Multiomics of GCN4-dependent replicative lifespan extension models reveals Gcn4 as a regulator of protein turnover in yeast

We have shown that multiple tRNA synthetase inhibitors can increase lifespan in both the nematode C. elegans and the budding yeast S. cerevisiae by acting through the conserved transcription factor Gcn4 (yeast) / ATF-4 (worms). To further understand the biology downstream of this conserved transcription factor in the yeast model system, we looked at two different yeast models known to have both upregulated Gcn4, and GCN4-dependent increased replicative lifespan. These two models are rpl31aΔ yeast, and yeast treated with the tRNA synthetase inhibitor borrelidin. We used both proteomic and RNAseq analysis of a block experimental design that included both of these models, to identify GCN4-dependent changes in these two longlived strains of yeast. Proteomic analysis of these yeast indicate that the longlived yeast have increased abundance of proteins involved in amino acid biosynthesis. RNASeq of these same yeast uncovered further regulation of protein turnover, identifying the differential expression of genes associated with both autophagy and with the ubiquitin proteasome system. The data presented here further underscore the important role that GCN4 and its orthologs play in the maintenance of protein homeostasis, which is itself an important hallmark of aging. Importantly, these changes could also have wider-ranging implications in the understanding and treatment of diseases of aging characterized by protein aggregation.我们已证实，多种tRNA合成酶抑制剂（tRNA synthetase inhibitor）可通过保守转录因子Gcn4（酵母）/ATF-4（线虫），分别延长秀丽隐杆线虫（Caenorhabditis elegans, C. elegans）和酿酒酵母（Saccharomyces cerevisiae, S. cerevisiae）的寿命。为进一步解析该保守转录因子在酿酒酵母模型系统中的下游生物学过程，我们选取了两种已被证实存在Gcn4上调、且经GCN4依赖性途径延长复制型寿命的酵母模型：rpl31aΔ缺失酵母，以及经tRNA合成酶抑制剂博罗利定（borrelidin）处理的酵母。我们采用包含这两种模型的区组实验设计，同时开展蛋白质组学（proteomic）与RNA测序（RNAseq）分析，以鉴定这两种长寿酵母菌株中GCN4依赖性的基因表达变化。蛋白质组学分析显示，长寿酵母中参与氨基酸生物合成的蛋白质丰度显著升高。对同一批酵母样本的RNA测序结果则揭示了蛋白质周转的更为复杂的调控机制，鉴定出与自噬（autophagy）及泛素-蛋白酶体系统（ubiquitin proteasome system）相关的基因存在差异表达。本研究数据进一步证实了GCN4及其同源物在维持蛋白质稳态（protein homeostasis）中的核心作用，而蛋白质稳态本身即是衰老的重要标志之一。值得关注的是，这些变化对于理解和治疗以蛋白质聚集为特征的衰老相关疾病，亦具有更为广泛的潜在价值。