Zhang Zhen, Wu Honglei, Chen Zhaosheng, Li Guangchun, Guo Jianqiang
The Second Hospital of Shandong University, Jinan 250033, China.
J Oncol. 2023 Mar 14;2023:1298312. doi: 10.1155/2023/1298312. eCollection 2023.
The long noncoding RNA (lncRNA) gene PTOV1-AS2 is a potentially oncogenic lncRNA gene. However, its role and regulatory mechanism in the occurrence and development of colon cancer are still unclear. In this study, the lncRNA PTOV1-AS2 was used as a starting point to investigate the role of competing endogenous RNA (ceRNA) regulatory mechanisms in colon cancer.
The expression of lncRNA PTOV1-AS2 mRNA in colon cancer tissues and cell lines was measured by quantitative real-time polymerase chain reaction (qRT-PCR) and screened for differential expression in cells. We examined the effects of lncRNA PTOV1-AS2 overexpression or downregulation of its expression on various cellular processes in HCT116 and SW620 cells after the transfection with an overexpression construct or PTOV1-AS2p-specific shRNA, respectively. In particular, we examined the effects on cell proliferation, migration, and invasion using the cell counting kit-8 CCK-8 assay and Transwell migration and invasion assays, respectively. In addition, the binding targets of lncRNA PTOV1-AS2/miR-145-5p and miR-145-5p/FSCN1 were predicted using various bioinformatics tools and validated by a dual luciferase assay. We also examined the effect of the lncRNA PTOV1-AS2/miR-145-5p axis on FSCN1 expression by qRT-PCR analysis. Furthermore, we investigated the effect of the PTOV1-AS2/miR-145-5p/FSCN1 axis on the biological function of colon cancer cells using an colon cancer cell model with reduced expression of PTOV1-AS2 and simultaneous transfection of a miR-145-5p inhibitor or FSCN1 vector. Additionally, we established a colon cancer xenograft tumor nude mouse model and used it to investigate the effect of locally injected lncRNA PTOV1-AS2 vector on the tumor growth and survival status of tumor-bearing mice.
We found that PTOV1-AS2 was highly expressed in colon cancer, which was associated with worse survival. High expression of PTOV1-AS2 promoted cell proliferation, migration, and invasion, while low expression of PTOV1-AS2 inhibited these processes in HCT116 and SW620 cells. The microRNA miR-145-5p was found to bind to the 3'-UTR region of both PTOV1-AS2 and FSCN1. In addition, miR-145-5p decreased the protein expression of its target gene FSCN1 and reduced the PTOV1-AS2-induced expression of FSCN1 in colon cancer cell lines. Also, silencing miR-145-5p or enhancing FSCN1 expression could partially restore the inhibition of cell proliferation, migration, invasion, and the tumorigenic capacity caused by silencing the expression of PTOV1-AS2 and .
PTOV1-AS2 promotes colon cancer progression by "sponging" miR-145-5p to upregulate FSCN1.
长链非编码RNA（lncRNA）基因PTOV1-AS2是一种潜在的致癌lncRNA基因。然而，其在结肠癌发生发展中的作用及调控机制仍不清楚。本研究以lncRNA PTOV1-AS2为切入点，探讨竞争性内源RNA（ceRNA）调控机制在结肠癌中的作用。
采用定量实时聚合酶链反应（qRT-PCR）检测lncRNA PTOV1-AS2 mRNA在结肠癌组织及细胞系中的表达，并筛选细胞中的差异表达情况。分别用过表达构建体或PTOV1-AS2特异性短发夹RNA（shRNA）转染HCT116和SW620细胞后，检测lncRNA PTOV1-AS2过表达或表达下调对细胞各种生物学过程的影响。具体而言，分别使用细胞计数试剂盒-8（CCK-8）法和Transwell迁移及侵袭实验检测对细胞增殖、迁移和侵袭的影响。此外，利用多种生物信息学工具预测lncRNA PTOV1-AS2/miR-145-5p和miR-145-5p/FSCN1的结合靶点，并通过双荧光素酶实验进行验证。我们还通过qRT-PCR分析检测lncRNA PTOV1-AS2/miR-145-5p轴对FSCN1表达的影响。此外，我们利用PTOV1-AS2表达降低且同时转染miR-145-5p抑制剂或FSCN1载体的结肠癌细胞模型，研究PTOV1-AS2/miR-145-5p/FSCN1轴对结肠癌细胞生物学功能的影响。另外，建立结肠癌异种移植瘤裸鼠模型，用于研究局部注射lncRNA PTOV1-AS2载体对荷瘤小鼠肿瘤生长及生存状态的影响。
我们发现PTOV1-AS2在结肠癌中高表达，这与较差的生存率相关。PTOV1-AS2的高表达促进了HCT116和SW620细胞的增殖、迁移和侵袭，而PTOV1-AS2的低表达则抑制了这些过程。发现微小RNA miR-145-5p与PTOV1-AS2和FSCN1的3'-非翻译区（3'-UTR）区域结合。此外，miR-145-5p降低了其靶基因FSCN1的蛋白表达，并降低了结肠癌细胞系中PTOV1-AS2诱导的FSCN1表达。而且，沉默miR-145-5p或增强FSCN1表达可部分恢复因沉默PTOV1-AS2表达而导致的细胞增殖、迁移、侵袭及致瘤能力的抑制。
PTOV1-AS2通过“吸附”miR-145-5p上调FSCN1，促进结肠癌进展。
