Proteomic Analysis of Dhh1 Complexes Reveals a Role for Hsp40 Chaperone Ydj1 in Yeast P-Body Assembly.
Proteomic Analysis of Dhh1 Complexes Reveals a Role for Hsp40 Chaperone Ydj1 in Yeast P-Body Assembly.
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DOI：
10.1534/g3.115.021444
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发表时间：
2015-09-21
期刊：
影响因子：
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通讯作者：
Dudley AM
中科院分区：
文献类型：
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作者：
Cary GA;Vinh DB;May P;Kuestner R;Dudley AM
p体（PB）是核糖核蛋白（RNP）复合物，当细胞暴露于应激时聚集成细胞质灶。尽管保守的mRNA衰变和翻译抑制机制是已知的PB的组成部分，但细胞如何以及为什么将RNP复合物组装成大的灶仍不清楚。利用质谱分析用核心PB蛋白Dhh1免疫分离的蛋白质，我们发现相当多的蛋白质含有低复杂性序列，类似于哺乳动物RNP颗粒中高度代表的蛋白质。我们还发现Hsp40的伴侣蛋白Ydj1，包含一个低复杂性结构域并控制朊病毒蛋白聚集，是形成Dhh1−GFP聚焦葡萄糖消耗所必需的。在PB诱导过程中，与Dhh1 - GFP可重复共富集的新一类蛋白质包括参与核苷酸或氨基酸代谢、糖酵解、转移RNA氨基酰化和蛋白质折叠的蛋白质。这些蛋白质中的许多已被证明在对其他压力的反应中形成病灶。最后，对Dhh1−GFP相关RNA的分析显示，编码PB蛋白Pat1和催化RNA及其相关线粒体RNA结合蛋白的mRNA富集。因此，对PB组成的全局表征揭示了对PB组装重要的蛋白质，并有证据表明RNA在PB功能中起积极作用。
P-bodies (PB) are ribonucleoprotein (RNP) complexes that aggregate into cytoplasmic foci when cells are exposed to stress. Although the conserved mRNA decay and translational repression machineries are known components of PB, how and why cells assemble RNP complexes into large foci remain unclear. Using mass spectrometry to analyze proteins immunoisolated with the core PB protein Dhh1, we show that a considerable number of proteins contain low-complexity sequences, similar to proteins highly represented in mammalian RNP granules. We also show that the Hsp40 chaperone Ydj1, which contains an low-complexity domain and controls prion protein aggregation, is required for the formation of Dhh1−GFP foci on glucose depletion. New classes of proteins that reproducibly coenrich with Dhh1−GFP during PB induction include proteins involved in nucleotide or amino acid metabolism, glycolysis, transfer RNA aminoacylation, and protein folding. Many of these proteins have been shown to form foci in response to other stresses. Finally, analysis of RNA associated with Dhh1−GFP shows enrichment of mRNA encoding the PB protein Pat1 and catalytic RNAs along with their associated mitochondrial RNA-binding proteins. Thus, global characterization of PB composition has uncovered proteins important for PB assembly and evidence suggesting an active role for RNA in PB function.