Long non-coding RNA TMPO-AS1 facilitates the progression of colorectal cancer cells via sponging miR-98-5p to upregulate BCAT1 expression.

BACKGROUND AND AIM: Colorectal cancer, as a common malignant carcinoma in the gastrointestinal tract, has a high mortality globally. However, the specific molecular mechanisms of long non-coding RNA (lncRNA) thymopoietin antisense transcript 1 (TMPO-AS1) in colorectal cancer were unclear. METHODS: We tested the expression level of TMPO-AS1 via qRT-PCR in colorectal cancer cells, while the protein levels of branched chain amino acid transaminase 1 (BCAT1) and the stemness-related proteins were evaluated by western blot analysis. Colony formation, EdU staining, TUNEL, flow cytometry, and sphere formation assays were to assess the biological behaviors of colorectal cancer cells. Then, luciferase reporter, RIP, and RNA pull down assay were applied for confirming the combination between microRNA-98-5p (miR-98-5p) and TMPO-AS1/BCAT1. RESULTS: TMPO-AS1 was aberrantly expressed at high levels in colorectal cancer cells. Silenced TMPO-AS1 restrained cell proliferation and stemness and promoted apoptosis oppositely, while overexpressing TMPO-AS1 exerted the adverse effects. Furthermore, miR-98-5p was proven to a target of TMPO-AS1 inhibit cell progression in colorectal cancer. Additionally, BCAT1 was proved to enhance cell progression as the target of miR-98-5p, and it offset the effect of silenced TMPO-AS1 on colorectal cancer cells. CONCLUSION: TMPO-AS1 promotes the progression of colorectal cancer cells via sponging miR-98-5p to upregulate BCAT1 expression.

miR­4306 inhibits the malignant behaviors of colorectal cancer by regulating lncRNA FoxD2­AS1.

MicroRNA (miR)­4306 and FoxD2­adjacent opposite strand RNA 1 (FOXD2­AS1) are cancer­related genes involved in tumor progression. However, the potential functional roles of miR­4306 and FoxD2­AS1 in colorectal cancer (CRC) development remain unknown. The present study aimed to investigate the biological functions and the molecular mechanisms of miR­4306 and FoxD2­AS1 in CRC. Reverse transcription­quantitative PCR analysis was performed to determine the expression levels of FoxD2­AS1 and miR­4306 in CRC tissues and cell lines. Functional experiments, including Cell Counting Kit­8, colony formation, cell cycle assays and western blotting, were conducted to examine the effects of FoxD2­AS1 and miR­4306 on the malignant behaviors of CRC cells. In addition, the relationship between FoxD2­AS1 and miR­4306 was assessed using a dual­luciferase reporter assay and Pearson's correlation analysis. Compared with normal samples and cells, FoxD2­AS1 expression was increased and miR­4306 expression was decreased in CRC tissues and cells. Functional experiments demonstrated that silencing FoxD2­AS1 inhibited proliferation and induced cell arrest at G0/G1 phase in CRC cells, while the overexpression of FoxD2­AS1 showed opposite results. Ki­67 and proliferating cell nuclear antigen expression levels were decreased after transfection with small interfering RNA FoxD2­AS1, but were increased after transfection with FoxD2­AS1 overexpression plasmid. Furthermore, investigations into the underling mechanism revealed that FoxD2­AS1 functioned as a molecular sponge of miR­4306. The inhibitory effects of FoxD2­AS1 silencing on CRC progression were reversed by miR­4306 knockdown. Collectively, the present study demonstrated that FoxD2­AS1 functioned as an oncogene in CRC progression, and that miR­4306 could inhibit the malignant behaviors of CRC by regulating FoxD2­AS1. Thus, the current study provided a promising therapeutic target for CRC treatment.

LINC00963 affects the development of colorectal cancer via MiR-532-3p/HMGA2 axis.

BACKGROUND: LINC00963 is high-expressed in various carcinomas, but its expression and function in colorectal cancer (CRC) have not been explored. This study explored the role and mechanism of LINC00963 in CRC. METHODS: The expression of LINC00963 in CRC and its relationship with prognosis were examined by starBase and survival analysis. The effects of LINC00963, miR-532-3p and HMGA2 on the biological characteristics and EMT-related genes of CRC cells were studied by RT-qPCR, CCK-8, clone formation experiments, flow cytometry, scratch test, Transwell, and Western blot. Xenograft assay and immunohistochemistry were performed to verify the effect of LINC00963 on tumor growth. The correlation among LINC00963, miR-532-3p, and HMGA2 was analyzed by bioinformatics analysis, luciferase assay, and Pearson test. RESULTS: LINC00963 was high-expressed in CRC, and this was associated with poor prognosis of CRC. Silencing LINC00963 inhibited the activity, proliferation, migration, and invasion of CRC cells, MMP-3 and MMP-9 expressions, moreover, it also blocked cell cycle progression, and inhibited tumor growth and Ki67 expression. However, overexpression of LINC00963 showed the opposite effects to silencing LINC00963. LINC00963 targeted miR-532-3p to regulate HMGA2 expression. Down-regulation of miR-532-3p promoted cell proliferation, migration and invasion, and expressions of MMP-3 and MMP-9, and knockdown of HMGA2 reversed the effect of miR-532-3p inhibitor. Up-regulation of miR-532-3p inhibited the biological functions of CRC cells, and overexpression of HMGA2 reversed the miR-532-3p mimic effect. CONCLUSION: LINC00963 affects the development of CRC through the miR-532-3p/HMGA2 axis.

Dkk1 inhibits malignant transformation induced by Bmi1 via the ß-catenin signaling axis in WB-F344 oval cells.

Dickkopf-1 (Dkk1) is an inhibitor of Wnt signaling involved in cancer cell proliferation, apoptosis, and migration and angiogenesis. It was previously reported that B cell-specific Moloney mouse leukemia virus integration site 1 (Bmi1) activates the Wnt pathway by inhibiting the expression of DKK1 in breast cancer cell lines and 293T cells. Bmi1 and DKK1 are highly expressed in liver samples taken by biopsy from patients with hepatitis B virus-related hepatocellular carcinoma (HCC), but the effect of both Bmi1 and DKK1 on the carcinogenesis of adult hepatic stem cells (oval cells) has not previously been reported. In this study, we used WB-F344 cells to explore the function and regulation of Dkk1 upon Bmi1 treatment. Overexpression of Dkk1 repressed differentiation, proliferation, and migration induced by Bmi1 but promoted the apoptosis of hepatic WB-F344 oval cells. In addition, Dkk1 reduced the enhancement of ß-catenin levels induced by Bmi1. Finally, we used transcriptome sequencing to perform a comprehensive evaluation of the transcriptome-related changes in WB-F344 oval cells induced by Dkk1 and Bmi1. These results may provide evidence for future studies of the pathogenesis of HCC and the design of possible therapies.

Mitofusin-2 (Mfn-2) Might Have Anti-Cancer Effect through Interaction with Transcriptional Factor SP1 and Consequent Regulation on Phosphatidylinositol Transfer Protein 3 (PITPNM3) Expression.

BACKGROUND The aim of this study was to explore the influence of mitofusin-2 (Mfn-2) on phosphatidylinositol transfer protein 3 (PITPNM3) and tumor growth and the potential mechanism behind the regulation of Mfn-2 on PITPNM3 in hepatic carcinoma cell line SMMC-7721. MATERIAL AND METHODS We obtained promoter sequence of PITPNM3 gene from University of Santa Cruz (UCSC) genomic database, and we predict transcriptional factor of PITPNM3 genes by JASPAR database. Target transcription factor was determined by comparison of binding sites number for promoter. SMMC-7721 cells were transfected with expression plasmid containing Mfn-2, transcription factor gene and PITPNM3. The cells transfected with empty vector were used as control. Real-time polymerase chain reaction was used to determine the mRNA level of target genes. Co-immunoprecipitation (Co-IP) assay was used to determine the interaction between Mfn-2 and target transcription factor. Chromatin immunoprecipitation assay (ChIP) assay was used to determine the binding of transcription factor with PITPNM3 promoter. Tumorigenicity assay was used to compare the effect of Mfn-2, SP1, and PITPNM3 on tumor development. RESULTS SP1 was selected as the target transcriptional factor. In the Co-IP assay, Mfn-2 was shown to interact with SP1. In the ChIP assay Mfn-2 transfection resulted in decreased binding number of SP1 with PITPNM3 promoter. Furthermore, PITPNM3 mRNA levels were significantly increased in SMMC-7721 cells transfected with SP1 but were decreased after transfection with Mfn-2. In nude mice, PITPNM3 and SP1 upregulation lead to larger tumor lump and conversely Mfn-2 upregulation lead to smaller tumor lump. CONCLUSIONS Mfn-2 could suppress expression of PITPNM3 through interaction with transcription factor SP1; Mfn-2 may have anti-tumor activity; SP1 and PITPNM3 may promote tumor development.