Department of Biochemistry and the Siebens-Drake Medical Research Institute, Schulich School of Medicine and Dentistry, The University of Western Ontario, London, Ontario N6A 5C1, Canada.
Int J Biol Sci. 2010 May 11;6(3):252-67. doi: 10.7150/ijbs.6.252.
NIP/DuoxA, originally cloned as a protein capable of binding to the cell fate determinant Numb in Drosophila, was recently identified as a modulator of reactive oxygen species (ROS) production in mammalian systems. Despite biochemical and cellular studies that link NIP/DuoxA to the generation of ROS through the dual oxidase (Duox) enzyme, the in vivo function of NIP/DuoxA has not been characterized to date. Here we report a genetic and functional characterization of nip in Drosophila melanogaster. We show that nip is essential for Drosophila development as nip null mutants die at the 1(st) larval instar. Expression of UAS-nip, but not UAS-Duox, rescued the lethality. To understand the function of nip beyond the early larval stage, we generated GAL4 inducible UAS-RNAi transgenes. da(G32)-GAL4 driven, ubiquitous RNAi-mediated silencing of nip led to profound abnormality in pre-adult development, crinkled wing and markedly reduced lifespan at 29 degrees C. Compared to wild type flies, da-GAL4 induced nip-RNAi transgenic flies exhibited significantly reduced ability to survive under oxidative stress and displayed impaired mitochondrial aconitase function. Our work provides in vivo evidence for a critical role for nip in the development and oxidative stress response in Drosophila.
NIP/DuoxA，最初被克隆为一种能够与果蝇中细胞命运决定因子 Numb 结合的蛋白质，最近被鉴定为哺乳动物系统中活性氧（ROS）产生的调节剂。尽管生化和细胞研究将 NIP/DuoxA 与通过双氧化酶（Duox）酶产生 ROS 联系起来，但迄今为止尚未对 NIP/DuoxA 的体内功能进行表征。在这里，我们报告了果蝇中 nip 的遗传和功能特征。我们表明，nip 对果蝇的发育是必不可少的，因为 nip 缺失突变体在第一幼虫期死亡。UAS-nip 的表达，但不是 UAS-Duox 的表达，挽救了致死性。为了了解 nip 在早期幼虫阶段之外的功能，我们生成了 GAL4 诱导的 UAS-RNAi 转基因。da(G32)-GAL4 驱动的普遍 RNAi 介导的 nip 沉默导致成年前发育严重异常，翅膀卷曲，在 29°C 下寿命明显缩短。与野生型果蝇相比，da-GAL4 诱导的 nip-RNAi 转基因果蝇在氧化应激下的存活能力显著降低，并且表现出受损的线粒体顺乌头酸酶功能。我们的工作为 nip 在果蝇发育和氧化应激反应中的关键作用提供了体内证据。
