Dataset Information

COPB2 gene silencing inhibits colorectal cancer cell proliferation and induces apoptosis via the JNK/c-Jun signaling pathway.

ABSTRACT:

Objectives

Colorectal cancer (CRC) is one of the most common malignant human tumors. It is associated with high morbidity and mortality rates. In recent years, tumor gene therapy has emerged as a promising new approach for colorectal cancer therapy. Herein, we identify and analyze the role of COPB2 (coatomer protein complex, subunit beta 2) in proliferation and apoptosis of CRC cells.

Methods

To investigate the role of COPB2 in the proliferation and apoptosis of CRC cells, a shCOPB2 vector and a shCtrl vector were constructed for transfection into RKO and HCT116 cells. Cells proliferation was subsequently measured via cell counting kit-8 (CCK8) assay and Celigo cell counting assay. Apoptosis was measured via flow cytometry. The activity level of Caspase 3/7 was measured. Finally, the level of several JNK/c-Jun apoptosis pathway-related proteins were measured to characterize the mechanism of apoptosis.

Results

Our results showed that the proliferation rate was decreased and the apoptosis rate was increased in shCOPB2-treated RKO and HCT116 cells compared to those in controls. After the silencing of COPB2, JNK/c-Jun signal pathway activation was increased, the expression levels of apoptosis pathway-related proteins, such as Bad, p53 and Caspase 3, were also increased.

Conclusion

COPB2 gene silencing can inhibit RKO and HCT116 cells proliferation and induce apoptosis via the JNK/c-Jun signaling pathway.

SUBMITTER: Wang Y

PROVIDER: S-EPMC7676692 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Publications

COPB2 gene silencing inhibits colorectal cancer cell proliferation and induces apoptosis via the JNK/c-Jun signaling pathway.

Wang Yan Y Xie Guangmei G Li Min M Du Juan J Wang Min M

PloS one 20201119 11

<h4>Objectives</h4>Colorectal cancer (CRC) is one of the most common malignant human tumors. It is associated with high morbidity and mortality rates. In recent years, tumor gene therapy has emerged as a promising new approach for colorectal cancer therapy. Herein, we identify and analyze the role of COPB2 (coatomer protein complex, subunit beta 2) in proliferation and apoptosis of CRC cells.<h4>Methods</h4>To investigate the role of COPB2 in the proliferation and apoptosis of CRC cells, a shCOP ...[more]

PMID: 33211699

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Project description:Emerging studies have reported that coatomer protein complex subunit β2 (COPB2) is overexpressed in several types of malignant tumor; however, to the best of our knowledge, no studies regarding COPB2 in gastric cancer have been published thus far. Therefore, the present study aimed to determine the significance and function of COPB2 in gastric cancer. COPB2 expression in gastric cancer cell lines was measured using reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) analysis. In addition, lentivirus‑short hairpin RNA (shRNA) COPB2 (Lv‑shCOPB2) was generated and used to infect BGC‑823 cells to analyze the effects of COPB2 on the cancerous phenotype. The effects of shRNA‑mediated COPB2 knockdown on cell proliferation were detected using MTT, 5‑bromo‑2‑deoxyuridine and colony formation assays. In addition, the effects of COPB2 knockdown on apoptosis were analyzed by flow cytometry. Nude mice and fluorescence imaging were used to characterize the regulation of tumor growth in vivo, and qPCR and immunohistochemistry were subsequently conducted to analyze COPB2 expression in xenograft tumor tissues. Furthermore, a receptor tyrosine kinase (RTK) signaling pathway antibody array was used to explore the relevant molecular mechanisms underlying the effects of COPB2 knockdown. The results revealed that COPB2 mRNA was abundantly overexpressed in gastric cancer cell lines, whereas knockdown of COPB2 significantly inhibited cell growth and colony formation ability, and led to increased cell apoptosis in vitro. The tumorigenicity assay revealed that knockdown of COPB2 reduced tumor growth in nude mice, and fluorescence imaging indicated that the total radiant efficiency of mice in the Lv‑shCOPB2‑infected group was markedly reduced compared with the mice in the Lv‑shRNA control‑infected group in vivo. The antibody array assay revealed that the levels of phosphorylation in 23 target RTKs were significantly reduced: In conclusion, COPB2 was highly expressed in gastric cancer cell lines, and knockdown suppressed colony formation and promoted cell apoptosis via inhibiting the RTK signaling and its downstream signaling cascade molecules. Therefore, COPB2 may present a valuable target for gene silencing strategy in gastric cancer.

Project description:ObjectiveTo investigate the effect of silencing GDP dissociation inhibitor 2 (GDI2) on colorectal cancer development and possible mechanisms based on transcriptomic analysis.MethodsThe differences in the expression levels of GDI2 in normal colorectal tissues and tumor tissues of colorectal cancer (CRC) patients were detected. The correlation of GDI2 expression levels with survival and clinical characteristics of CRC patients was analyzed. The effects of GDI2 expression levels on the biological functions of CRC cells were examined by CCK-8 assay, plate clone formation assay, wound healing assay, and Transwell assay. The effect of GDI2 on the proliferation and growth of xenograft tumors was investigated by a xenograft tumor model of CRC in nude mice. Based on transcriptomics, we explored the possible mechanisms and associated pathways of the effect of silencing GDI2 on CRC cells. Cellular experiments and Western blot assays were performed to verify the potential mechanisms and related pathway of GDI2 action on CRC.ResultsThe expression levels of GDI2 in CRC tissues and cells were higher than those in normal tissues and cells. The expression level of GDI2 correlated with clinical characteristics such as lymphatic metastasis, tumor stage, tumor volume, and lymphocyte count. Silencing of GDI2 reduced the proliferative activity and migration and invasion ability of CRC cells, as well as inhibited the proliferation of CRC xenograft tumors. The differentially expressed genes were significantly enriched in biological processes such as cell cycle arrest and the p53 signaling pathway after GDI2 silencing. The percentage of G0/G1 phase cells in CRC cells was increased after silencing GDI2 as verified by flow cytometry. RAB5A was highly associated with the p53 pathway and could interact with TP53 via the ZFYVE20 protein. The mutual binding between GDI2 protein and RAB5A protein was verified by immunoprecipitation assay. Silencing GDI2 while overexpressing RAB5A reversed the reduced proliferation, migration, and invasion ability as well as cell cycle arrest of CRC cells. Meanwhile, the addition of p53 signaling pathway inhibitor Pifithrin-α (PFT-α) also reversed the biological effects of silencing GDI2 on CRC cells. The p-p21 and p-p53 protein expression levels were significantly greater in the sh-GDI2 group than in the sh-NC group. However, the p-p21 and p-p53 protein expression levels were reduced after silencing GDI2 while overexpressing RAB5A.ConclusionSilencing GDI2 activates the p53 signaling pathway by regulating RAB5A expression levels, which in turn induces cell cycle arrest and ultimately affects the proliferative activity, migration, and invasive ability of CRC cells.

Dataset Information

COPB2 gene silencing inhibits colorectal cancer cell proliferation and induces apoptosis via the JNK/c-Jun signaling pathway.

Objectives

Methods

Results

Conclusion

Publications

COPB2 gene silencing inhibits colorectal cancer cell proliferation and induces apoptosis via the JNK/c-Jun signaling pathway.

Similar Datasets

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