突触小泡蛋白 2B 作为 BACE1 的新型相互作用蛋白负调控 AβPP 的淀粉样蛋白生成过程。

Synaptic Vesicle Protein 2B Negatively Regulates the Amyloidogenic Processing of AβPP as a Novel Interaction Partner of BACE1.

机构信息

Department of Neurology, Graduate School of Medicine, Kyoto University, Kyoto, Japan.

Department of Human Health Science, Graduate School of Medicine, Kyoto University, Kyoto, Japan.

出版信息

J Alzheimers Dis. 2020;75(1):173-185. doi: 10.3233/JAD-200071.

DOI:10.3233/JAD-200071

PMID:32280101

Abstract

BACKGROUND

Given that amyloid-β (Aβ) peptide is produced and released at synapses, synaptic Aβ is one of the promising therapeutic targets to prevent synaptic dysfunction in Alzheimer's disease (AD). Although Aβ production begins with the cleavage of the amyloid-β protein precursor (AβPP) by β-site AβPP cleaving enzyme 1 (BACE1), the mechanism on how BACE1 is involved in AβPP processing at synapses remains unclear.

OBJECTIVE

This study aimed to identify novel BACE1 interacting proteins regulating Aβ production at the synapse.

METHODS

BACE1 interacting proteins were pulled down using a mass spectrometry-based proteomics of wild-type (WT) rat brain synaptoneurosome lysates utilizing anti-BACE1 antibody. Then, a novel BACE1 interactor was identified and characterized using experimental systems that utilized transfected cells and knockout (KO) mice.

RESULTS

Synaptic vesicle protein 2B (SV2B) was identified as a novel presynaptic interaction partner of BACE1. In HEK293 cells, co-overexpression of SV2B with BACE1 significantly reduced the sAβPPβ and Aβ levels released in the media; thus, SV2B overexpression negatively affected the AβPP cleavage by BACE1. Compared with those of WT mice, the hippocampal lysates of SV2B knockout mice had significantly elevated Aβ levels, whereas the β-secretase activity and the AβPP and BACE1 protein levels remained unchanged. Finally, a fractionation assay revealed that BACE1 was mislocalized in SV2B KO mice; hence, SV2B may be involved in BACE1 trafficking downregulating the amyloidogenic pathway of AβPP.

CONCLUSION

SV2B has a novel role of negatively regulating the amyloidogenic processing of AβPP at the presynapses.

摘要

背景

由于淀粉样蛋白-β（Aβ）肽在突触中产生和释放，突触 Aβ 是预防阿尔茨海默病（AD）中突触功能障碍的有希望的治疗靶标之一。尽管 Aβ 的产生始于β-位点 Aβ 蛋白前体（AβPP）被β-位点 AβPP 裂解酶 1（BACE1）切割，但 BACE1 如何参与突触中 AβPP 的加工机制尚不清楚。

目的

本研究旨在鉴定新的 BACE1 相互作用蛋白，以调节突触中 Aβ 的产生。

方法

利用抗 BACE1 抗体从野生型（WT）大鼠脑突触体裂解物中进行基于质谱的蛋白质组学分析，以拉下 BACE1 相互作用蛋白。然后，利用转染细胞和敲除（KO）小鼠的实验系统，对一种新的 BACE1 相互作用蛋白进行鉴定和表征。

结果

突触小泡蛋白 2B（SV2B）被鉴定为 BACE1 的一种新的突触前相互作用伙伴。在 HEK293 细胞中，SV2B 与 BACE1 的共表达显著降低了培养基中 sAβPPβ 和 Aβ 的释放水平；因此，SV2B 过表达负调控了 BACE1 对 AβPP 的切割。与 WT 小鼠相比，SV2B 敲除小鼠海马裂解物中的 Aβ 水平显著升高，而β-分泌酶活性以及 AβPP 和 BACE1 蛋白水平保持不变。最后，一项分馏实验表明，BACE1 在 SV2B KO 小鼠中发生了定位错误；因此，SV2B 可能参与 BACE1 的运输，从而下调 AβPP 的淀粉样蛋白形成途径。