Nitric oxide scavenging causes remodeling of the endoplasmic reticulum, Golgi apparatus and mitochondria in pulmonary arterial endothelial cells.
Nitric oxide scavenging causes remodeling of the endoplasmic reticulum, Golgi apparatus and mitochondria in pulmonary arterial endothelial cells.
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DOI：
10.1016/j.niox.2013.06.005
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发表时间：
2013-09-01
期刊：
影响因子：
--
通讯作者：
Sehgal PB
中科院分区：
文献类型：
--
作者：
Lee JE;Yuan H;Liang FX;Sehgal PB
内皮细胞细胞质细胞器的结构和功能对组成型细胞内一氧化氮（NO）的依赖在很大程度上仍未被探索。我们之前报道了暴露于NO清除剂或sirna介导的eNOS敲低后的细胞中高尔基体的断裂。其他研究报告称，NO供体增加了线粒体裂变。在功能上，我们之前报道了暴露于NO清除剂2-（4-羧基苯基）-4,4,5,5-四甲基氨基唑啉-1-氧-3-氧化物（c-PTIO）的牛肺动脉内皮细胞（PAECs）表现出分泌可溶性蛋白延长的促分泌表型。在本研究中，我们研究了NO清除是否导致内质网（ER）的重塑。活细胞DAF-2DA成像证实细胞内NO与BODIPY C5-神经酰胺标记的高尔基体相关。未经治疗的人paec表现为外周管网状内质网的模式，并伴有核旁内质网片的积累。通过内质网结构蛋白reticulon-4b/Nogo-B和内质网驻留GTPase atlastin-3的免疫荧光检测、RTN4-GFP和KDEL-mCherry的活细胞荧光检测以及电镜检测，暴露于c-PTIO的细胞显示内质网片显著增加。这些内质网变化被NO供体二乙胺NONOate抑制，也由L-NAME产生，但不由D-NAME或8-br-cGMP产生。内质网重塑伴随着高尔基体断裂和线粒体纤原性和功能的增加（四甲基罗丹明，TMRE的摄取）。尽管高尔基体断裂，但功能性内质网/高尔基体运输单元被保留下来，这可以从Sec31A内质网出口位点邻近分散的高尔基体元件的积累和可溶性货物分泌增加1.8倍看出。Western blotting和免疫计划数据显示，在c-PTIO暴露后，RTN4b的泛素化程度越来越高，尤其是在蛋白酶体抑制剂MG132存在的情况下。目前的数据完成了三个紧密并列的细胞器-高尔基体，内质网和线粒体-的结构完整性依赖于一氧化氮的显著见解。
The dependence of the structure and function of cytoplasmic organelles in endothelial cells on constitutively produced intracellular nitric oxide (NO) remains largely unexplored. We previously reported fragmentation of the Golgi apparatus in cells exposed to NO scavengers or after siRNA-mediated knockdown of eNOS. Others have reported increased mitochondrial fission in response to an NO donor. Functionally, we previously reported that bovine pulmonary arterial endothelial cells (PAECs) exposed to the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5- tetramethylimidazoline-1-oxyl-3-oxide (c-PTIO) developed a prosecretory phenotype characterized by prolonged secretion of soluble proteins. In the present study, we investigated whether NO scavenging led to remodeling of the endoplasmic reticulum (ER). Live-cell DAF-2DA imaging confirmed the presence of intracellular NO in association with the BODIPY C5- ceramide-labelled Golgi apparatus. Untreated human PAECs displayed a pattern of peripheral tubulo-reticular ER with a juxtanuclear accumulation of ER sheets. Cells exposed to c-PTIO showed a dramatic increase in ER sheets as assayed using immunofluoresence for the ER structural protein reticulon-4b/Nogo-B and the ER-resident GTPase atlastin-3, live-cell fluorescence assays using RTN4-GFP and KDEL-mCherry, and electron microscopy methods. These ER changes were inhibited by the NO donor diethylamine NONOate, and also produced by L-NAME, but not D-NAME or 8-br-cGMP. This ER remodeling was accompanied by Golgi fragmentation and increased fibrillarity and function of mitochondria (uptake of tetramethyl- rhodamine, TMRE). Despite Golgi fragmentation the functional ER/Golgi trafficking unit was preserved as seen by the accumulation of Sec31A ER exit sites adjacent to the dispersed Golgi elements and a 1.8-fold increase in secretion of soluble cargo. Western blotting and immunopanning data showed that RTN4b was increasingly ubiquitinated following c-PTIO exposure, especially in the presence of the proteasomal inhibitor MG132. The present data complete the remarkable insight that the structural integrity of three closely juxtaposed cytoplasmic organelles - Golgi apparatus, endoplasmic reticulum and mitochondria -is dependent on nitric oxide.
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DOI：
10.1111/j.1600-0854.2010.01134.x
发表时间：
2011-01
期刊：
Traffic (Copenhagen, Denmark)
影响因子：
--
作者：
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DOI：
10.1152/ajplung.00086.2009
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2009-10-01
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2013-02-19
影响因子：
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作者：
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影响因子：
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DOI：
10.1124/jpet.108.144113
发表时间：
2009-04-01
影响因子：
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作者：
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