A Positive Feedback Loop of lncRNA HOXD-AS2 and SMYD3 Facilitates Hepatocellular Carcinoma Progression via the MEK/ERK Pathway

Jin Sun; Yingnan Li; Mengjiao Shi; Hongwei Tian; Jun Li; Kai Zhu; Ying Guo; Yanhua Mu; Jing Geng; Zongfang Li

doi:10.2147/JHC.S416946

A Positive Feedback Loop of lncRNA HOXD-AS2 and SMYD3 Facilitates Hepatocellular Carcinoma Progression via the MEK/ERK Pathway

J Hepatocell Carcinoma. 2023 Jul 27:10:1237-1256. doi: 10.2147/JHC.S416946. eCollection 2023.

Authors

Jin Sun^{1

2

3}, Yingnan Li^{1

2

3}, Mengjiao Shi^{1

2}, Hongwei Tian^{1

2

3}, Jun Li^{1

2

3}, Kai Zhu^{1

2

3}, Ying Guo^{1

2

3}, Yanhua Mu^{1

2

3}, Jing Geng^{1

2

3}, Zongfang Li^{1

2

3

4}

Affiliations

¹ National-Local Joint Engineering Research Center of Biodiagnostics and Biotherapy, Xi'an Jiaotong University, Xi'an, Shaanxi, People's Republic of China.
² Shaanxi Provincial Clinical Research Center for Hepatic & Splenic Diseases, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, People's Republic of China.
³ Center for Tumor and Immunology, the Precision Medical Institute, Xi'an Jiaotong University, Xi'an, Shaanxi, People's Republic of China.
⁴ Department of Geriatric General Surgery, the Second Affiliated Hospital of Xi' an Jiaotong University, Xi'an, Shaanxi, People's Republic of China.

Abstract

Purpose: HOX cluster-embedded long noncoding RNAs (HOX-lncRNAs) have been shown to be tightly related to hepatocellular carcinoma (HCC). However, the potential biological roles and underlying molecular mechanism of HOX-lncRNAs in HCC largely remains to be elucidated.

Methods: The expression signature of eighteen HOX-lncRNAs in HCC cell lines were measured by qRT-PCR. HOXD-AS2 expression and its clinical significance in HCC was investigated by bioinformatics analysis utilizing the TCGA data. Subcellular localization of HOXD-AS2 in HCC cells was observed by RNA-FISH. Loss‑of‑function experiments in vitro and in vivo were conducted to probe the roles of HOXD-AS2 in HCC. Potential HOXD-AS2-controlled genes and signaling pathways were revealed by RNA-seq. Rescue experiments were performed to validate that SMYD3 mediates HOXD-AS2 promoting HCC progression. The positive feedback loop of HOXD-AS2 and SMYD3 was identified by luciferase reporter assay and ChIP-qPCR.

Results: HOXD-AS2 was dramatically elevated in HCC, and its up-regulation exhibited a positive association with aggressive clinical features (T stage, pathologic stage, histologic grade, AFP level, and vascular invasion) and unfavorable prognosis of HCC patients. HOXD-AS2 was distributed both in the nucleus and the cytoplasm of HCC cells. Knockdown of HOXD-AS2 restrained the proliferation, migration, invasion of HCC cells in vitro, as well as tumor growth in subcutaneous mouse model. Transcriptome analysis demonstrated that SMYD3 expression and activity of MEK/ERK pathway were impaired by silencing HOXD-AS2 in HCC cells. Rescue experiments revealed that SMYD3 as downstream target mediated oncogenic functions of HOXD-AS2 in HCC cells through altering the expression of cyclin B1, cyclin E1, MMP2 as well as the activity of MEK/ERK pathway. Additionally, HOXD-AS2 was uncovered to be positively regulated at transcriptional level by its downstream gene of SMYD3.

Conclusion: HOXD-AS2, a novel oncogenic HOX-lncRNA, facilitates HCC progression by forming a positive feedback loop with SMYD3 and activating the MEK/ERK pathway.

Keywords: HOXD-AS2; MEK/ERK signaling; SMYD3; hepatocellular carcinoma.

Grants and funding

This study was supported by the National Natural Science Foundation of China (81502136 and 81802456), the Natural Science Foundation Research Program of Shaanxi Province of China (2023-JC-YB-750), the Science Research Foundation of the Second Affiliated Hospital of Xi’an Jiaotong University (2020YJ(ZYTS)546-09 and 2020YJ(ZYTS)645).