Department of Neurosurgery, Shengjing Hospital of China Medical University, Shenyang, China.
Key Laboratory of Neuro-oncology in Liaoning Province, Shenyang, China.
CNS Neurosci Ther. 2023 Oct;29(10):2811-2825. doi: 10.1111/cns.14218. Epub 2023 Apr 17.
The isocitrate dehydrogenase (IDH) phenotype is associated with reprogrammed energy metabolism in glioblastoma (GBM) cells. Small nucleolar RNAs (snoRNAs) are known to exert an important regulatory role in the energy metabolism of tumor cells. The purpose of this study was to investigate the role of C/D box snoRNA U3 and transcription factor zinc finger and BTB domain-containing 7A (ZBTB7A) in the regulation of aerobic glycolysis and the proliferative capacity of IDH1 wild-type (IDH1 ) GBM cells.
Quantitative reverse transcription PCR and western blot assays were utilized to detect snoRNA U3 and ZBTB7A expression. U3 promoter methylation status was analyzed via bisulfite sequencing and methylation-specific PCR. Seahorse XF glycolysis stress assays, lactate production and glucose consumption measurement assays, and cell viability assays were utilized to detect glycolysis and proliferation of IDH1 GBM cells.
We found that hypomethylation of the CpG island in the promoter region of U3 led to the upregulation of U3 expression in IDH1 GBM cells, and the knockdown of U3 suppressed aerobic glycolysis and the proliferation ability of IDH1 GBM cells. We found that small nucleolar-derived RNA (sdRNA) U3-miR, a small fragment produced by U3, was able to bind to the ZBTB4 3'UTR region and reduce ZBTB7A mRNA stability, thereby downregulating ZBTB7A protein expression. Furthermore, ZBTB7A transcriptionally inhibited the expression of hexokinase 2 (HK2) and lactate dehydrogenase A (LDHA), which are key enzymes of aerobic glycolysis, by directly binding to the HK2 and LDHA promoter regions, thereby forming the U3/ZBTB7A/HK2 LDHA pathway that regulates aerobic glycolysis and proliferation of IDH1 GBM cells.
U3 enhances aerobic glycolysis and proliferation in IDH1 GBM cells via the U3/ZBTB7A/HK2 LDHA axis.
异柠檬酸脱氢酶（IDH）表型与胶质母细胞瘤（GBM）细胞中重新编程的能量代谢有关。小核仁 RNA（snoRNA）已知在肿瘤细胞的能量代谢中发挥重要的调节作用。本研究旨在探讨 C/D 框 snoRNA U3 和转录因子锌指和 BTB 结构域包含 7A（ZBTB7A）在调节 IDH1 野生型（IDH1）GBM 细胞有氧糖酵解和增殖能力中的作用。
利用定量逆转录 PCR 和 Western blot 检测 snoRNA U3 和 ZBTB7A 的表达。通过亚硫酸氢盐测序和甲基化特异性 PCR 分析 U3 启动子甲基化状态。利用 Seahorse XF 糖酵解应激测定、乳酸生成和葡萄糖消耗测定以及细胞活力测定来检测 IDH1 GBM 细胞的糖酵解和增殖。
我们发现 U3 启动子区域 CpG 岛的低甲基化导致 IDH1 GBM 细胞中 U3 的表达上调，而 U3 的敲低抑制了 IDH1 GBM 细胞的有氧糖酵解和增殖能力。我们发现，由 U3 产生的小片段 snoRNA U3-miR 能够与 ZBTB4 3'UTR 区域结合，并降低 ZBTB7A mRNA 的稳定性，从而下调 ZBTB7A 蛋白表达。此外，ZBTB7A 通过直接结合 HK2 和 LDHA 启动子区域，转录抑制有氧糖酵解的关键酶己糖激酶 2（HK2）和乳酸脱氢酶 A（LDHA）的表达，从而形成调节 IDH1 GBM 细胞有氧糖酵解和增殖的 U3/ZBTB7A/HK2 LDHA 通路。
U3 通过 U3/ZBTB7A/HK2 LDHA 轴增强 IDH1 GBM 细胞的有氧糖酵解和增殖。
