Sam68 promotes aerobic glycolysis in colorectal cancer by regulating PKM2 alternative splicing
Sam68 promotes aerobic glycolysis in colorectal cancer by regulating PKM2 alternative splicing
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Sam68通过调节PKM2选择性剪接促进结直肠癌的有氧糖酵解
DOI：
10.21037/atm.2020.03.108
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发表时间：
2020-04-01
影响因子：
--
通讯作者：
Huang, Zhaohui
中科院分区：
文献类型：
--
作者：
Zhao, Jing;Li, Jiuming;Huang, Zhaohui
Sam 68是一种RNA结合蛋白，在多种癌症中发挥致癌作用。然而，Sam 68在结直肠癌（CRC）中的具体功能和机制尚未阐明。丙酮酸激酶肌肉（PKM）2是糖酵解中的关键限速酶，PKM 2维持大多数癌细胞中糖酵解主导的能量代谢。方法CCK-8法检测Sam 68对细胞生长的影响。进行丙酮酸激酶活性和乳酸检测测定以分析Sam 68对有氧糖酵解的影响。RNA免疫沉淀（RIP）用于检测Sam 68与PKM 2序列的结合。Western blot和实时荧光定量PCR分析Sam 68对PKM 2的调控作用。结果功能获得和功能丧失研究表明，异位表达的Sam 68促进大肠癌细胞的糖酵解和细胞增殖，而敲低Sam 68则抑制大肠癌细胞的糖酵解和细胞增殖。Sam 68通过调节丙酮酸激酶（PKM 2或PKM 1）的选择性剪接来调节其表达谱。Sam 68的过表达与PKM 1/PKM 2比值降低相关，这对糖酵解过程有积极贡献。Sam 68可显著促进CRC细胞增殖，降低PKM 1/PKM 2比值，导致CRC细胞葡萄糖代谢由氧化磷酸化转变为糖酵解。此外，Sam 68还能促进PKM 2 mRNA从细胞核向细胞质的转运，增加PKM 2蛋白的表达，从而提高丙酮酸激酶活性和乳酸产量。结论Sam 68可能通过调节PKM 2的选择性剪接和表达，影响结直肠癌细胞的生长和糖酵解途径。
Background Sam68, an RNA-binding protein, exerts oncogenic functions in several types of cancer. However, the specific functions and mechanisms of Sam68 in colorectal cancer (CRC) had not been previously clarified. Pyruvate kinase muscle (PKM)2 is the key rate-limiting enzyme in glycolysis, and PKM2 maintains the glycolysis-dominant energy metabolism in most cancer cells. Methods CCK8 assay was performed to show the effect of Sam68 on cell growth. Pyruvate kinase activity and lactate detection assays were performed to analyze the effects of Sam68 on aerobic glycolysis. RNA immunoprecipitation (RIP) was used to detect the binding of Sam68 to the PKM2 sequence. Western blot and real-time PCR were executed to analyze the regulation of PKM2 by Sam68. Results Gain-of-function and loss-of-function studies showed that ectopic expression of Sam68 promoted glycolysis and cell proliferation in CRC cells, whereas Sam68 knockdown inhibited glycolysis and cell proliferation. Mechanically, Sam68 modulated the expression profile of pyruvate kinase (PKM2 or PKM1) by regulating its alternative splicing. Overexpression of Sam68 was associated with decreased PKM1/PKM2 ratio, which positively contributed to the glycolysis procedure. Sam68 significantly promoted cell proliferation and caused a decrease of PKM1/PKM2 ratio, resulting in the metabolism of glucose switched from oxidative phosphorylation to glycolysis in CRC cells. Besides, Sam68 enhanced PKM2 mRNA transport from the nucleus to cytoplasm and increased the expression of PKM2 protein, resulting in elevated pyruvate kinase activity and lactate production. Conclusions These findings suggested that Sam68 affected cell growth and glycolysis pathway by regulating the alternative splicing and expression of PKM2 in CRC.