GCN5L1介导的线粒体乙酰化在乳腺癌的发生和转移中的功能研究

Historically mitochondrial research explored its primal role in cellular energy transduction. More recently the role of mitochondrial metabolism in retrograde signal transduction to control gene expression, cytosolic signaling and biosynthetic pathways have been recognized. These cellular regulatory roles of mitochondria are especially well defined during tumorigenesis where mitochondrial glutaminolysis, in parallel with aerobic glycolysis (Warburg effect), function as a favored oncogenic metabolic pathway. Glutaminolysis plays a pivotal role in the maintenance of mitochondrial bioenergetics, redox control and macromolecule biosynthesis. In parallel, mitochondrial protein acetylation, as a post-translational control node, regulates glutamine metabolism and mitochondrial lysine-deacetylase enzyme SIRT3 functions as a tumor suppressor. Our laboratory has focused on a novel protein GCN5L1 which has found to both facilitate mitochondrial protein acetylation. In a subsequent unbiased metabolomic steady-state and flux studies, I found that the genetic ablation of GCN5L1 in primary hepatocytes evoked a canonical cancer metabolic signature with the parallel induction of glycolysis and glutaminolysis. In addition, I found that GCN5L1 levels are diminished in metastatic breast cancer cell line and GCN5L1 deletion enhanced glutaminase activity. Collectively these data support the hypothesis that GCN5L1 functions as a metabolic oncogenic regulator to control mitochondrial enzymes acetylation to govern glucose and glutamine metabolism during tumorigenesis to facilitate tumor growth and metastasis. To interrogate this hypothesis, I propose the following specific aims: 1. To identify and characterize the role of GCN5L1 in regulating glutamine metabolism. 2. Role of GCN5L1-mediated mitochondrial acetylation in mitochondrial metabolic rewiring. 3. Functional characterization of GCN5L1 in the regulation of breast cancer development and metastasis.

线粒体不仅是细胞内的能量工厂，也通过胞内的信号转导调控胞质内的生物合成和细胞核中的基因表达。线粒体谷氨酰胺代谢增强和有氧糖酵解被证实为肿瘤代谢重要标志。已知线粒体蛋白乙酰化作为线粒体内最重要的翻译后修饰，调控线粒体代谢通路；线粒体去乙酰化酶SIRT3有抑癌功能，但是线粒体乙酰化是否能通过调控代谢重编程影响肿瘤的发生和发展还不清楚。我们前期工作发现了线粒体乙酰转移调控因子GCN5L1，它的缺失促进了谷氨酰胺酶活性和谷氨酰胺代谢。此外，我们发现转移性乳腺癌细胞系中GCN5L1水平降低。因此我们提出假设：GCN5L1通过调控线粒体乙酰化以控制谷氨酰胺代谢和肿瘤发生和转移。为了证实这一假设，本项目将研究以下内容：1. GCN5L1在调节谷氨酰胺代谢中的作用；2. GCN5L1介导的线粒体乙酰化在线粒体代谢重编程中的作用；3. GCN5L1在乳腺癌发生和转移中的功能。

一直以来的研究认为产能是线粒体的主要功能。近年来的研究表明，线粒体逆向信号能够调控细胞信号转导和基因表达。线粒体在肿瘤代谢重编程中的重要作用也逐渐被认识，包括谷氨酰胺代谢增加，有氧糖酵解增加都是肿瘤代谢的特征。谷氨酰胺代谢能够促进线粒体生物合成，调控细胞的氧化还原平衡。GCN5L1在细胞内有着与Sirt3相反的功能，正向调控线粒体蛋白乙酰化，我们通过代谢组学发现GCN5L1缺失的细胞出现了肿瘤代谢的特征，我们建立GCN5L1敲除的细胞系，发现缺失了GCN5L1的肝癌细胞增殖迅速，其机制是GCN5L1通过乙酰化调控了谷氨酰胺酶的活性，进而调控了mTORC1信号和细胞的增殖。在肝细胞敲除GCN5L1的小鼠中，我们用化学诱变剂建立了小鼠肝癌模型，结合腺相关病毒过表达系统，证明了GCN5L1缺失促进肝癌的发生，而GCN5L1高表达则抑制肝癌的发生。同时，我们应用了谷氨酰胺酶抑制剂，发现能够很好的抑制小鼠肝癌模型中肿瘤的生长。在临床样本中，我们发现了GCN5L1表达与谷氨酰胺酶活性以及mTORC1活性的相关性，说明了GCN5L1对肝癌发生的调控作用和机制，同时也说明了酶活性的调控而非转录调控对肿瘤代谢重编程有着重要的作用。