. The study was designed to determine how these mutants can be rescued.METHODS: We expressed wt and mutant ABCA4 in HEK293 cells and studied the effect of the mutations on trafficking and processing and the ability of correctors to rescue them. We used a combination of western blotting, confocal microscopy and surface biotinylation coupled with pulldown of plasma membrane proteins.RESULTS: G1961E is sensitive to inhibitors of the aggresome, tubacin and the lysosome, bafilomycin A. Both mutants cause a reduction in heat shock protein, Hsp27. Incubation of HEK293 cells expressing the mutants with VX-809, an FDA approved drug for the treatment of cystic fibrosis, increased the levels of A1038V and G1961E by 2- to 3-fold. Importantly, VX-809 increased the levels of both mutants at the plasma membrane suggesting that trafficking had been restored. Transfecting additional Hsp27 to the cells also increased the steady state levels of both mutants. However, in combination with VX-809 the addition of Hsp27 caused a dramatic increase in the protein expression particularly in the G1961 mutant which increased approximately 5-fold.CONCLUSION: Our results provide a new mechanism for the rescue of ABCA4 trafficking mutants based on the restoration of Hsp27. Our results provide a pathway for the treatment of Stargardt disease.