肿瘤细胞的快速增殖和持续分裂依赖膜脂的加倍。因此旺盛的脂质合成被认为是肿瘤细胞代谢的关键特征之一【1-3】。ATP-柠檬酸裂解酶（ATP-citrate lyase, ACLY）是脂肪酸合成代谢途径的第一个关键酶，催化底物柠檬酸与CoA形成脂肪酸从头合成所需的二碳单位乙酰辅酶A。ACLY的表达和活性受到磷酸化、乙酰化和泛素化的调节【4-7】；而近期研究发现，ACLY也可以发生O-GlcNAc糖基化修饰。O-GlcNAc修饰被广泛认为是一种可以感受营养波动的“感受器”（sensor）【8-12】，但ACLY的O-GlcNAc糖基化修饰如何感知营养丰度及其生物学意义亟待揭示。

    2024年10月10日，东北师范大学分子表观遗传学教育部重点实验室魏民、冯云鹏团队在PNAS上发表的题为“O-GlcNAcylation of ATP-citrate lyase couples glucose supply to lipogenesis for rapid tumor cell proliferation”的研究论文，报道了O-GlcNAc糖基化修饰在调节ACLY活性中的关键作用。 东北师范大学刘佳博士、王杨博士和田苗苗博士为该研究的共同第一作者；冯云鹏教授、魏民教授和金鑫博士为共同通讯作者。

     该研究阐释了ACLY通过 O-GlcNAc糖基化修饰感知葡萄糖浓度的升高，进而增强脂类合成促进肿瘤细胞快速增殖的分子机制。蛋白质谱分析显示，979位丝氨酸（S979）是ACLY的主要O-GlcNAc糖基化修饰位点。该位点的O-GlcNAc糖基化修饰能够显著增强底物CoA与ACLY的结合，从而提高ACLY的酶活性，促进脂肪酸及脂质的生成。基于 [U-13C] 葡萄糖的代谢流示踪分析发现， ACLY S979的O-GlcNAc糖基化修饰可以根据葡萄糖浓度的变化，对进入脂肪酸从头合成代谢途径的葡萄糖来源的碳单位进行动态调节。因此，ACLY S979的O-GlcNAc糖基化修饰可以响应葡萄糖浓度的变化，从而将葡萄糖供给与脂质合成连接在一起。研究人员发现，ACLY的O-GlcNAc修饰水平在肿瘤细胞/组织中显著升高。小鼠体内荷瘤实验显示，破坏ACLY S979的O-GlcNAc糖基化修饰能够显著抑制葡萄糖以及EGF对肿瘤生长产生的促进作用。 

    值得注意的是，已知生长因子EGF可以通过提高ACLY S455的磷酸化修饰，增强底物柠檬酸与ACLY的结合，提高ACLY的酶活性【6,13-17】。而该研究阐释的是ACLY S979的O-GlcNAc糖基化修饰增强另一底物底物CoA与ACLY的结合，且这两个位点的修饰对于产物的生成缺一不可。尽管ACLY S455的磷酸化与S979的糖基化两种修饰之间并不存在“cross-talk”，但EGF信号通过促进葡萄糖的吸收，进而上调ACLY的O-GlcNAc糖基化修饰。显然，决定ACLY活性的营养依赖的糖基化修饰和丝裂原信号指导的磷酸化修饰直接或间接地受EGF信号调控。该研究一方面阐释了细胞如何感知糖类物质的丰度指导脂肪酸合成进而促进细胞增殖的机制；另一方面也为从演化视角思考多细胞复杂有机体的细胞增殖调控机制增添了新的理解：虽然多细胞复杂有机体的增殖受丝裂原信号的控制，但决定单细胞生物增殖的古老的营养感知机制是保守的，丝裂原信号促进细胞增殖的过程依然需要通过特定的营养“检查点”（checkpoint）的核验。

  摘要：Abstract

Elevated lipid synthesis is one of the best-characterized metabolic alterations in cancer and crucial for membrane expansion. As a key rate-limiting enzyme in de novo fatty acid synthesis, ATP-citrate lyase (ACLY) is frequently up-regulated in tumors and regulated by posttranslational modifications (PTMs). Despite emerging evidence showing O-GlcNAcylation on ACLY, its biological function still remains unknown. Here, we observed a significant upregulation of ACLY O-GlcNAcylation in various types of human tumor cells and tissues and identified S979 as a major O-GlcNAcylation site. Importantly, S979 O-GlcNAcylation is required for substrate CoA binding and crucial for ACLY enzymatic activity. Moreover, it is sensitive to glucose fluctuation and decisive for fatty acid synthesis as well as tumor cell proliferation. In response to EGF stimulation, both S979 O-GlcNAcylation and previously characterized S455 phosphorylation played indispensable role in the regulation of ACLY activity and cell proliferation; however, they functioned independently from each other. In vivo, streptozocin treatment- and EGFR overexpression-induced growth of xenograft tumors was mitigated once S979 was mutated. Collectively, this work helps comprehend how cells interrogate the nutrient enrichment for proliferation and suggests that although mammalian cell proliferation is controlled by mitogen signaling, the ancient nutrition-sensing mechanism is conserved and still efficacious in the cells of multicellular organisms.

Significance

Cancer cells are characterized by a high rate of fatty acid synthesis required for membrane expansion during rapid proliferation. As a key rate-limiting enzyme in de novo fatty acid synthesis, ACLY is frequently deregulated by PTMs in tumors. Despite recently identified O-GlcNAcylation on ACLY, its biological function is still unknown. Here, we identify ACLY S979 as a key O-GlcNAcylation site that is crucial for ACLY function. By improving CoA association, S979 O-GlcNAcylation enhances ACLY activity and in turn promotes lipogenesis and tumor cell proliferation. Also, it is capable of sensing nutritional alterations and EGF stimulation and fine-tunes fatty acid synthesis accordingly. Together, this work brings insights into the dynamic coordination between glucose supply and lipogenesis for rapid tumor cell proliferation.

DOI： https://doi.org/10.1073/pnas.2402674121 

原文链接：https://www.pnas.org/doi/10.1073/pnas.2402674121 

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