RTN3调节BACE1轴突转运的机制研究

发布时间：2019-05-12 00:54

【摘要】：阿茨海默症(Alzheimer's disease, AD)的病理特征之一是患者脑部的老年斑(Senile plaque),其中包含淀粉样蛋白斑、活化的星形胶质细胞、激活的小胶质细胞和大量营养不良的神经元。淀粉样蛋白Aβ的沉积形成淀粉样蛋白斑,这种肽段是由其前体蛋白(amyloid precursor protein, APP)经过β-和γ-secretase两步依次剪切而成。BACE1是唯一在β-位点切割APP的蛋白酶,它起始了Aβ的生成。增加BACE1的活性即会引起Aβ产量的增加,并最终导致AD病人脑中淀粉样蛋白的沉积。因此,在AD患者中BACE1的活性为何会增加,又如何增加,一直是近年来比较热门的研究课题。APP主要在神经元中表达,而人脑中Aβ的沉积与其轴突释放高度相关。所以我们进一步探讨了BACE1的突触定位是如何受到调节的。Reticulon3(RTN3)与BACE1相互作用,抑制其酶活性和对APP的剪切。这种调节功能的机制可能是1)：BACE1酶活性的发挥需要偏酸性的环境,而RTN3与BACE1相互作用后使得后者被滞留在内质网中,无法进去pH相对偏低的细胞器如早期内吞体,从而不能发挥其最大酶活性；2)：RTN3与BACE1的相互作用本身在空间阻碍了BACE1与APP的结合。在本研究中,我们对RNT3是否调节BACE1的突触定位进行研究,并且进一步明确其发生机制。所以本研究的主要目的是：研究在神经元中,BACE1在神经元中的定位和转运方式。更重要的是,了解RTN3对于BACE1的轴突转运和APP剪切的影响。我们的结果显示BACE1不仅分布于晚期高尔基体、转运高尔基体和早期内吞体,还存在于轴突和树突中。我们发现BACE1与突触前蛋白Synaptophysin存在共定位,提示BACE1定位于突触。突触位置的Ap过度释放与淀粉样蛋白沉积的增加有关,所以我们还探讨了BACE1在突触位置的定位是否受到其负向调控蛋白reticulon3(RTN3)的调控。我们发现在原代培养神经元中同时过表达RTN3与BACE1使得BACE1与Synaptophysin和PSD95的共定位下降,且这种下降是由于BACE1的顺式轴突转运被RTN3的过表达抑制。我们的结果说明RTN3的水平可以影响BACE1在轴突中的转运,且调控BACE1的轴突转运可以减少Aβ的轴突末端释放和淀粉样蛋白沉积,因此或可成为阿茨海默症的一条治疗途径。
[Abstract]:One of the pathological features of Alzheimer's disease (Alzheimer's disease, AD) is the Alzheimer's disease (Senile plaque), which contains amyloid plaques, activated astrocytes, activated microglia, and a large number of malnourished neurons. The deposition of amyloid protein A 尾 forms amyloid plaques, which are formed by the cleavage of the precursor protein (amyloid precursor protein, APP) through 尾-and 纬-secretase. BACE1 is the only protease that cleaves APP at the 尾-site. It initiated the formation of A 尾. Increasing the activity of BACE1 can increase the production of A 尾 and eventually lead to the deposition of starch protein in the brain of AD patients. Therefore, why the activity of BACE1 increases and how to increase in AD patients has been a hot research topic in recent years. App is mainly expressed in neurons, and the deposition of A 尾 in human brain is highly related to its axon release. Therefore, we further discussed how the synaptic localization of BACE1 is regulated. Reticulon3 (RTN3) interacts with BACE1 to inhibit its enzyme activity and shear to APP. The mechanism of this regulatory function may be 1): the activity of BACE1 requires an acidic environment, and the interaction between RTN3 and BACE1 makes the latter trapped in the endoplasmic reticulum and cannot enter organelles with relatively low pH, such as early endocytosis. Therefore, it can not give full play to its maximum enzyme activity. 2): the interaction between RTN3 and BACE1 hinders the combination of BACE1 and APP in space. In this study, we studied whether RNT3 regulated the synaptic localization of BACE1 and further clarified its mechanism. Therefore, the main purpose of this study is to study the localization and transport of BACE1 in neurons. More importantly, understand the effect of RTN3 on axonal transport and APP shear of BACE1. Our results show that BACE1 is not only distributed in the late Golgi apparatus, transport Golgi apparatus and early endocytosis, but also exists in axons and dendrites. We found that BACE1 was co-located with presynaptic protein Synaptophysin, suggesting that BACE1 was located in synapses. The over-release of Ap at synaptic position is related to the increase of amyloid deposition, so we also discussed whether the localization of BACE1 at synaptic position is regulated by its negative regulatory protein reticulon3 (RTN3). We found that the co-localization of BACE1, Synaptophysin and PSD95 was decreased by co-expression of RTN3 and BACE1 in primary cultured neurons, and this decrease was due to the inhibition of the over-expression of RTN3 on the cis-axon transport of BACE1. Our results suggest that the level of RTN3 can affect the transport of BACE1 in axons, and the regulation of axon transport of BACE1 can reduce the axon terminal release and amyloid deposition of A 尾, so it may be a therapeutic approach for Alzheimer's disease.
【学位授予单位】：中南大学
【学位级别】：博士
【学位授予年份】：2013
【分类号】：R749.16

【共引文献】