N -methyladenosine-modified CircSTX6 Promotes Hepatocellular Carcinoma Progression by Regulating the HNRNPD/ATF3 Axis and Encoding a 144 Amino Acid Polypeptide

Overview

Journal Clin Transl Med

Publisher Wiley

Specialty General Medicine

Date 2023 Oct 25

PMID 37877357

Authors

Jiahua Lu

Junnan Ru

Yunhao Chen

Zhenan Ling

Hanqing Liu

Bo Ding

Yifan Jiang

Jun Ma

Deguo Zhang

Jiangzhen Ge

Yu Li

Fei Sun

Diyu Chen

Shusen Zheng

Jian Wu

Affiliations

Soon will be listed here.

Abstract

Background: Circular RNAs (circRNAs) play a significant role in the initiation and progression of various cancers, including hepatocellular carcinoma (HCC). Circular syntaxin 6 (circSTX6, also known as hsa_circ_0007905) has been identified as a microRNA (miRNA) sponge in pancreatic adenocarcinoma. However, its full range of functions in terms of protein scaffold and translation remain largely unexplored in the context of HCC.

Methods: The expression of circSTX6 and its encoded protein was examined in HCC tumour tissues. N -methyladenosine (m A) on circSTX6 was verified and quantified by methylated RNA immunoprecipitation (Me-RIP), RIP and dual luciferase reporter assays. The biological functions of circSTX6 and its encoded protein in HCC were clarified by in vitro and in vivo experiments. Mechanistically, the interaction between circSTX6 and heterogeneous nuclear ribonucleoprotein D (HNRNPD) was investigated by RNA pull-down, RIP and fluorescence in situ hybridization (FISH)/IF. The regulatory effects of circSTX6 and HNRNPD on activating transcription factor 3 (ATF3) mRNA were determined by mRNA stability and RIP assays. Furthermore, the presence of circSTX6-encoded protein was verified by mass spectrometry.

Results: CircSTX6 and its encoded 144 amino acid polypeptide, circSTX6-144aa, were highly expressed in HCC tumour tissues and served as independent risk factors for overall survival in HCC patients. The expression of circSTX6 was regulated by METTL14 in an m A-dependent manner. Functionally, circSTX6 accelerated HCC proliferation and tumourigenicity and reinforced tumour metastasis in vitro and in vivo. Mechanistically, circSTX6 acted as a sponge for HNRNPD protein, facilitating its binding to ATF3 mRNA, consequently promoting ATF3 mRNA decay. Meanwhile, circSTX6-144aa promoted HCC proliferation, migration and invasion independent of circSTX6 itself.

Conclusion: Collectively, our study reveals that m A-modified circSTX6 drives malignancy in HCC through the HNRNPD/ATF3 axis, while its encoded circSTX6-144aa contributes to HCC progression independent of circSTX6. CirSTX6 and its encoded protein hold promise as potential biomarkers and therapeutic targets in HCC.

Citing Articles

Mitigating effects of Jiawei Chaihu Shugan decoction on necroptosis and inflammation of hippocampal neurons in epileptic mice.

Wang Q, Qin B, Yu H, Zeng J, Fan J, Wu Q Sci Rep. 2025; 15(1):4649.

PMID: 39920301 PMC: 11805973. DOI: 10.1038/s41598-025-89275-8.

The Mechanism and Latest Progress of m6A Methylation in the Progression of Pancreatic Cancer.

Liu Z, Ma P, He Y, Zhang Y, Mou Z, Fang T Int J Biol Sci. 2025; 21(3):1187-1201.

PMID: 39897038 PMC: 11781182. DOI: 10.7150/ijbs.104407.

m6A-modified circSTX6 as a key regulator of cervical cancer malignancy via SPI1 and IL6/JAK2/STAT3 pathways.

Han X, Xia L, Wu Y, Chen X, Wu X Oncogene. 2025; .

PMID: 39814850 DOI: 10.1038/s41388-024-03260-5.

RNA-binding protein HNRNPD promotes chondrocyte senescence and osteoarthritis progression through upregulating FOXM1.

Jiang H, Zhang Y, Hu G, Ji P, Ming J, Li Y Commun Biol. 2024; 7(1):1695.

PMID: 39719453 PMC: 11668876. DOI: 10.1038/s42003-024-07407-8.

Downregulation of circSTX6 suppresses tumor progression while facilitating radiosensitivity in cervical squamous cell carcinoma.

Hu X, Xing F, Yin Y, Zhao N, Xing L, Dong G Heliyon. 2024; 10(21):e39262.

PMID: 39524808 PMC: 11544058. DOI: 10.1016/j.heliyon.2024.e39262.

References

Park O, Ha H, Lee Y, Boo S, Kwon D, Song H . Endoribonucleolytic Cleavage of mA-Containing RNAs by RNase P/MRP Complex. Mol Cell. 2019; 74(3):494-507.e8. DOI: 10.1016/j.molcel.2019.02.034. View

Liu P, Zhong Q, Song Y, Guo D, Ma D, Chen B . Long noncoding RNA Linc01612 represses hepatocellular carcinoma progression by regulating miR-494/ATF3/p53 axis and promoting ubiquitination of YBX1. Int J Biol Sci. 2022; 18(7):2932-2948. PMC: 9066125. DOI: 10.7150/ijbs.69514. View

Zhang C, Gao Y, Du C, Markowitz G, Fu J, Zhang Z . Hepatitis B-Induced IL8 Promotes Hepatocellular Carcinoma Venous Metastasis and Intrahepatic Treg Accumulation. Cancer Res. 2021; 81(9):2386-2398. DOI: 10.1158/0008-5472.CAN-20-3453. View

Zaccara S, Ries R, Jaffrey S . Reading, writing and erasing mRNA methylation. Nat Rev Mol Cell Biol. 2019; 20(10):608-624. DOI: 10.1038/s41580-019-0168-5. View

Xu J, Wan Z, Tang M, Lin Z, Jiang S, Ji L . N-methyladenosine-modified CircRNA-SORE sustains sorafenib resistance in hepatocellular carcinoma by regulating β-catenin signaling. Mol Cancer. 2020; 19(1):163. PMC: 7681956. DOI: 10.1186/s12943-020-01281-8. View

Gong X, Tian M, Cao N, Yang P, Xu Z, Zheng S . Circular RNA circEsyt2 regulates vascular smooth muscle cell remodeling via splicing regulation. J Clin Invest. 2021; 131(24). PMC: 8670847. DOI: 10.1172/JCI147031. View

Hu Z, Zhou S, Li J, Zhou Z, Wang P, Xin H . Circular RNA Sequencing Identifies CircASAP1 as a Key Regulator in Hepatocellular Carcinoma Metastasis. Hepatology. 2019; 72(3):906-922. DOI: 10.1002/hep.31068. View

Chen X, Han P, Zhou T, Guo X, Song X, Li Y . circRNADb: A comprehensive database for human circular RNAs with protein-coding annotations. Sci Rep. 2016; 6:34985. PMC: 5057092. DOI: 10.1038/srep34985. View

Xuan J, Sun W, Lin P, Zhou K, Liu S, Zheng L . RMBase v2.0: deciphering the map of RNA modifications from epitranscriptome sequencing data. Nucleic Acids Res. 2017; 46(D1):D327-D334. PMC: 5753293. DOI: 10.1093/nar/gkx934. View

10.

Wang Y, Chen D, Xie H, Jia M, Sun X, Peng F . AUF1 protects against ferroptosis to alleviate sepsis-induced acute lung injury by regulating NRF2 and ATF3. Cell Mol Life Sci. 2022; 79(5):228. PMC: 11072094. DOI: 10.1007/s00018-022-04248-8. View

11.

Wagner B, DeMaria C, Sun Y, WILSON G, Brewer G . Structure and genomic organization of the human AUF1 gene: alternative pre-mRNA splicing generates four protein isoforms. Genomics. 1998; 48(2):195-202. DOI: 10.1006/geno.1997.5142. View

12.

Liang L, Zhu Y, Li J, Zeng J, Wu L . ALKBH5-mediated m6A modification of circCCDC134 facilitates cervical cancer metastasis by enhancing HIF1A transcription. J Exp Clin Cancer Res. 2022; 41(1):261. PMC: 9413927. DOI: 10.1186/s13046-022-02462-7. View

13.

Huang H, Weng H, Chen J . mA Modification in Coding and Non-coding RNAs: Roles and Therapeutic Implications in Cancer. Cancer Cell. 2020; 37(3):270-288. PMC: 7141420. DOI: 10.1016/j.ccell.2020.02.004. View

14.

Zhang W, Wagner B, Ehrenman K, Schaefer A, DeMaria C, Crater D . Purification, characterization, and cDNA cloning of an AU-rich element RNA-binding protein, AUF1. Mol Cell Biol. 1993; 13(12):7652-65. PMC: 364837. DOI: 10.1128/mcb.13.12.7652-7665.1993. View

15.

DeMaria C, Sun Y, Long L, Wagner B, Brewer G . Structural determinants in AUF1 required for high affinity binding to A + U-rich elements. J Biol Chem. 1997; 272(44):27635-43. DOI: 10.1074/jbc.272.44.27635. View

16.

Chen L . The biogenesis and emerging roles of circular RNAs. Nat Rev Mol Cell Biol. 2016; 17(4):205-11. DOI: 10.1038/nrm.2015.32. View

17.

Llovet J, Pinyol R, Kelley R, El-Khoueiry A, Reeves H, Wang X . Molecular pathogenesis and systemic therapies for hepatocellular carcinoma. Nat Cancer. 2022; 3(4):386-401. PMC: 9060366. DOI: 10.1038/s43018-022-00357-2. View

18.

Pan Y, Chen H, Kilberg M . Interaction of RNA-binding proteins HuR and AUF1 with the human ATF3 mRNA 3'-untranslated region regulates its amino acid limitation-induced stabilization. J Biol Chem. 2005; 280(41):34609-16. PMC: 3600371. DOI: 10.1074/jbc.M507802200. View

19.

Tsitsipatis D, Grammatikakis I, Driscoll R, Yang X, Abdelmohsen K, Harris S . AUF1 ligand circPCNX reduces cell proliferation by competing with p21 mRNA to increase p21 production. Nucleic Acids Res. 2021; 49(3):1631-1646. PMC: 7897478. DOI: 10.1093/nar/gkaa1246. View

20.

Zhou C, Molinie B, Daneshvar K, Pondick J, Wang J, Van Wittenberghe N . Genome-Wide Maps of m6A circRNAs Identify Widespread and Cell-Type-Specific Methylation Patterns that Are Distinct from mRNAs. Cell Rep. 2017; 20(9):2262-2276. PMC: 5705222. DOI: 10.1016/j.celrep.2017.08.027. View