YTHDF2通过诱导IGFBP7的mRNA衰变激活PI3K/AKT信号转导通路促进胶质母细胞瘤进展的研究

doi:10.19401/j.cnki.1007-3639.2022.03.004

摘要/Abstract

摘要：

背景与目的：N6-甲基腺苷（N6-methyladenosine,m6A）甲基化修饰识别蛋白YTHDF2被证实在癌症进展中扮演重要角色。探讨YTHDF2是否通过促进胰岛素样生长因子结合蛋白7（insulin-like growth factor binding protein 7,IGFBP7）的mRNA衰变,激活磷脂酰肌醇3-激酶（phosphoinositide 3-kinase,PI3K）-蛋白激酶B（protein kinase,AKT）信号转导通路,调控胶质母细胞瘤（glioblastoma,GBM）的细胞周期和凋亡。方法：采用实时荧光定量聚合酶链反应（real-time fluorescence quantitative polymerase chain reaction,RTFQ-PCR）检测YTHDF2和IGFBP7在GBM组织和细胞中的表达。通过流式细胞术检测细胞周期和凋亡率。采用蛋白质印迹法（Western blot）检测p-AKT、AKT、p-PI3K和PI3K蛋白表达。采用RNA免疫沉淀（RNA immunoprecipitation,RIP）实验和RNA稳定性实验验证YTHDF2与IGFBP7之间的关系。结果：与癌旁组织和正常星形胶质细胞系（NHA细胞）相比,YTHDF2在GBM组织和细胞中表达升高,IGFBP7表达降低（P均<0.05）。敲减YTHDF2可诱导GBM细胞周期阻滞和凋亡率升高（P<0.05）。在GBM细胞中,YTHDF2可通过识别m6A修饰的IGFBP7进而促进IGFBP7 mRNA降解（P<0.05）。抑制IGFBP7可部分地挽救si-YTHDF2对GBM细胞周期和凋亡的影响（P均<0.05）。此外,YTHDF2还可通过调控IGFBP7激活PI3K/AKT信号转导通路促进GBM恶性进展（P<0.05）。结论：YTHDF2通过m6A修饰的方式调控IGFBP7的表达,激活PI3K/AKT信号转导通路以促进GBM细胞恶性生物学行为。

关键词: YTHDF2, 胶质母细胞瘤, 细胞周期, 细胞凋亡

Abstract:

Background and purpose: N6-methyladenosine (m6A) methylation modification recognition protein YTHDF2 has been shown to play an important role in cancer progression. This research was designed to explore whether YTHDF2 regulated cell cycle and apoptosis of glioblastoma (GBM) via promoting mRNA decay of insulin-like growth factor binding protein 7 (IGFBP7) and activating phosphoinositide 3-kinase (PI3K)-protein kinase (AKT) signaling pathway. Methods: The expressions of YTHDF2 and IGFBP7 in GBM tissues and cells were detected by real-time fluorescence quantitative polymerase chain reaction (RTFQ-PCR). The cell cycle and apoptosis rate were detected by flow cytometry. Protein expressions of p-AKT, AKT, p-PI3K and PI3K were detected by Western blot. RNA immunoprecipitation (RIP) assay and RNA stability experiment were used to verify the relationship between YTHDF2and IGFBP7. Results: Compared with adjacent tissues and normal astrocyte line (NHA cells) expression of YTHDF2 was increased while expression of IGFBP7 was decreased in GBM tissues and cells (all P<0.05). Knockdown of YTHDF2 could induce GBM cell cycle arrest and increase apoptotic rate (P<0.05). In GBM cells, YTHDF2 could induce IGFBP7 mRNA decay via recognizing m6A modified IGFBP7 (P<0.05). Inhibition of IGFBP7 partially rescued the effects of si-YTHDF2 on GBM cell cycle and apoptosis (all P<0.05). In addition, YTHDF2 promoted the malignant progression of GBM by regulating IGFBP7 to activate PI3K/AKT signaling pathway (P<0.05). Conclusion: YTHDF2 promotes the malignant biological behavior of GBM cells through regulating IGFBP7 expression by means of m6A modification and activating PI3K/AKT signaling pathway.

Key words: YTHDF2, Glioblastoma, Cell cycle, Cell apoptosis

中图分类号:

R739.41

刁新峰, 李新茂, 候亮, 魏志玄. YTHDF2通过诱导IGFBP7的mRNA衰变激活PI3K/AKT信号转导通路促进胶质母细胞瘤进展的研究[J]. 中国癌症杂志, 2022, 32(3): 218-227.

Diao Xinfeng, Li Xinmao, Hou Liang, Wei Zhixuan. YTHDF2 promotes progression of glioblastoma via inducing mRNA decay of IGFBP7 and activating PI3K/AKT signaling pathway[J]. China Oncology, 2022, 32(3): 218-227.

图/表 5

参考文献 20

[1]	张良龙, 安宏伟, 赵立智, 等. LncRNA LINC00152过表达在替莫唑胺诱导脑胶质瘤干细胞周期阻滞中的作用机制探讨[J]. 中国癌症杂志, 2020, 30(5): 328-334.
	ZHANG L L, AN H W, ZHAO L Z, et al. Mechanism of overexpression of lncRNA LINC00152 in temozolomide-induced stem cell cycle arrest in glioma[J]. China Oncol, 2020, 30(5): 328-334.
[2]	CHAVDA V, PATEL V, YADAV D, et al. Therapeutics and research related to glioblastoma: advancements and future targets[J]. Curr Drug Metab, 2020, 21(3): 186-198. doi: 10.2174/1389200221666200408083950
[3]	董明皓, 张霓, 何锋. N6-腺苷酸甲基化修饰在肿瘤发生发展中的作用[J]. 中华实验外科杂志, 2020, 37(9): 1770-1774.
	DONG M H, ZHANG N, HE F. The role of N6-methyladenosine modification in tumorigenesis and development[J]. Chin J Exp Surg, 2020, 37(9): 1770-1774.
[4]	ZHANG C Z, HUANG S Z, ZHUANG H K, et al. YTHDF2 promotes the liver cancer stem cell phenotype and cancer metastasis by regulating OCT4 expression via m6A RNA methylation[J]. Oncogene, 2020, 39(23): 4507-4518. doi: 10.1038/s41388-020-1303-7
[5]	RUPP C, SCHERZER M, RUDISCH A, et al. IGFBP7, a novel tumor stroma marker, with growth-promoting effects in colon cancer through a paracrine tumor-stroma interaction[J]. Oncogene, 2015, 34(7): 815-825. doi: 10.1038/onc.2014.18
[6]	PEN A, DUROCHER Y, SLINN J, et al. Insulin-like growth factor binding protein 7 exhibits tumor suppressive and vessel stabilization properties in U87MG and T98G glioblastoma cell lines[J]. Cancer Biol Ther, 2011, 12(7): 634-646. doi: 10.4161/cbt.12.7.17171
[7]	CUI Q, SHI H L, YE P, et al. m6A RNA methylation regulates the self-renewal and tumorigenesis of glioblastoma stem cells[J]. Cell Rep, 2017, 18(11): 2622-2634. doi: 10.1016/j.celrep.2017.02.059
[8]	DU J Y, HOU K Y, MI S, et al. Malignant evaluation and clinical prognostic values of m6A RNA methylation regulators in glioblastoma[J]. Front Oncol, 2020, 10: 208. doi: 10.3389/fonc.2020.00208
[9]	FANG R P, CHEN X, ZHANG S C, et al. EGFR/SRC/ERK-stabilized YTHDF2 promotes cholesterol dysregulation and invasive growth of glioblastoma[J]. Nat Commun, 2021, 12(1): 177. doi: 10.1038/s41467-020-20379-7
[10]	LI J F, XIE H Y, YING Y F, et al. YTHDF2 mediates the mRNA degradation of the tumor suppressors to induce AKT phosphorylation in N6-methyladenosine-dependent way in prostate cancer[J]. Mol Cancer, 2020, 19(1): 152. doi: 10.1186/s12943-020-01267-6
[11]	HONG L, PU X W, GAN H L, et al. YTHDF2 inhibit the tumorigenicity of endometrial cancer via downregulating the expression of IRS1 methylated with m6A[J]. J Cancer, 2021, 12(13): 3809-3818. doi: 10.7150/jca.54527
[12]	LI Y F, XI Y F, ZHU G Q, et al. Downregulated IGFBP7 facilitates liver metastasis by modulating epithelial-mesenchymal transition in colon cancer[J]. Oncol Rep, 2019, 42(5): 1935-1945.
[13]	ZHAO Q Y, ZHAO R L, SONG C H, et al. Increased IGFBP7 expression correlates with poor prognosis and immune infiltration in gastric cancer[J]. J Cancer, 2021, 12(5): 1343-1355. doi: 10.7150/jca.50370
[14]	AKIEL M, GUO C Q, LI X, et al. IGFBP7 deletion promotes hepatocellular carcinoma[J]. Cancer Res, 2017, 77(15): 4014-4025. doi: 10.1158/0008-5472.CAN-16-2885
[15]	TIAN X Y, ZHANG L, SUN L G, et al. Low expression of insulin-like growth factor binding protein 7 associated with poor prognosis in human glioma[J]. J Int Med Res, 2014, 42(3): 651-658. doi: 10.1177/0300060513503926
[16]	ZHANG C, ZHANG M Q, GE S, et al. Reduced m6A modification predicts malignant phenotypes and augmented Wnt/PI3K/AKT signaling in gastric cancer[J]. Cancer Med, 2019, 8(10): 4766-4781. doi: 10.1002/cam4.v8.10
[17]	YANG J B, DING W L, WANG X Y, et al. Knockdown of DNA polymerase ζ relieved the chemoresistance of glioma via inhibiting the PI3K/AKT signaling pathway[J]. Bioengineered, 2021, 12(1): 3924-3933. doi: 10.1080/21655979.2021.1944027
[18]	EVDOKIMOVA V, TOGNON C E, BENATAR T, et al. IGFBP7 binds to the IGF-1 receptor and blocks its activation by insulin-like growth factors[J]. Sci Signal, 2012, 5(255): ra92.
[19]	ZHANG L, LIAN R, ZHAO J J, et al. IGFBP7 inhibits cell proliferation by suppressing AKT activity and cell cycle progression in thyroid carcinoma[J]. Cell Biosci, 2019, 9: 44. doi: 10.1186/s13578-019-0310-2
[20]	HADDADI N, LIN Y G, TRAVIS G, et al. PTEN/PTENP1: ‘regulating the regulator of RTK-dependent PI3K/AKT signalling’, new targets for cancer therapy[J]. Mol Cancer, 2018, 17(1): 37. doi: 10.1186/s12943-018-0803-3