From the Section of Neurobiology and.
J Biol Chem. 2013 Sep 13;288(37):26668-77. doi: 10.1074/jbc.M113.476689. Epub 2013 Aug 7.
Increased processing of amyloid precursor protein (APP) and accumulation of neurotoxic amyloid β peptide (Aβ) in the brain is central to the pathogenesis of Alzheimer's disease (AD). Therefore, the identification of molecules that regulate Aβ generation is crucial for future therapeutic approaches for AD. We demonstrated previously that RanBP9 regulates Aβ generation in a number of cell lines and primary neuronal cultures by forming tripartite protein complexes with APP, low-density lipoprotein-related protein, and BACE1, consequently leading to increased amyloid plaque burden in the brain. RanBP9 is a scaffold protein that exists and functions in multiprotein complexes. To identify other proteins that may bind RanBP9 and regulate Aβ levels, we used a two-hybrid analysis against a human brain cDNA library and identified COPS5 as a novel RanBP9-interacting protein. This interaction was confirmed by coimmunoprecipitation experiments in both neuronal and non-neuronal cells and mouse brain. Colocalization of COPS5 and RanBP9 in the same subcellular compartments further supported the interaction of both proteins. Furthermore, like RanBP9, COPS5 robustly increased Aβ generation, followed by increased soluble APP-β (sAPP-β) and decreased soluble-APP-α (sAPP-α) levels. Most importantly, down-regulation of COPS5 by siRNAs reduced Aβ generation, implying that endogenous COPS5 regulates Aβ generation. Finally, COPS5 levels were increased significantly in AD brains and APΔE9 transgenic mice, and overexpression of COPS5 strongly increased RanBP9 protein levels by increasing its half-life. Taken together, these results suggest that COPS5 increases Aβ generation by increasing RanBP9 levels. Thus, COPS5 is a novel RanBP9-binding protein that increases APP processing and Aβ generation by stabilizing RanBP9 protein levels.
淀粉样前体蛋白(APP)的加工增加和神经毒性淀粉样β肽(Aβ)在大脑中的积累是阿尔茨海默病(AD)发病机制的核心。因此,鉴定调节 Aβ生成的分子对于 AD 的未来治疗方法至关重要。我们之前已经证明,RanBP9 通过与 APP、低密度脂蛋白相关蛋白和 BACE1 形成三组分蛋白复合物,在许多细胞系和原代神经元培养物中调节 Aβ的产生,从而导致大脑中的淀粉样斑块负担增加。RanBP9 是一种支架蛋白,存在于多蛋白复合物中并发挥作用。为了鉴定可能与 RanBP9 结合并调节 Aβ水平的其他蛋白质,我们使用针对人脑 cDNA 文库的双杂交分析,鉴定出 COPS5 是一种新的 RanBP9 相互作用蛋白。在神经元和非神经元细胞以及小鼠脑中的共免疫沉淀实验证实了这种相互作用。COPS5 和 RanBP9 在相同亚细胞区室中的共定位进一步支持了这两种蛋白质的相互作用。此外,与 RanBP9 一样,COPS5 强烈增加 Aβ的产生,随后可溶性 APP-β(sAPP-β)增加,可溶性 APP-α(sAPP-α)减少。最重要的是,siRNAs 下调 COPS5 可减少 Aβ的产生,表明内源性 COPS5 调节 Aβ的产生。最后,AD 大脑和 APΔE9 转基因小鼠中 COPS5 的水平显著增加,并且 COPS5 的过表达通过增加其半衰期强烈增加了 RanBP9 蛋白水平。总之,这些结果表明 COPS5 通过增加 RanBP9 水平来增加 Aβ的产生。因此,COPS5 是一种新的 RanBP9 结合蛋白,通过稳定 RanBP9 蛋白水平来增加 APP 的加工和 Aβ的产生。