Comparative proteome analysis of changes in the 26S proteasome during oocyte maturation in goldfish
Comparative proteome analysis of changes in the 26S proteasome during oocyte maturation in goldfish
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DOI：
10.1002/pmic.200600055
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发表时间：
2006-07-01
期刊：
影响因子：
3.4
通讯作者：
Tokumoto, Toshinobu
中科院分区：
文献类型：
--
作者：
Horiguchi, Ryo;Dohra, Hideo;Tokumoto, Toshinobu
蛋白酶体是一种大的、多亚单位的颗粒，是真核细胞中大多数受调控的细胞内蛋白质降解的蛋白质分解机制。蛋白酶体功能的改变对减数分裂细胞周期的调节可能是重要的。为了研究26S蛋白酶体在减数分裂成熟过程中亚单位水平的变化，我们从金鱼未成熟和成熟的卵母细胞中纯化了26S蛋白酶体。采用双向聚丙烯酰胺凝胶电泳法分离蛋白质。通过基质辅助激光解吸/电离飞行时间质谱仪和数据库分析鉴定了金鱼卵母细胞26S蛋白酶体的全部斑点，用于差异分析。明确识别了四个不同的斑点(仅在成熟卵母细胞265蛋白酶体中检测到，而在未成熟卵母细胞中未检测到)和四个上调或下调的蛋白质斑点。成熟特异点不是26S蛋白酶体组分，而是它们的相互作用蛋白，并被鉴定为含有伴侣蛋白的TCP-1亚基和肌球蛋白轻链。20S核心颗粒的3个亚基和19S调节颗粒之一的次要斑点显示出依赖于减数分裂细胞周期的变化。这些结果表明，在金鱼卵母细胞成熟过程中，蛋白酶体亚基的改变和蛋白质与蛋白酶体的细胞周期相依赖的相互作用发生。
Proteasomes are large, multi-subunit particles that act as the proteolytic machinery for most of the regulated intracellular protein degradation in eukaryotic cells. An alteration of proteasome function may be important for the regulation of the meiotic cell cycle. To study the change at the subunit level of the 26S proteasome during meiotic maturation, we purified 26S proteasomes from immature and mature oocytes of goldfish. Two-dimensional polyacrylamide gel electrophoresis was used to separate proteins. For differential analysis, whole spots of the 26S proteasome from goldfish oocytes were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry and database analysis. Four spots that were different (only detected in mature oocyte 265 proteasomes and not in immature ones) and four protein spots that were up- or down-regulated were identified unambiguously. The mature-specific spots were not 26S proteasome components but rather their interacting proteins, and were identified as chaperonin-containing TCP-1 subunits and myosin light chain. Minor spots of three subunits of the 20S core particle and one of the 19S regulatory particle showed meiotic cell cycle-dependent changes. These results demonstrate that modifications of proteasomal subunits and cell cycle phase-dependent interactions of proteins with proteasomes occur during oocyte maturation in goldfish.