The deubiquitinating enzyme UBPy/USP8 interacts with TrkA and inhibits neuronal differentiation in PC12 cells
The deubiquitinating enzyme UBPy/USP8 interacts with TrkA and inhibits neuronal differentiation in PC12 cells
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DOI：
10.1016/j.yexcr.2015.01.019
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发表时间：
2015-04-10
影响因子：
3.7
通讯作者：
Martegani, Enzo
中科院分区：
文献类型：
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作者：
Ceriani, Michela;Amigoni, Loredana;Martegani, Enzo
受体酪氨酸激酶的原肌球蛋白相关激酶（Irk）家族控制突触功能、可塑性并维持分化、形态和神经元细胞存活。了解Trk受体下调和再循环是指出交感神经元和感觉神经元功能和存活的关键一步。来源于大鼠肾上腺髓质嗜铬细胞瘤的PC 12细胞已被广泛用作研究神经元分化的模型系统，因为它们响应于神经生长因子（NGF），表型发生显著变化，并获得交感神经元的许多特性。在这项研究中，我们证明了在PC 12细胞中，TrkA受体与去泛素化酶USP 8/UBPy以NGF依赖性方式相互作用，并且它在体内和体外被USP 8去泛素化。USP 8过表达阻断了NGF诱导的神经突生长，而催化失活突变体USP 8/UBPYC 748 A的过表达引起了细胞分化的显著增加。定位和生化实验已经指出，在NGF刺激后，USP 8和TrkA部分共定位于内体中。最后，我们研究了USP 8对TrkA周转所起的作用：使用针对USP 8的特异性siRNA，我们发现USP 8敲低增加了TrkA半衰期，这表明USP 8的去泛素化活性促进了TrkA降解。(C)2015 Elsevier Inc. All rights reserved.
The tropomyosin-related kinase (Irk) family of receptor tyrosine kinases controls synaptic function, plasticity and sustains differentiation, morphology, and neuronal cell survival. Understanding Trk receptors down-regulation and recycling is a crucial step to point out sympathetic and sensory neuron function and survival. PC12 cells derived from pheochromocytoma of the rat adrenal medulla have been widely used as a model system for studies of neuronal differentiation as they respond to nerve growth factor (NGF) with a dramatic change in phenotype and acquire a number of properties characteristic of sympathetic neurons. In this study we demonstrated that in PC12 cells the TrkA receptor interacts with the deubiquitinating enzyme USP8/UBPy in a NGF-dependent manner and that it is deubiquitinated in vivo and in vitro by USP8. USP8 overexpression blocked NGF-induced neurites outgrowth while the overexpression of the catalytically inactive mutant USP8/UBPYC748A caused a marked increase of cell differentiation. Localization and biochemical experiments have point out that USP8 and TrkA partially co-localize in endosomes after NGF stimulation. Finally we have studied the role played by USP8 on TrkA turnover: using specific siRNA for USP8 we found that USP8 knockdown increases TrkA half-life, suggesting that the deubiquitinating activity of USP8 promotes TrkA degradation. (C) 2015 Elsevier Inc. All rights reserved.