蛋白酶体降解细胞内大多数的蛋白质，并在细胞分裂、免疫应答和细胞死亡等各种细胞活动中发挥关键作用。真核生物蛋白酶体由一个20S核心颗粒和不同种类的蛋白酶体激活因子组成。根据蛋白酶体激活因子的不同，蛋白酶体降解底物的方式可分为依赖或不依赖泛素化。以19S调节颗粒为激活因子的26S蛋白酶体在细胞中含量最为丰富，它能够识别并降解泛素化的底物蛋白质。以PA200为激活因子的蛋白酶体可以在精子发生和体细胞DNA损伤修复的过程中以依赖于乙酰化、而非泛素化的方式介导核心组蛋白的降解。但是，PA200-蛋白酶体的非核心组蛋白底物仍有待揭秘。

通过对野生型和PA200基因敲除的小鼠胚胎成纤维细胞进行蛋白质谱分析，我们课题组发现PA200基因敲除后可引起二千余种蛋白水平上调。本文选择其中的SCF复合型泛素连接酶的底物结合组分F-box蛋白2（F-Box only protein 2，FBXO2）作为研究对象。

本研究利用蛋白质免疫印迹实验发现，PA200基因敲除会引起FBXO2 蛋白降解减慢；而通过实时荧光定量PCR检测其mRNA水平时发现，PA200基因敲除并不会引起FBXO2转录水平的改变，提示PA200-蛋白酶体能够促进 FBXO2 的蛋白降解。去乙酰化酶抑制剂TSA可以促进FBXO2的降解。通过查阅蛋白质乙酰化数据库（http://pail.biocuckoo.org/wsresult.php），发现FBXO2的第166位的赖氨酸（Lys166）为其潜在乙酰化位点。当把FBXO2的Lys166突变为精氨酸后，PA200-蛋白酶体对于FBXO2的降解减弱。进一步研究表明，PA200-蛋白酶体通过其类似溴区结构域（Bromodomain-like domain）介导FBXO2的降解。在细胞中过表达野生型 PA200 时，可以明显促进 FBXO2 的降解，但过表达含有突变 BRDL 的PA200 则不能明显促进 FBXO2 的降解。此外，PA200-蛋白酶体还可以通过降解FBXO2促进突变型亨廷顿蛋白的积聚。

本研究确认了PA200-蛋白酶体通过依赖于乙酰化的方式降解非核心组蛋白底物FBXO2，同时提供了通过靶向PA200-FBXO2通路来防治亨廷顿病等神经退行性疾病的可能性。

Proteasomes degrade most intracellular proteins and play key roles in various cellular activities, such as cell division, immune response and cell death. The eukaryotic proteasome consists of a 20S core particle and different types of proteasome activators. Depending on the proteasome activator, the proteasome degrades substrates in a manner that can be classified as ubiquitination-dependent or ubiquitination-independent. The 26S proteasome, which uses the 19S regulatory particle as the activator,  degrades ubiquitinated proteins. The PA200-proteasome which uses PA200 as activator, degrades the core histones in an acetylation-dependent manner during spermatogenesis and somatic DNA repair. However, the non-core histone substrates of the PA200-proteasome remain to be uncovered.

By using mass spectrometric analysis of the proteins from wild-type and PA200-deficient mouse embryonic fibroblasts, our group found that deletion of PA200 can cause upregulation of more than two thousand protein. In this thesis, we selected F-box only protein 2 (FBXO2), a substrate binding component of SCF ubiquitin ligase complex for further study.

This study revealed that deletion of PA200 caused a slowdown of FBXO2 protein degradation through protein immunoblotting experiments, while the detection of its mRNA level by quantitative real-time PCR revealed that deletion of PA200 did not change FBXO2 transcript levels, suggesting that PA200-proteasome can promote FBXO2 protein degradation. The deacetylase inhibitor TSA could accelerate the degradation of FBXO2. By searching the Protein Acetylation Database (http://pail.biocuckoo.org/wsresult.php), lysine at position 166 (Lys166) of FBXO2 was found as a potential acetylation site. The degradation of FBXO2 by the PA200-proteasome was attenuated when Lys166 of FBXO2 was mutated to arginine. Meanwhile, the PA200-proteasome mediate the degradation of FBXO2 via its bromodomain-like domain. Overexpression of the wild-type PA200 promoted the degradation of FBXO2, but mutation of BRDL abolished this effect. In addition, PA200-proteasome can promote the accumulation of mutant Huntington protein by degrading FBXO2.

These results suggest that the PA200-proteasome promotes the degradation of FBXO2 in an acetylation-dependent manner, providing a possibility to modulate the PA200-FBXO2 pathway in prevention and treatment of neurodegenerative diseases, such as Huntington's disease.