苏木化调节BACE1磷酸化和泛素化参与AD发生的机制研究

Aβ is the major component of AD-associated senile plaques and the cleavage of APP by BACE1 is the rate-limiting step in Aβ generation during AD pathogenesis. Several post-translational modifications of BACE1 regulate its intracellular trafficking and its APP β site-mediated cleavage, but the relationship between these processes is not fully understood. We have recently found that BACE1 SUMOylation is enhanced in APP/PS1 mouse brain. We have also found that SUMOylation at Lys501 modulates BACE1 to accumulate in endocytic compartments and increases its protease activity thus aggravating Aβ deposition and AD pathology. SUMOylation at Lys501 inhibits phosphorylation of BACE1 on Ser498. We speculate that BACE1 SUMOylation is upregulated in AD, and this not only inhibits the ubiquitination of BACE1 on K501 and its degradation, but also inhibits the phosphorylation of BACE1 by blocking the interaction between BACE1 and its related kinases, resulting in BACE1 accumulation and over-activation and therefore increasing Aβ deposition and AD pathology. In the present study, we will make use of adeno-associated virus infection, biochemical molecular biology and electrophysiology to first confirm how SUMOylation mediates the inhibition of BACE1 phosphorylation. Then, we will investigate that SUMOylation of BACE1 aggregates amyloid deposition and AD pathology through inhibiting its phosphorylation and ubiquition. The results will reveal novel mechanisms underlying amyloid deposition, which will provide a new target for therapeutic intervention in AD.

老年斑是AD的主要病理特征之一，其主要成分Aβ的产生依赖于BACE1对APP的剪切，BACE1活性与翻译后修饰密切相关，但机制不明。申请者最近发现，APP/PS1小鼠脑内BACE1苏木化显著上调；苏木化导致BACE1在内涵体分布增加，促进APP-β剪切，加剧AD病程；BACE1 K501苏木化抑制S498位点磷酸化。鉴于此，我们推测，AD风险因素上调BACE1的苏木化，一方面阻遏与上游激酶结合，抑制BACE1磷酸化，提高其酶活性；另一方面抑制BACE1泛素化，逃逸蛋白酶体降解，增加其稳定，二者共同促进Aβ生成，导致神经毒性及认知障碍，促使AD发生。本项目拟联合运用腺相关病毒感染、生化分子生物学、行为学和电生理技术，在细胞和AD动物水平，确定BACE1苏木化与磷酸化的相互关系；阐明BACE1苏木化通过抑制磷酸化和泛素化参与AD病程。研究结果将为AD发病机制提供新线索，为药物研发提供新靶标。

细胞外老年斑（senile plaques, SPs）是阿尔茨海默病(Alzheimer’s disease, AD)的主要病理特征之一，其主要成分淀粉样蛋白（Aβ）的产生依赖于限速酶β分泌酶 （BACE1）对淀粉样前体蛋白APP的剪切，BACE1 活性与翻译后修饰密切相关，但其翻译后修饰的调控机制不明。本项目明确了BACE1苏木化与磷酸化、泛素化的相互关系，研究了苏木特异蛋白酶2（SUMO-specific proteases-2，SENP2）介导BACE1去苏木化与AD病变的分子机制，探讨了高脂饲养促进BACE1苏木化和去磷酸化加剧AD病变的相关机制。发现：1. BACE1的苏木化修饰抑制其磷酸化；2. BACE1的磷酸化修饰抑制其苏木化；3. BACE1的苏木化修饰抑制其泛素化；4. BACE1 苏木化抑制磷酸化和泛素化参与AD病理；5. SENP2介导BACE1 去苏木化降低其稳定性；6. 在AD模型动物中，SENP2表达水平与年龄呈负相关；7. 过表达SENP2 减少APP/PS1 小鼠的 Aβ斑块沉积以及缓解突触损伤和认知障碍。8. 在AD模型动物中，高脂饲养促进BACE1的苏木化和去磷酸化，加剧AD病变。本研究成果为BACE1苏木化异常修饰参与AD发病机制提供了新线索，为AD药物开发提供有效特异的分子靶点。