Qian Chong-Sheng, Li Ling-Jie, Huang Hai-Wen, Yang Hai-Fei, Wu De-Pei
1Department of Hematology, The First Affiliated Hospital of Soochow University, No. 188, Shizi Street, Suzhou, 215006 People's Republic of China.
2Jiangsu Institute of Hematology, The First Affiliated Hospital of Soochow University, Suzhou, 215006 People's Republic of China.
Cancer Cell Int. 2020 Mar 19;20:87. doi: 10.1186/s12935-020-1158-6. eCollection 2020.
LncRNA NEAT1 has been identified as a tumour driver in many human cancers. However, the underlying mechanism of lncRNA NEAT1 in diffuse large B-cell lymphoma (DLBCL) progression is unclear.
The expression levels of NEAT1, GLI1 and miR-34b-5p were detected by RT-qPCR and Western blotting in DLBCL tissues and cell lines. MTT and colony formation assays were performed to examine cell proliferation, while annexin-V staining and TUNEL assays were performed to measure cell apoptosis. The effect of NEAT1, GLI1 and miR-34b-5p on cell cycle-associated proteins was evaluated by Western blotting. Dual-luciferase reporter and RNA immunoprecipitation (RIP) assays were employed to investigate the interaction between NEAT1 and miR-34b-5p or GLI1 and miR-34b-5p. Moreover, chromatin immunoprecipitation (ChIP) was performed to demonstrate the interaction between MYC and NEAT1.
NEAT1 and GLI1 were upregulated while miR-34b-5p was downregulated in DLBCL tissues and cell lines compared to normal controls. Knockdown of NEAT1 or overexpression of miR-34b-5p inhibited cell proliferation but promoted cell apoptosis. Overexpression of NEAT1 reversed GLI1-knockdown induced attenuation of cell proliferation. In other words, NEAT1 acted as a competing endogenous RNA (ceRNA), regulating the miR-34b-5p-GLI1 axis, further affecting the proliferation of DLBCL. Moreover, MYC modulated NEAT1 transcription by directly binding to the NEAT1 promoter.
We revealed that MYC-regulated NEAT1 promoted DLBCL proliferation via the miR-34b-5p-GLI1 pathway, which could provide a novel therapeutic target for DLBCL.
长链非编码RNA NEAT1已被确定为多种人类癌症中的肿瘤驱动因子。然而，lncRNA NEAT1在弥漫性大B细胞淋巴瘤（DLBCL）进展中的潜在机制尚不清楚。
采用RT-qPCR和蛋白质免疫印迹法检测DLBCL组织和细胞系中NEAT1、GLI1和miR-34b-5p的表达水平。进行MTT和集落形成试验以检测细胞增殖，同时进行膜联蛋白V染色和TUNEL试验以检测细胞凋亡。通过蛋白质免疫印迹法评估NEAT1、GLI1和miR-34b-5p对细胞周期相关蛋白的影响。采用双荧光素酶报告基因和RNA免疫沉淀（RIP）试验研究NEAT1与miR-34b-5p或GLI1与miR-34b-5p之间的相互作用。此外，进行染色质免疫沉淀（ChIP）试验以证明MYC与NEAT1之间的相互作用。
与正常对照相比，DLBCL组织和细胞系中NEAT1和GLI1上调，而miR-34b-5p下调。敲低NEAT1或过表达miR-34b-5p可抑制细胞增殖，但促进细胞凋亡。NEAT1的过表达逆转了GLI1敲低诱导的细胞增殖减弱。换句话说，NEAT1作为一种竞争性内源RNA（ceRNA），调节miR-34b-5p-GLI1轴，进一步影响DLBCL的增殖。此外，MYC通过直接结合NEAT1启动子调节NEAT1转录。
我们发现MYC调节的NEAT1通过miR-34b-5p-GLI1途径促进DLBCL增殖，这可能为DLBCL提供一个新的治疗靶点。
