METTL3 和 STAT3 形成正反馈环促进肝癌细胞转移。

METTL3 and STAT3 form a positive feedback loop to promote cell metastasis in hepatocellular carcinoma.

机构信息

Laboratory of Infection and Immunology, School of Medical Technology, Xinxiang Medical University, Xinxiang, Henan, 453003, People's Republic of China.

Key Laboratory of Laboratory Medicine, Ministry of Education of China, Zhejiang Provincial Key Laboratory of Medical Genetics, School of Laboratory Medicine and Life Sciences, Wenzhou Medical University, Wenzhou, 325035, China.

出版信息

Cell Commun Signal. 2023 May 25;21(1):121. doi: 10.1186/s12964-023-01148-7.

DOI:10.1186/s12964-023-01148-7

PMID:37231451

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC10210303/

Abstract

BACKGROUND

It is well-established that most Hepatocellular carcinoma (HCC) patients die of metastasis, yet the potential mechanisms orchestrating metastasis remain poorly understood. Current evidence suggests that the dysregulation of METTL3-mediated m6A methylation modification is closely associated with cancer progression. STAT3 is an oncogenic transcription factor that reportedly plays a central role in the occurrence and development of HCC. However, the relationship between METTL3 and STAT3 in HCC metastasis remains unclear.

METHODS

The relationship between METTL3 expression and the survival of HCC patients was assessed by online tools GEPIA and Kaplan-Meier Plotter. Western blotting, Tissue microarray (TMA), and immunohistochemistry (IHC) staining were used to evaluate the expression levels of METTL3 and STAT3 in HCC cell lines and metastatic and non-metastatic tissues. Methylated RNA immunoprecipitation (MeRIP), MeRIP sequencing (MeRIP-seq), qRT-PCR, RNA immunoprecipitation (RIP), Western blotting and luciferase reporter gene assay were utilized to clarify the mechanism of METTL3 regulating STAT3 expression. Immunofluorescence staining, Western blotting, qRT-PCR, Co-immunoprecipitation (Co-IP), IHC staining, TMA and Chromatin immunoprecipitation (ChIP) assay were performed to explore the mechanism of STAT3 modulating METTL3 localization. Cell viability, wound healing and transwell assay, and orthotopic xenograft model were used to evaluate the role of METTL3-STAT3 feedback loop in the promotion of HCC metastasis in vitro and in vivo.

RESULTS

METTL3 and STAT3 are both abundantly expressed in high-metastatic HCC cells and tissues. Moreover, a positive correlation was found between the expression of STAT3 and METTL3 in HCC tissues. Mechanistically, METTL3 could induce the m6A modification of STAT3 mRNA, and then promote the translation of m6A-contained STAT3 mRNA by interacting with the translation initiation machinery. In contrast, STAT3 promoted nuclear localization of METTL3 via transcriptionally upregulating WTAP, a vital member of the methyltransferase complex, and facilitated the methyltransferase function of METTL3. METTL3 and STAT3 form a positive feedback loop to accelerate HCC metastasis in vitro and in vivo.

CONCLUSIONS

Our findings reveal a novel mechanism of HCC metastasis and uncover the METTL3-STAT3 feedback signaling as a potential target for the anti-metastatic treatment of HCC. Video Abstract.

摘要

背景

众所周知，大多数肝细胞癌（HCC）患者死于转移，但协调转移的潜在机制仍知之甚少。目前的证据表明，METTL3 介导的 m6A 甲基化修饰的失调与癌症的进展密切相关。STAT3 是一种致癌转录因子，据报道在 HCC 的发生和发展中起核心作用。然而，METTL3 和 STAT3 在 HCC 转移中的关系尚不清楚。

方法

通过在线工具 GEPIA 和 Kaplan-Meier Plotter 评估 METTL3 表达与 HCC 患者生存的关系。Western blot、组织微阵列（TMA）和免疫组织化学（IHC）染色用于评估 HCC 细胞系和转移性和非转移性组织中 METTL3 和 STAT3 的表达水平。利用甲基化 RNA 免疫沉淀（MeRIP）、MeRIP 测序（MeRIP-seq）、qRT-PCR、RNA 免疫沉淀（RIP）、Western blot 和荧光素酶报告基因检测来阐明 METTL3 调节 STAT3 表达的机制。免疫荧光染色、Western blot、qRT-PCR、免疫共沉淀（Co-IP）、IHC 染色、TMA 和染色质免疫沉淀（ChIP）检测用于研究 STAT3 调节 METTL3 定位的机制。细胞活力、划痕愈合和 Transwell 检测以及原位异种移植模型用于评估 METTL3-STAT3 反馈回路在体外和体内促进 HCC 转移的作用。

结果

METTL3 和 STAT3 在高转移性 HCC 细胞和组织中均大量表达。此外，在 HCC 组织中发现 STAT3 的表达与 METTL3 的表达呈正相关。机制上，METTL3 可以诱导 STAT3 mRNA 的 m6A 修饰，然后通过与翻译起始机制相互作用促进 m6A 包含的 STAT3 mRNA 的翻译。相反，STAT3 通过转录上调甲基转移酶复合物的重要成员 WTAP 促进 METTL3 的核定位，并促进 METTL3 的甲基转移酶功能。METTL3 和 STAT3 形成正反馈回路，加速 HCC 在体外和体内的转移。