山茱萸环烯醚萜苷对蛋白磷酸酶2A催化亚基C磷酸化的调节机制

doi:10.3969/j.issn.1006-7795.2016.06.012

首都医科大学学报 ›› 2016, Vol. 37 ›› Issue (6): 777-783.doi: 10.3969/j.issn.1006-7795.2016.06.012

• 神经系统疾病的基础研究 • 上一篇下一篇

山茱萸环烯醚萜苷对蛋白磷酸酶2A催化亚基C磷酸化的调节机制

李雪莲^1,2,3,4,5, 杨翠翠^2,3,4,5, 张兰^2,3,4,5, 石京山¹

1. 遵义医学院基础药理省部共建教育部重点实验室, 贵州遵义 563000;
2. 首都医科大学宣武医院药物研究室, 北京 100053;
3. 北京脑重大疾病研究院, 北京 100053;
4. 北京市神经药物工程研究中心, 北京 100053;
5. 神经变性病教育部重点实验室, 北京 100053

收稿日期:2016-10-14 出版日期:2016-12-21 发布日期:2016-12-16
通讯作者: 张兰, 石京山 E-mail:lanizhg@hotmail.com;shijs@zmc.edu.cn
基金资助:
国家自然科学基金（81473373），北京市自然科学基金（7132110），“重大新药创制”科技重大专项（2015ZX09101016001），北京市卫生系统高层次卫生技术人才（2014-2-014），北京市新世纪百千万人才工程（008-0014），北京市教委新医药学科群（XK100270569）

Regulatory mechanism of Cornel iridoid glycoside on protein phosphatase 2A catalytic subunit C phosphorylation

Li Xuelian^1,2,3,4,5, Yang Cuicui^2,3,4,5, Zhang Lan^2,3,4,5, Shi Jingshan¹

1. Key Laboratory for Basic Pharmacology of Ministry of Education, Zunyi Medical College, Zunyi 563000, Guizhou Province, China;
2. Department of Pharmacology, Xuanwu Hospital, Capital Medical University, Beijing 100053, China;
3. Beijing Institute for Brain Disorders, Beijing 100053, China;
4. Beijing Engineering Research Center for Nervous System Drugs, Beijing 100053, China;
5. Key Laboratory for Neurodegenerative Diseases of Ministry of Education, Beijing 100053, China

Received:2016-10-14 Online:2016-12-21 Published:2016-12-16
Supported by:
This study was supported by National Natural Science Foundation of China(81473373),Natural Science Foundation of Beijing(7132110),Major Project for Essential Drug Research and Development(2015ZX09101016001),Beijing Health and Technical High-level Personal Plan(2014-2-014),Beijing New Century Talented Person Project(008-0014),New Medical Disciplines Project of Beijing Education Committee(XK100270569).

摘要/Abstract

摘要： 目的探讨山茱萸环烯醚萜苷（Cornel iridoid glycoside，CIG）上调蛋白磷酸酶2A（protein phosphatase 2A，PP2A），PP2A活性抑制tau蛋白磷酸化的作用机制。方法 1确定最佳转染条件：将Src质粒DNA（0.2、0.4、0.6、0.8 μg）瞬时转染入小鼠神经瘤母细胞（Neuro-2A cell，N2a细胞），观察不同量Src对PP2A催化亚基C磷酸化和tau蛋白磷酸化的影响；2将0.6 μg Src质粒DNA转染入N2a细胞，24 h后加入CIG（50、100、200 μg/mL）共同孵育24 h，观察CIG对Src、蛋白酪氨酸磷酸酯酶1B（protein tyrosine phosphatase 1B，PTP1B）、p-PP2Ac及tau蛋白磷酸化的作用。结果 1转染Src（0.2、0.4、0.6 μg）质粒DNA到N2a细胞，Src蛋白表达明显增加，p-PP2Ac水平上调，PP2Ac蛋白总量表达无变化，tau蛋白在Ser 199/202、Ser 396位点磷酸化显著增加；转染0.8 μg Src质粒DNA到N2a细胞与转染0.6 μg Src相比，p-PP2Ac表达下降，PP2Ac蛋白总量表达无变化，tau蛋白在Ser 199/202、Ser 396位点磷酸化降低。2转染0.6 μg Src质粒DNA到N2a细胞，Src蛋白表达增加，p-PP2Ac表达增加，tau蛋白在Ser 199/202、Thr 205、Thr 217以及Ser 396位点的磷酸化明显增加；CIG对Src蛋白表达无影响，能够抑制p-PP2Ac的表达，抑制tau蛋白在Ser 199/202、Thr 205、Thr 217以及Ser 396位点的磷酸化。此外，CIG能上调PTP1B蛋白表达。结论 CIG对Src蛋白无明显调节作用，但能通过增加PTP1B的表达，降低PP2A催化亚基C磷酸化，从而升高PP2A活性，进一步降低tau的过度磷酸化。CIG对tau蛋白过度磷酸化的抑制作用，将会给阿尔茨海默病的治疗带来广阔的应用前景。

关键词: 山茱萸环稀醚萜苷, 蛋白酪氨酸激酶Src, 蛋白磷酸酶2A, 蛋白酪氨酸磷酸酯酶1B, tau蛋白, 阿尔茨海默病

Abstract: Objective To investigate the mechanism of Cornel iridoid glycoside (CIG) inhibiting tau phosphorylation by up-regulating protein phosphatase 2A (PP2A) activity. Methods ① To determine optimal transfection conditions:transfecting Src plasmid DNA (0.2, 0.4, 0.6, 0.8 μg) into mouse neuro-2A cell (N2a cells) was performed to observe the effects of different quantity Src on phosphorylation of PP2A catalytic subunit C and tau phosphorylation. ② After transfecting 0.6 μg Src plasmid DNA into N2a cells 24 hours,the cells were incubated with CIG (50, 100, 200 μg/mL) 24h,then the effects of CIG on Src, PTP1B, p-PP2Ac and tau phosphorylation were observed. Results ① Expression of Src protein was significantly increased, the expression of p-PP2Ac was up-regulated and the expression of PP2A did not change when Src (0.2, 0.4, 0.6 μg) plasmid transfected into N2a cells, and the tau phosphorylation at the sites of Ser 199/202, Ser 396 increased significantly; In the N2a cells transfected with 0.8 μg Src, the expression of p-PP2Ac was increased apparently, the expression of PP2A was not changed, and the phosphorylation of tau at Ser 199/202 and Ser 396 sites was decreased. ② In the N2a cells transfected with 0.6 μg Src, the expression of Src was significantly increased, the expression of p-PP2Ac was significantly increased, and the tau phosphorylation at the sites of Ser 199/202, Thr 205, Thr 217 and Ser 396 sites increased significantly; CIG could inhibit the expression of p-PP2Ac, the expression of tau phosphorylation at the sites of Ser 199/202, Thr 205, Thr 217, and Ser 396. In addition, CIG can up-regulate PTP1B protein expression. Conclusion CIG had no obvious regulation effect on Src, but it could decrease the phosphorylation of PP2A catalytic subunit C by increasing the expression of PTP1B, and then increases the activity of PP2A, and further reduces the level of tau hyperphosphorylation. The inhibition of CIG on tau hyperphosphorylation, will bring broad application prospects on the treatment of AD.

Key words: Cornel iridoid glycoside, protein tyrosine kinase Src, protein phosphatase 2A, protein tyrosine phosphatase 1B, tau protein, Alzheimer's disease

中图分类号:

李雪莲, 杨翠翠, 张兰, 石京山. 山茱萸环烯醚萜苷对蛋白磷酸酶2A催化亚基C磷酸化的调节机制[J]. 首都医科大学学报, 2016, 37(6): 777-783.

Li Xuelian, Yang Cuicui, Zhang Lan, Shi Jingshan. Regulatory mechanism of Cornel iridoid glycoside on protein phosphatase 2A catalytic subunit C phosphorylation[J]. Journal of Capital Medical University, 2016, 37(6): 777-783.

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山茱萸环烯醚萜苷对蛋白磷酸酶2A催化亚基C磷酸化的调节机制

Regulatory mechanism of Cornel iridoid glycoside on protein phosphatase 2A catalytic subunit C phosphorylation

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