Bip介导内质网应激诱导的tau蛋白过度磷酸化及伴侣蛋白Sil1的保护作用

发布时间：2018-04-26 21:02

本文选题：AD + 内质网应激　；参考：《华中科技大学》2012年博士论文

【摘要】：老年痴呆,又称为阿尔茨海默病(Alzheimer's Disease, AD)是一种进行性发展的神经退行性疾病,其临床表现为认知能力下降、记忆功能降低甚至丧失。随着老龄社会的到来,患阿尔茨海默病的病人会越来越多,给社会和家庭带来沉重的经济负担。AD病人脑中两大病理学特征是：神经元外由Aβ短肽形成的老年斑(Senile plaques,SP)的沉积；神经元内神经纤维缠结(Neurofibrillary tangles, NFTs)的聚集。神经纤维缠结是由过度磷酸化的tau蛋白构成。并且研究发现,由过度磷酸化的tau蛋白形成的NTFs的数量和AD病人的临床痴呆程度成正相关。内质网(Endoplasmic reticulum, ER)是真核细胞中最重要的细胞器之一,主要参与维持胞内钙稳态以及新合成的膜蛋白和分泌型蛋白质的翻译后处理过程。多种病理条件,如缺血、缺氧或中毒等刺激下,诱导细胞发生以伴侣蛋白表达及非折叠蛋白反应启动为特征的内质网应激(ER Stress)。有很多研究报道称ER应激参与了AD的发病过程,但具体机制有待深入研究。内质网分子伴侣(ER molecular chaperones) Bip (Immunoglobulin-binding protein)是内质网中最重要的分子伴侣之一,在内质网应激时表达增高,被认为是内质网应激时最重要的标志之一。Bip在AD病人的大脑皮质和海马中表达明显增高；本课题组以前研究发现持续光照会造成大鼠海马中Bip水平增高,tau蛋白过度磷酸化。Bip与tau过度磷酸化之间的关系及相关机制尚无报道。糖原合酶激酶-3β(Glycogen synthase kinase-3p, GSK-3p)是在AD发病过程中促使tau蛋白发生过度磷酸化的最重要的磷酸激酶之一,在AD病人脑中GSK-3β过度激活。本课题前期研究发现,ER应激在诱导Bip水平增高、tau蛋白过度磷酸化的同时,激活了GSK-3β;有研究发现,在突变Bip的转基因小鼠中,Bip失去正常功能作用的同时,GSK-3β活性受也到了抑制,上述研究提示Bip和GSK-3β也可能存在着某些必然的联系,但是具体机制不清楚。因此,本研究利用ER应激诱导剂毒胡萝卜素(Thapsigargin, TG)在整体或细胞水平诱导ER应激,发现GSK-3β激活、tau在Ser396、Ser198/199/202位点发生过度磷酸化；抑制GSK-3β激活,tau磷酸化水平显著降低。进一步利用RNA干扰技术下调Bip水平,TG诱导的tau过度磷酸化显著降低；ER应激及Bip过表达时,Bip可激活GSK-3β、促进GSK-3β与tau特异性结合而诱导tau过度磷酸化。核酸交换因子(Nucleotide exchange factor) Sil1是Bip的重要的共同伴侣分子。本研究发现在ER应激、Bip过表达的细胞中及AD模型小鼠(Tg2576小鼠)脑中Sil1表达明显降低,在细胞水平过表达Sil1通过抑制GSK-3β激活、减少Bip与GSK-3β、tau结合而显著改善TG和Bip过表达诱导的tau过度磷酸化；下调Sil1则Bip水平增高及tau发生过度磷酸化。此外,在培养的大鼠原代海马神经元过表达Bip或下调Sill水平,神经元的轴突生长明显抑制。本研究系统证明了Bip可通过激活GSK-3β、促进GSK-3β与tau特异性结合而诱导tau过度磷酸化,Sil1对ER应激及Bip过表达诱导的tau蛋白过度磷酸化具有保护作用。研究结果为进一步认识内质网相关的分子伴侣在AD发病中的作用及防治研究提供了依据。
[Abstract]:Alzheimer's disease, also known as Alzheimer's Disease (AD), is a progressive neurodegenerative disease. Its clinical manifestation is a decline in cognitive ability and a decrease in memory function. With the arrival of an aging society, patients with Alzheimer's disease are becoming more and more serious, bringing heavy economic burden to society and families. The two major pathological features in the brain of.AD patients are: the deposition of Senile plaques (SP) formed by A beta short peptide in neurons; aggregation of Neurofibrillary tangles, NFTs in neurons. The neurofibrillary tangles are constructed from the overphosphorylated tau protein. And the study found that the tau protein form is overphosphorylated. The number of NTFs was positively correlated with the degree of dementia in AD patients.
Endoplasmic reticulum (ER) is one of the most important organelles in eukaryotic cells. It is mainly involved in the maintenance of intracellular calcium homeostasis and the post-processing of newly synthesized membrane proteins and secretory proteins. A variety of pathological conditions, such as ischemia, hypoxia or poisoning, induce cells to occur with chaperone protein expression and unfolded eggs. A number of studies have reported that ER stress is involved in the pathogenesis of AD, but the specific mechanisms need to be studied in depth. The specific mechanisms need to be further studied. The endoplasmic reticulum molecular chaperone (ER molecular chaperones) Bip (Immunoglobulin-binding protein) is one of the most important molecular chaperones in the endoplasmic reticulum and is stressed by endoplasmic reticulum. The high expression of.Bip is considered to be one of the most important markers of endoplasmic reticulum stress. The expression of.Bip in the cerebral cortex and hippocampus of AD patients is significantly higher. The previous study found that the continuous light irradiation caused the increase of Bip in the hippocampus of rats, and the relationship between the excessive phosphorylation of tau protein and the phosphorylation of tau over the hippocampus and the related mechanisms. Report.
The glycogen synthase kinase -3 beta (Glycogen synthase kinase-3p, GSK-3p) is one of the most important phosphorylation of tau protein in the pathogenesis of AD. GSK-3 beta in the brain of AD patients is excessively activated. The previous study found that ER stress was induced by the increase of Bip level and the activation of tau protein over phosphorylation. It has been found that in transgenic mice with mutant Bip, Bip lost normal function and GSK-3 beta activity was also inhibited. The above studies suggest that Bip and GSK-3 beta may also have certain connections, but the specific mechanism is not clear.
Therefore, this study uses ER stress inducer (Thapsigargin, TG) to induce ER stress at the whole or cell level. It is found that GSK-3 beta activates, tau is over phosphorylated at Ser396, Ser198/199/202 site, and GSK-3 beta activation, and tau phosphorylation level is reduced significantly. Further downregulation of Bip level by RNA interference technique is used. Over expression of ER stress and Bip overexpression, Bip activates GSK-3 beta, promotes GSK-3 beta and tau specific binding and induces tau overphosphorylation. Nucleic acid exchange factor (Nucleotide exchange factor) Sil1 is the important co chaperone of Bip. In 576 mice, the expression of Sil1 in the brain was obviously reduced. The overexpression of Sil1 in the cell level by inhibiting the activation of GSK-3 beta, reducing the combination of Bip and GSK-3 beta, tau to significantly improve the tau overexpression induced by TG and Bip overexpression, and down regulated Sil1 Bip level and tau occur over phosphorylation. In addition, the primary hippocampal neurons in the cultured rat were overexpressed. Or down regulation of Sill, the axonal growth of neurons was significantly inhibited.
This study shows that Bip can induce the excessive phosphorylation of tau by activating GSK-3 beta and promoting the specific binding of GSK-3 beta to tau. Sil1 has protective effect on ER stress and the excessive phosphorylation of tau protein induced by Bip overexpression. The research results provide a further understanding of the role of endoplasmic reticulum related partners in the pathogenesis of AD and the prevention and treatment of AD. It's the basis.

【学位授予单位】：华中科技大学
【学位级别】：博士
【学位授予年份】：2012
【分类号】：R749.16

【共引文献】