A278C mutation of dihydropteridine reductase decreases autophagy via mTOR signaling
A278C mutation of dihydropteridine reductase decreases autophagy via mTOR signaling
复制标题
二氢蝶啶还原酶的 A278C 突变通过 mTOR 信号传导降低自噬
DOI：
10.1093/abbs/gmx061
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发表时间：
2017-08-01
影响因子：
3.7
通讯作者：
Gu, Yanting
中科院分区：
文献类型：
--
作者：
Si, Qin;Sun, Sifan;Gu, Yanting
二氢蝶啶还原酶（Dihydropteridine reductase，QDPR）在BH 4的再循环过程中起重要作用，与氧化应激密切相关。我们之前曾报道过，人肾脏HEK 293 T细胞中QDPR的过表达可以显着保护细胞免受氧化应激，而这些有益作用被A278 C突变所消除。为了研究野生型和突变型QDPR对HEK 293 T细胞自噬的影响及其机制，我们构建了野生型和突变型QDPR表达质粒，并将其转染HEK 293 T细胞。3 d后收集细胞，观察融合蛋白的表达及细胞内活性氧（ROS）的产生。Western blot分析mTOR和核糖体蛋白S6激酶B1（S6 K1）信号通路的变化以及LC-I、LC-II、Bcl-1、Bcl-2、p62和p53的表达。结果表明，外源性野生型QDPR显著降低了mTOR的表达，并降低了mTOR和S6 K1的磷酸化。QDPR突变抑制了mTOR的调节，提示QDPR可能通过抑制mTOR信号通路而对自噬起正性调节作用。野生型QDPR组p62、LC 3-II和Beclin 1的表达明显增强，突变后均逆转。此外，QDPR的突变改变了Beclin 2上QDPR的上调。因此可以得出结论，QDPR似乎在增强自噬中起重要作用，并且其突变有助于自噬的失调。
Dihydropteridine reductase (QDPR) plays an important role in the recycling of BH4 and is closely related to oxidative stress. We have previously reported that the overexpression of QDPR in human kidney HEK293T cells significantly protected against oxidative stress, and these beneficial effects were abolished by A278C mutation. To evaluate the effect of wild-type and mutant QDPR on autophagy and its mechanism in HEK293T cells, we constructed the wild-type and mutant QDPR expression plasmids and transfected them into HEK293T cells. Three days later, cells were collected to observe the expression of fusion protein and the intracellular production of reactive oxygen species (ROS). Western blot analysis was employed to evaluate the change of mTOR and ribosomal protein S6 kinase B1 (S6K1) signaling and the expression of LC-I, LC-II, Bcl-1, Bcl-2, p62, and p53. The results showed that the exogenous wild-type QDPR significantly decreased the expression of mTOR and phosphorylation of the mTOR and S6K1. Mutation of QDPR inhibited the regulation of mTOR, suggesting that QDPR is a positive regulator of autophagy via suppressing mTOR signaling. The expressions of p62, LC3-II and Beclin 1 were dramatically enhanced in wild-type QDPR group, which were reversed after QDPR mutation. Additionally, mutation of QDPR altered the upregulation of QDPR on Beclin 2. It is therefore concluded that QDPR appears to play an important role in enhancing autophagy, and its mutation contributes to dysregulation of autophagy.