Research paperCircFOXO3 functions as a molecular sponge for miR-143-3p to promote the progression of gastric carcinoma via upregulating USP44
Introduction
Gastric carcinoma (GC) ranks among the five cancers with the highest morbidity and ranks third in terms of cancer-related deaths. GC also causes enormous economic problems for the global healthcare system (Nasrollahzadeh-Khakiani et al., 2017). The diagnosis of GC is often delayed, and clinical management can be inefficient. Therefore, most GC cases are initially evident as local diffusion or metastasis(Chen et al., 2012, Wu et al., 2017, Dan et al., 2018). However, little is known about early-stage molecular biomarkers of GC. It is imperative to improve our understanding regarding the essential mechanisms of GC initiation and progression for developing better targeted GC treatment.
Circular RNAs (circRNAs) are a type of noncoding RNA which characterized by covalently closed loops resulting from back splicing events(Ledford, 2013, Patop et al., 2019). Their functions are typically revealed by high-throughput sequencing and bioinformatic analysis. In the cell, circRNAs are primarily located in cytoplasm. And it has been found that circRNAs contain abundant miRNA competing binding sites indicating competing endogenous RNA function of circRNAs(Denzler et al., 2014, Thomson and Dinger, 2016). Furthermore, The crosslinking immunoprecipitation assays show that circRNAs are able to act as protein sponges due to the ability of interacting with abundant RBPs(Dudekula et al., 2016, Abdelmohsen et al., 2017). Therefore, circRNAs may be involved in crucial biological events(Chen and Yang, 2015, Barrett and Salzman, 2016, Beermann et al., 2016, Cortes-Lopez and Miura, 2016, Ebbesen et al., 2017, Li et al., 2018), including the formation and progression of cancers(Meng et al., 2017, Kristensen et al., 2018). CircRNAs were reported to play roles in the initiation and progression of GC. Zhang X et al found that circNRIP1 promotes GC via AKT1/mTOR pathway by targeting of miR-149-5p (Zhang et al., 2019b), Liu j et al demonstrated the molecular function of circ-SERPINE2 in the development of GC(Liu et al., 2019), Chen J et al found that circPVT1 is a proliferative factor and prognostic marker in GC(Chen et al., 2017), and Rong D et al revealed that circPSMC3 suppresses GC by sponging miR-296-5p(Rong et al., 2019). Recently, the molecular functions of circular RNA circFOXO3 in cancers have been investigated.
CircFOXO3 (hsa_circ_0006404) contains 1435 nucleotides and is derived from an exon of the FOXO3. CircFOXO3 involves in the progression of many cancers, including lung cancer(Zhang et al., 2018b), breast cancer(Lu, 2017), glioblastoma(Zhang et al., 2019a), prostate cancer(Shen et al., 2020), and esophageal squamous cell cancer(Xing et al., 2019). Nonetheless, the role of circFOXO3 in the progression of GC remains unelucidated, and neither are other molecular mechanisms of action of circFOXO3. The aim of this study was to address these issues.
Here, we hypothesized that circFOXO3 involves in the progression of GC and relates to the tumor growth and metastasis of GC. By performing a serial in vitro and vivo experiments. CircFOXO3 was shown to upregulate the expression of ubiquitin-specific peptidase 44 (USP44) in GC cell lines by interacting with microRNA (miRNA,miR)-143-3p. CircFOXO3 also enhanced GC cell proliferation and migration through the miR-143-3p-USP44 pathway and promoted tumor growth in vivo. These finding provide a new perspective regarding the function of circFOXO3 and highlight a potential target for studies directed at developing GC therapies.
Section snippets
Cell culture and transfection
All GC cell lines (GES1, AGS, MKN45, SNU5, MKN74, MGC803, NCI-N87) and 293 T cells were purchased from the American Type Culture Collection (Manassas, VA, USA). Those cells were cultured with Dulbecco’s Modified Eagle medium (DMEM) (Gibco) medium supplemented with 10% fetal bovine serum (FPS)(Gibco), and were cultivated according to the vendor’s recommendations at 37 °C in a humidified atmosphere containing 5% CO2 and 95% air. A circFOXO3-expressing plasmid - the human circ-FOXO3 cDNA
CircFOXO3 is upregulated in GC cells
Initially, we investigated the expression of circFOXO3 in GC cells. we hypothesized that circFOXO3 might be involved in the progression of GC. Subsequently, we quantified circFOXO3 levels in GC cell lines (Fig. 1A). The expression of circFOXO3 was found to be significantly upregulated in MGC803 cells and downregulated in NCI-N87 cells. Then, we tested the efficiency of circFOXO3- PcDNA transfection and its NC as well as si-circFOXO3 transfection and its NC in NCI-N87 cells and MGC803 cells (
Discussion
In the present study, we found that circFOXO3 is overexpressed in GC tumors and that it is involved in the progression of GC via the miR-143-3p-USP44 axis. We performed a serial in vitro and vivo experiments to determine the role of circFOXO3 in GC progression, and the molecular mechanisms underlying it. In the past few years, the biological functions and pathological roles of circRNA in the formation and progression of cancers were determined by performing high-throughput sequencing and
Conclusions
• The results collectively reveal the role of circFOXO3 in GC.
• We clarified the mechanisms by which circFOXO3 participates in GC progression in vivo and in vitro.
• Our findings highlight the crucial function of circRNAs in GC progression, and reveal a new potential therapeutic target in GC.
CRediT authorship contribution statement
Tian Xiang: Validation, Formal analysis, Visualization, Software, Writing - review & editing. Hong-sheng Jiang: Validation, Formal analysis, Visualization, Software, Writing - review & editing. Bi-tao Zhang: Resources, Writing - review & editing, Supervision, Data curation. Gao Liu: Conceptualization, Methodology, Software, Investigation, Writing - original draft, Resources, Supervision, Data curation, Writing - review & editing.
Declaration of Competing Interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgement
N.A.
Data availability statement
The data sets used and/or analyzed during the current study are available from the corresponding author on reasonable request.
Funding
This work was supported by the National Natural Science Foundation of China [grant number 81760540]. The sponsor had no role in study design; in the collection, analysis, and interpretation of data; in the writing of the report; and in the decision to submit the article for publication.
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