‡Biochemistry, Molecular and Structural Biology section, KU Leuven, Celestijnenlaan 200G, B-3001 Leuven-Heverlee, Flanders, Belgium.
Biochem J. 2013 Nov 1;455(3):295-306. doi: 10.1042/BJ20130417.
Accumulation of aggregated forms of αSyn (α-synuclein) into Lewy bodies is a known hallmark associated with neuronal cell death in Parkinson's disease. When expressed in the yeast Saccharomyces cerevisiae, αSyn interacts with the plasma membrane, forms inclusions and causes a concentration-dependent growth defect. We have used a yeast mutant, cog6Δ, which is particularly sensitive to moderate αSyn expression, for screening a mouse brain-specific cDNA library in order to identify mammalian proteins that counteract αSyn toxicity. The mouse ribosomal and chaperone protein RPS3A was identified as a suppressor of αSyn [WT (wild-type) and A53T] toxicity in yeast. We demonstrated that the 50 N-terminal amino acids are essential for this function. The yeast homologues of RPS3A were not effective in suppressing the αSyn-induced growth defect, illustrating the potential of our screening system to identify modifiers that would be missed using yeast gene overexpression as the first screening step. Co-expression of mouse RPS3A delayed the formation of αSyn-GFP inclusions in the yeast cells. The results of the present study suggest that the recently identified extraribosomal chaperonin function of RPS3A also acts on the neurodegeneration-related protein αSyn and reveal a new avenue for identifying promising candidate mammalian proteins involved in αSyn functioning.
α-突触核蛋白（α-synuclein）聚集形式的积累是帕金森病中与神经元细胞死亡相关的已知标志。当在酵母酿酒酵母中表达时，α-突触核蛋白与质膜相互作用，形成包含物并导致浓度依赖性的生长缺陷。我们使用了一种酵母突变体 cog6Δ，它对适度的α-突触核蛋白表达特别敏感，用于筛选小鼠大脑特异性 cDNA 文库，以鉴定对抗α-突触核蛋白毒性的哺乳动物蛋白。鼠核糖体和伴侣蛋白 RPS3A 被鉴定为酵母中 α-突触核蛋白[WT（野生型）和 A53T]毒性的抑制剂。我们证明，50 个 N 端氨基酸对于该功能是必需的。RPS3A 的酵母同源物在抑制 α-突触核蛋白诱导的生长缺陷方面没有效果，这说明了我们的筛选系统有可能识别到使用酵母基因过表达作为第一步筛选时会错过的修饰因子。鼠 RPS3A 的共表达延迟了酵母细胞中 α-突触核蛋白-GFP 包含物的形成。本研究的结果表明，最近发现的 RPS3A 的核糖体外伴侣蛋白功能也作用于与神经退行性变相关的蛋白α-突触核蛋白，并为鉴定参与α-突触核蛋白功能的有前途的候选哺乳动物蛋白开辟了新途径。
