Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ottawa, Canada.
Ottawa Institute of Systems Biology, Ottawa, Canada.
PLoS Genet. 2020 Nov 30;16(11):e1009220. doi: 10.1371/journal.pgen.1009220. eCollection 2020 Nov.
Cellular metabolism is tightly regulated by many signaling pathways and processes, including lysine acetylation of proteins. While lysine acetylation of metabolic enzymes can directly influence enzyme activity, there is growing evidence that lysine acetylation can also impact protein localization. As the Saccharomyces cerevisiae lysine acetyltransferase complex NuA4 has been implicated in a variety of metabolic processes, we have explored whether NuA4 controls the localization and/or protein levels of metabolic proteins. We performed a high-throughput microscopy screen of over 360 GFP-tagged metabolic proteins and identified 23 proteins whose localization and/or abundance changed upon deletion of the NuA4 scaffolding subunit, EAF1. Within this, three proteins were required for glycogen synthesis and 14 proteins were associated with the mitochondria. We determined that in eaf1Δ cells the transcription of glycogen biosynthesis genes is upregulated resulting in increased proteins and glycogen production. Further, in the absence of EAF1, mitochondria are highly fused, increasing in volume approximately 3-fold, and are chaotically distributed but remain functional. Both the increased glycogen synthesis and mitochondrial elongation in eaf1Δ cells are dependent on Bcy1, the yeast regulatory subunit of PKA. Surprisingly, in the absence of EAF1, Bcy1 localization changes from being nuclear to cytoplasmic and PKA activity is altered. We found that NuA4-dependent localization of Bcy1 is dependent on a lysine residue at position 313 of Bcy1. However, the glycogen accumulation and mitochondrial elongation phenotypes of eaf1Δ, while dependent on Bcy1, were not fully dependent on Bcy1-K313 acetylation state and subcellular localization of Bcy1. As NuA4 is highly conserved with the human Tip60 complex, our work may inform human disease biology, revealing new avenues to investigate the role of Tip60 in metabolic diseases.
细胞代谢受到许多信号通路和过程的严格调控，包括蛋白质赖氨酸乙酰化。虽然代谢酶的赖氨酸乙酰化可以直接影响酶活性，但越来越多的证据表明赖氨酸乙酰化也可以影响蛋白质定位。由于酿酒酵母赖氨酸乙酰转移酶复合物 NuA4 被牵连到多种代谢过程中，我们探索了 NuA4 是否控制代谢蛋白的定位和/或蛋白质水平。我们对超过 360 个 GFP 标记的代谢蛋白进行了高通量显微镜筛选，发现有 23 种蛋白质的定位和/或丰度在缺失 NuA4 支架亚基 EAF1 时发生了变化。其中，有三种蛋白质与糖原合成有关，有 14 种蛋白质与线粒体有关。我们确定在 eaf1Δ 细胞中，糖原生物合成基因的转录上调，导致蛋白质和糖原的产生增加。此外，在没有 EAF1 的情况下，线粒体高度融合，体积增加约 3 倍，分布混乱，但仍保持功能。eaf1Δ 细胞中糖原合成和线粒体伸长的增加都依赖于 Bcy1，即酵母 PKA 的调节亚基。令人惊讶的是，在没有 EAF1 的情况下，Bcy1 的定位从核内变为细胞质，PKA 的活性也发生了改变。我们发现，NuA4 依赖的 Bcy1 定位依赖于 Bcy1 位置 313 上的赖氨酸残基。然而，eaf1Δ 细胞中糖原积累和线粒体伸长的表型虽然依赖于 Bcy1，但并不完全依赖于 Bcy1-K313 乙酰化状态和 Bcy1 的亚细胞定位。由于 NuA4 与人类 Tip60 复合物高度保守，我们的工作可能为人类疾病生物学提供信息，揭示研究 Tip60 在代谢疾病中的作用的新途径。
