Nek5 interacts with mitochondrial proteins and interferes negatively in mitochondrial mediated cell death and respiration
Nek5 interacts with mitochondrial proteins and interferes negatively in mitochondrial mediated cell death and respiration
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DOI：
10.1016/j.cellsig.2015.02.021
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发表时间：
2015-06-01
影响因子：
4.8
通讯作者：
Kobarg, Joerg
中科院分区：
文献类型：
--
作者：
Melo Hanchuk, Talita D.;Papa, Priscila Ferreira;Kobarg, Joerg
线粒体参与能量供应、信号传递、细胞死亡和细胞分化，并与多种人类疾病有关。NeKs(NIMA相关蛋白)代表哺乳动物蛋白激酶家族，在细胞周期进程中发挥重要作用，但最近还涉及到其他功能。通过酵母双杂交(Y2H)筛选Nek5相互作用伙伴，回收线粒体蛋白Cox11、MTX-2和BCLAF1。细胞凋亡实验显示稳定的hNek5表达对thapsigargin(2MU)处理后Hek293-T的细胞死亡有保护作用。在thapsigargin处理4h后，Nek5使细胞沉默以及表达“激酶死亡”形式的Nek5的细胞显示ROS形成增加。稳定表达hNek5后，线粒体呼吸链活性降低。另一方面，沉默hNek5的细胞表现出1.7倍的基础呼吸速率，特别是在从TMPD到细胞色素c和复合体II的电子转移步骤。综上所述，我们的数据首次表明了线粒体对Nek5的定位和功能及其参与细胞死亡和细胞呼吸调节的作用。HNek5在Hek293T细胞中的稳定表达可提高细胞存活率，降低细胞死亡率和耐药性，而shRNA缺失hNek5可克服肿瘤细胞的耐药性，并在体外诱导细胞凋亡。HNek5的稳定表达还抑制thapsigargin促进HEK293T细胞的凋亡和呼吸链复合体IV。(C)2015 Elsevier Inc.保留所有权利。
Mitochondria are involved in energy supply, signaling, cell death and cellular differentiation and have been implicated in several human diseases. Neks (NIMA-related kinases) represent a family of mammal protein kinases that play essential roles in cell-cycle progression, but other functions have recently been related. A yeast two-hybrid (Y2H) screen was performed to identify and characterize Nek5 interaction partners and the mitochondrial proteins Cox11, MTX-2 and BCLAF1 were retrieved. Apoptosis assay showed protective effects of stable hNek5 expression from Hek293-T's cell death after thapsigargin treatment (2 mu M). Nek5 silenced cells as well as cells expressing a "kinase dead" version of Nek5, displayed an increase in ROS formation after 4 h of thapsigargin treatment. Mitochondrial respiratory chain activity was found decreased upon stable hNek5expression. Cells silenced for hNek5 on the other hand presented 1.7 fold increased basal rates of respiration, especially at the electrons transfer steps from TMPD to cytochrome c and at the complex II. In conclusion, our data suggest for the first time mitochondrial localization and functions for Nek5 and its participation in cell death and cell respiration regulation. Stable expression of hNek5 in Hek293T cells resulted in enhanced cell viability, decreased cell death and drug resistance, while depletion of hNek5by shRNA overcame cancer cell drug resistance and induced apoptosis in vitro. Stable expression of hNek5 also inhibits thapsigargin promoted apoptosis and the respiratory chain complex IV in HEK293T cells. (C) 2015 Elsevier Inc. All rights reserved.