TRIM22 activates NF-κB signaling in glioblastoma by accelerating the degradation of IκBα

AbstractNF-κB signaling plays a critical role in tumor growth and treatment resistance in GBM as in many other cancers. However, the molecular mechanisms underlying high, constitutive NF-κB activity in GBM remains to be elucidated. Here, we screened a panel of tripartite motif (TRIM) family proteins and identified TRIM22 as a potential activator of NF-κB using an NF-κB driven luciferase reporter construct in GBM cell lines. Knockout of TRIM22 using Cas9-sgRNAs led to reduced GBM cell proliferation, while TRIM22 overexpression enhanced proliferation of cell populations, in vitro and in an orthotopic xenograft model. However, two TRIM22 mutants, one with a critical RING-finger domain deletion and the other with amino acid changes at two active sites of RING E3 ligase (C15/18A), were both unable to promote GBM cell proliferation over controls, thus implicating E3 ligase activity in the growth-promoting properties of TRIM22. Co-immunoprecipitations demonstrated that TRIM22 bound a negative regulator of NF-κB, NF-κB inhibitor alpha (IκBα), and accelerated its degradation by inducing K48-linked ubiquitination. TRIM22 also formed a complex with the NF-κB upstream regulator IKKγ and promoted K63-linked ubiquitination, which led to the phosphorylation of both IKKα/β and IκBα. Expression of a non-phosphorylation mutant, srIκBα, inhibited the growth-promoting properties of TRIM22 in GBM cell lines. Finally, TRIM22 was increased in a cohort of primary GBM samples on a tissue microarray, and high expression of TRIM22 correlated with other clinical parameters associated with progressive gliomas, such as wild-type IDH1 status. In summary, our study revealed that TRIM22 activated NF-κB signaling through posttranslational modification of two critical regulators of NF-κB signaling in GBM cells.

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microRNA-802 inhibits epithelial-mesenchymal transition through targeting flotillin-2 in human prostate cancer

Bioscience Reports ◽

10.1042/bsr20160521 ◽

2017 ◽

Vol 37 (2) ◽

Cited By ~ 14

Author(s):

Dawei Wang ◽

Guoliang Lu ◽

Yuan Shao ◽

Da Xu

Keyword(s):

Prostate Cancer ◽

Cell Proliferation ◽

Cell Lines ◽

Cell Apoptosis ◽

Epithelial Mesenchymal Transition ◽

Xenograft Model ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Mesenchymal Transition

miRNAs are a class of non-coding RNAs that exert critical roles in various biological processes. The aim of the present study was to identify the functional roles of miR-802 in regulating epithelial–mesenchymal transition (EMT) in prostate cancer (PCa). miR-802 expression was detected in 73 pairs of PCa samples and PCa cell lines (PC3 and DU145 cells) by qRT-PCR. Cell proliferation was detected using MTT assay, and cell apoptosis was evaluated using flow cytometry. Transwell assay was conducted to investigate cell migration and invasion. Expression analysis of a set of EMT markers was performed to explore whether miR-802 is involved in EMT program. Xenograft model was established to investigate the function of miR-802 in carcinogenesis in vivo. The direct regulation of Flotillin-2 (Flot2) by miR-802 was identified using luciferase reporter assay. miR-802 was remarkably down-regulated in PCa tissues and cell lines. Gain-of-function trails showed that miR-802 serves as an ‘oncosuppressor’ in PCa through inhibiting cell proliferation and promoting cell apoptosis in vitro. Overexpression of miR-802 significantly suppressed in vivo PCa tumor growth. Luciferase reporter analysis identified Flot2 as a direct target of miR-802 in PCa cells. Overexpressed miR-802 significantly suppressed EMT, migration and invasion in PCa cells by regulating Flot2. We identified miR-802 as a novel tumor suppressor in PCa progression and elucidated a novel mechanism of the miR-802/Flot2 axis in the regulation of EMT, which may be a potential therapeutic target.

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SNHG9 Promotes the Hepatoblastoma Tumorigenesis via miR-23a-5p/Wnt3a Axis

10.21203/rs.3.rs-335750/v1 ◽

2021 ◽

Author(s):

Ramesh Bhandari ◽

Sun Gui Feng ◽

Liu Ya ◽

Bian Zhixuan ◽

Pan Quihui ◽

...

Keyword(s):

Cell Proliferation ◽

Cell Lines ◽

Molecular Mechanisms ◽

Pearson Correlation ◽

Direct Interaction ◽

Luciferase Reporter ◽

Kaplan Meier ◽

Rna Immunoprecipitation ◽

Cellular Apoptosis

Abstract Background: Hepatoblastoma is common hepatic tumors occurring children between 0 – 5 years. Accumulating studies has shown lncRNA potential role in distinct cancers progression and development including the hepatoblastoma. SnoRNA host gene 9 (SNHG9) is associated the progression of distinct human cancers but, it`s specific molecular mechanisms in hepatoblastoma not unknown. Methods:In this study, we estimated SNHG9 expression on hepatoblastoma tissue and cell lines by quantitative Real-Time Polymerase Chain Reaction (qRT-PCR). Next, we downregulated and upregulated the SNHG9 expression in hepatoblastoma cell lines and then determined the cell proliferation (CCK-8), colony formation, cellular apoptosis activity. The dual luciferase reporter activity, RNA immunoprecipitation (RIP), biotin RNA pulls down and Spemann’s Pearson correlation coefficient assay were performed to establish the interaction between the SNHG9, WNt3a and miR-23a-5p. Xenograft in-vivo tumorgenicity test was performed to elucidate therole of SNHG9 hepatoblastoma in tumorigenesis. SNHG9 role in Cisplatin drugs resistance in hepatoblastoma was also determined. Results:SNHG9 was significantly upregulated in hepatoblastoma tissue and cell lines. SNHG9 overexpression on HUH6 & HepG2 resulted in a significant increase in cell proliferation and clonogeneic while SNHG9 knock down resulted in a sustained inhibition of cell proliferation and clonogenic activity. Dual luciferase activity, RNA immunoprecipitation and biotin pull down confirmed the direct interaction of miR-23a-5p with SNHG9. In Xenograft tumorgenicity test showed SNHG9 downregulation significantly reduced the tumor growth on mice. ROC and Kaplan-Meier analysis showed potential prognostic and diagnostic importance of SNHG9 in hepatoblastoma.Conclusion: We concluded that SNHG9/miR-23a-5p/Wnt3a axis promotes the progression hepatoblastoma tumor.

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Hsa_circ_0026416 promotes proliferation and migration in colorectal cancer via miR-346/NFIB axis

Cancer Cell International ◽

10.1186/s12935-020-01593-1 ◽

2020 ◽

Vol 20 (1) ◽

Author(s):

Yahang Liang ◽

Jingbo Shi ◽

Qingsi He ◽

Guorui Sun ◽

Lei Gao ◽

...

Keyword(s):

Colorectal Cancer ◽

Cell Proliferation ◽

Molecular Mechanisms ◽

Xenograft Model ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Potential Biomarker ◽

Mouse Xenograft Model

Abstract Background Colorectal cancer (CRC) is one of the most common cancers worldwide. Circular RNAs (circRNAs), a novel class of non-coding RNAs, have been confirmed to be key regulators of many diseases. With many scholars devoted to studying the biological function and mechanism of circRNAs, their mysterious veil is gradually being revealed. In our research, we explored a new circRNA, hsa_circ_0026416, which was identified as upregulated in CRC with the largest fold change (logFC = 3.70) of the evaluated circRNAs via analysing expression profiling data by high throughput sequencing of members of the GEO dataset (GSE77661) to explore the molecular mechanisms of CRC. Methods qRT-PCR and western blot analysis were utilized to assess the expression of hsa_circ_0026416, miR-346 and Nuclear Factor I/B (NFIB). CCK-8 and transwell assays were utilized to examine cell proliferation, migration and invasion in vitro, respectively. A luciferase reporter assay was used to verify the combination of hsa_circ_0026416, miR-346 and NFIB. A nude mouse xenograft model was also utilized to determine the role of hsa_circ_0026416 in CRC cell growth in vivo. Results Hsa_circ_0026416 was markedly upregulated in CRC patient tissues and plasma and was a poor prognosis in CRC patients. In addition, the area under the curve (AUC) of hsa_circ_0026416 (0.767) was greater than the AUC of CEA (0.670), CA19-9 (0.592) and CA72-4 (0.575). Functionally, hsa_circ_0026416 promotes cell proliferation, migration and invasion both in vitro and in vivo. Mechanistically, hsa_circ_0026416 may function as a ceRNA via competitively absorbing miR-346 to upregulate the expression of NFIB. Conclusions In summary, our findings demonstrate that hsa_circ_0026416 is an oncogene in CRC. Hsa_circ_0026416 promotes the progression of CRC via the miR-346/NFIB axis and may represent a potential biomarker for diagnosis and therapy in CRC.

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The proliferation and invasion of osteosarcoma are inhibited by miR-101 via targetting ZEB2

Bioscience Reports ◽

10.1042/bsr20181283 ◽

2019 ◽

Vol 39 (2) ◽

Cited By ~ 5

Author(s):

Haopeng Lin ◽

Xiaodong Zheng ◽

Ting Lu ◽

Yang Gu ◽

Canhao Zheng ◽

...

Keyword(s):

Cell Proliferation ◽

Cell Lines ◽

Molecular Mechanisms ◽

Bone Cell ◽

Luciferase Reporter ◽

Osteosarcoma Cell ◽

Osteosarcoma Cells ◽

Human Osteosarcoma ◽

Direct Target ◽

Proliferation And Invasion

AbstractHaving a better grasp of the molecular mechanisms underlying carcinogenesis and progression in osteosarcoma would be helpful to find novel therapeutic targets. Different types of cancers have presented abnormal expression of miRNA-101 (miR-101). Nevertheless, we still could not figure out what expression of miR-101 in human osteosarcoma is and its biological function. Thus, we conducted the present study to identify its expression, function, and molecular mechanism in osteosarcoma. We detected the expression of miR-101 in osteosarcoma samples and cell lines. The effects of miR-101 on osteosarcoma cells’ proliferation and invasion were evaluated. Luciferase reporter assay was applied to identify the direct target of miR-101. Compared with adjacent normal specimens and normal bone cell line by using qPCR, the expression levels of miR-101 in osteosarcoma specimens and human osteosarcoma cell lines distinctly decreased. According to function assays, we found that overexpression of miR-101 significantly inhibited the cell proliferation and invasion in osteosarcoma cells. Moreover, we confirmed that zinc finger E-box binding homeobox 2 (ZEB2) was a direct target of miR-101. In addition, overexpression of ZEB2 could rescue the inhibition effect of proliferation and invasion induced by miR-101 in osteosarcoma cells. MiR-101 has been proved to be down-regulated in osteosarcoma and has the ability to suppress osteosarcoma cell proliferation and invasion by directly targetting ZEB2.

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miR-140-5p induces cell apoptosis and decreases Warburg effect in chronic myeloid leukemia by targeting SIX1

Bioscience Reports ◽

10.1042/bsr20190150 ◽

2019 ◽

Vol 39 (4) ◽

Cited By ~ 9

Author(s):

Zi-Yuan Nie ◽

Xiao-Jun Liu ◽

Ying Zhan ◽

Meng-Han Liu ◽

Xiao-Yan Zhang ◽

...

Keyword(s):

Cell Proliferation ◽

Chronic Myeloid Leukemia ◽

Tumor Suppressor ◽

Cell Lines ◽

Cell Apoptosis ◽

Warburg Effect ◽

Myeloid Leukemia ◽

Critical Role ◽

Luciferase Reporter ◽

Protein Levels

Abstract microRNAs (miRNA), as tumor suppressors or oncogenes, are involved in modulating cancer cell behavior, including cell proliferation and apoptosis. The miR-140-5p acts as a tumor suppressor in several tumors, but the role of miR-140-5p in chronic myeloid leukemia (CML) remains unclear. Here, we investigated the suppression of miR-140-5p in CML patients and CML cell lines using quantitative PCR (qPCR) and fluorescence in situ hybridization (FISH). Overexpression miR-140-5p in CML cells significantly inhibited cell proliferation as revealed by the CCK-8 assay and promoted cell apoptosis as revealed by flow cytometry. Moreover, the sine oculis homeobox 1 (SIX1) gene had been confirmed as a direct target of miR-140-5p using bioinformatics analysis and luciferase reporter assays. Overexpression of miR-140-5p decreased the SIX1 protein level in CML cells. SIX1 mRNA and protein levels were significantly up-regulated in CML patients and CML cell lines. Knockdown of SIX1 expression significantly inhibited CML cell proliferation and promoted cell apoptosis. Furthermore, SIX1 as a transcriptional factor positively regulated pyruvate kinase isozyme type M2 (PKM2) expression and played an important role in the Warburg effect. In addition, these findings indicated that miR-140-5p functions as a tumor suppressor and plays a critical role in CML cell apoptosis and metabolism by targeting SIX1. Moreover, the miR-140-5p/SIX1 axis may be a potential therapeutic target in CML.

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RNA m6A reader YTHDF2 facilitates lung adenocarcinoma cell proliferation and metastasis by targeting the AXIN1/Wnt/β-catenin signaling

Cell Death and Disease ◽

10.1038/s41419-021-03763-z ◽

2021 ◽

Vol 12 (5) ◽

Author(s):

Yin Li ◽

Hao Sheng ◽

Feng Ma ◽

Qiang Wu ◽

Jianfang Huang ◽

...

Keyword(s):

Cell Proliferation ◽

Lung Adenocarcinoma ◽

Colony Formation ◽

Critical Role ◽

Xenograft Model ◽

Inhibitory Effect ◽

Negative Regulator ◽

Tumor Xenograft ◽

And Migration

AbstractLung adenocarcinoma (LUAD) remains a leading cause of cancer-related deaths worldwide. YTHDF2 is a reader of N6-methyladenosine (m6A) on RNA and plays a critical role in the initiation and propagation of myeloid leukemia; however, whether YTHDF2 controls the development of LUAD remains to be explored. Here, we found that YTHDF2 was significantly upregulated in LUAD compared with paracancerous normal tissues, and YTHDF2 knockdown drastically inhibited, while its overexpression promoted, cell growth, colony formation and migration of LUAD cells in vitro. In addition, YTHDF2 knockdown significantly inhibited tumorigenesis in a murine tumor xenograft model. Through the integrative analysis of RNA-seq, m6A-seq, CLIP-seq, and RIP-seq datasets, we identified a set of potential direct targets of YTHDF2 in LUAD, among which we confirmed AXIN1, which encodes a negative regulator of the Wnt/β-catenin signaling, as a direct target of YTHDF2. YTHDF2 promoted AXIN1 mRNA decay and subsequently activated the Wnt/β-catenin signaling. Knockout of AXIN1 sufficiently rescued the inhibitory effect of YTHDF2 depletion on lung cancer cell proliferation, colony-formation, and migration. These results revealed YTHDF2 to be a contributor of LUAD development acting through the upregulation of the AXIN1/Wnt/β-catenin signaling, which can be a potential therapeutic target for LUAD.

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Circular RNA Hsa_Circ_0007444 Inhibits Tumor Progression Through MiR-23a-3p/DICER1 in Ovarian Cancer

10.21203/rs.3.rs-955306/v1 ◽

2021 ◽

Author(s):

Min Zhang ◽

Yu Sun ◽

Hanzi Xu ◽

Yaqian Shi ◽

Rong Shen ◽

...

Keyword(s):

Ovarian Cancer ◽

Cell Proliferation ◽

Tumor Growth ◽

Cell Lines ◽

Xenograft Model ◽

Cell Counting ◽

Luciferase Reporter ◽

Ovarian Cancer Cells ◽

Mouse Xenograft Model

Abstract Background: Circular RNAs are a class of non-coding regulatory RNAs reported to be involved in cancer development and progression. Previous studies, including our own, have indicated that hsa_circ_0007444 was downregulated in ovarian cancer (OC) tissues. Herein, we demonstrated another mechanism of hsa_circ_0007444 in ovarian cancer.Methods: The expression of hsa_circ_0007444, miR-23a-3p, and DICER1 were determined by quantitative real-time PCR. Cell proliferation, invasion, migration, and apoptosis were examined by cell counting kit 8, transwell, and flow cytometry assays. The roles of hsa_circ_0007444 in tumor growth and metastasis were assessed in vivo using a nude mouse xenograft model. The bioinformatics tools were employed to predict the binding sites, which were then verified by dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assays. DICER1 protein level was measured by western blot. Results: Hsa_circ_0007444 was downregulated in ovarian cancer cell lines compared with normal ovarian epithelial cell lines. Also, gain- and loss-of-function results indicated that hsa_circ_0007444 inhibited cell proliferation, invasion, migration, increased cell apoptosis of ovarian cancer cells in vitro, and impaired tumor growth and lung metastasis in vivo. Additionally, the results of the bioinformatics analysis, RIP, dual-luciferase reporter, and rescue assays confirmed that hsa_circ_0007444 could interact with AGO2 and sponge miR-23a-3p, thereby upregulating DICER1 expression, which was an important tumor suppressor in ovarian cancer.Conclusion: We found that overexpressed hsa_circ_0007444 could inhibit ovarian cancer progression through the hsa_circ_0007444/miR-23a-3p/DICER1 axis.

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PTBP3 Modulates of P53 Expression and Promotes Colorectal Cancer Cell Proliferation by Maintaining UBE4A mRNA Stability

10.21203/rs.3.rs-827706/v1 ◽

2021 ◽

Author(s):

Canbin Xie ◽

Liang Li ◽

Xiaorong Li ◽

Min Ma ◽

Fei Long ◽

...

Keyword(s):

Colorectal Cancer ◽

Cell Proliferation ◽

Cell Lines ◽

Molecular Mechanisms ◽

Rna Binding ◽

Binding Protein ◽

Critical Role ◽

Cell Counting ◽

P53 Expression ◽

Polypyrimidine Tract Binding Protein

Abstract Background Colorectal cancer (CRC) is the most common malignancy worldwide and has become the second leading cause of cancer-related death worldwide. The RNA-binding protein polypyrimidine tract-binding protein 3 (PTBP3) was recently reported to play a critical role in multiple cancers, and its molecular mechanisms involve RNA splicing, 3′ end processing and translation. However, the role of PTBP3 in CRC is unclear. Methods We analyzed the expression levels of PTPB3 using The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) datasets and clinical tissues. The effect of PTBP3 on CRC cell proliferation was measured by Cell Counting Kit-8 (CCK-8), colony formation, flow cytometry and tumor xenograft assays. A series of experiments were conducted to reveal the mechanisms by which PTBP3 promotes CRC proliferation. Results We showed that PTBP3 was upregulated in CRC and associated with a poor prognosis. Knockdown of PTBP3 in CRC cell lines restricted CRC proliferative capacities in vitro and in vivo. Mechanistically, we found that PTBP3 regulated the expression of the E3 ubiquitin ligase ubiquitination factor E4A (UBE4A) by binding the 3' untranslated region (UTR) of its mRNA, thereby preventing its degradation. We also discovered that UBE4A participated in the degradation of P53, and knockdown of PTBP3 in CRC cell lines increased P53 expression. Overexpression of UBE4A rescued PTBP3 knockdown-induced inhibition of CRC cell proliferation and P53 expression. Conclusions PTBP3 plays an essential role in CRC cell proliferation by stabilizing UBE4A to regulate P53 expression and may serve as a new prognostic biomarker and effective therapeutic target for CRC.

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M6A-mediated upregulation of LINC00958 increases lipogenesis and acts as a nanotherapeutic target in hepatocellular carcinoma

Journal of Hematology & Oncology ◽

10.1186/s13045-019-0839-x ◽

2020 ◽

Vol 13 (1) ◽

Cited By ~ 24

Author(s):

Xueliang Zuo ◽

Zhiqiang Chen ◽

Wen Gao ◽

Yao Zhang ◽

Jinguo Wang ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Cell Lines ◽

Molecular Mechanisms ◽

Xenograft Model ◽

Systemic Administration ◽

Luciferase Reporter ◽

Loss Of Function ◽

Poor Overall Survival

Abstract Background Long non-coding RNAs (lncRNAs) possess significant regulatory functions in multiple biological and pathological processes, especially in cancer. Dysregulated lncRNAs in hepatocellular carcinoma (HCC) and their therapeutic applications remain unclear. Methods Differentially expressed lncRNA profile in HCC was constructed using TCGA data. LINC00958 expression level was examined in HCC cell lines and tissues. Univariate and multivariate analyses were performed to demonstrate the prognostic value of LINC00958. Loss-of-function and gain-of-function experiments were used to assess the effects of LINC00958 on cell proliferation, motility, and lipogenesis. Patient-derived xenograft model was established for in vivo experiments. RNA immunoprecipitation, dual luciferase reporter, biotin-labeled miRNA pull-down, fluorescence in situ hybridization, and RNA sequencing assays were performed to elucidate the underlying molecular mechanisms. We developed a PLGA-based nanoplatform encapsulating LINC00958 siRNA and evaluated its superiority for systemic administration. Results We identified a lipogenesis-related lncRNA, LINC00958, whose expression was upregulated in HCC cell lines and tissues. High LINC00958 level independently predicted poor overall survival. Functional assays showed that LINC00958 aggravated HCC malignant phenotypes in vitro and in vivo. Mechanistically, LINC00958 sponged miR-3619-5p to upregulate hepatoma-derived growth factor (HDGF) expression, thereby facilitating HCC lipogenesis and progression. METTL3-mediated N6-methyladenosine modification led to LINC00958 upregulation through stabilizing its RNA transcript. A PLGA-based nanoplatform loaded with si-LINC00958 was developed for HCC systemic administration. This novel drug delivery system was controlled release, tumor targeting, safe, and presented satisfactory antitumor efficacy. Conclusions Our results delineate the clinical significance of LINC00958 in HCC and the regulatory mechanisms involved in HCC lipogenesis and progression, providing a novel prognostic indicator and promising nanotherapeutic target.

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MicroRNA-140-5p Inhibits the Progression of Colorectal Cancer by Targeting VEGFA

Cellular Physiology and Biochemistry ◽

10.1159/000430237 ◽

2015 ◽

Vol 37 (3) ◽

pp. 1123-1133 ◽

Cited By ~ 41

Author(s):

Wenbo Zhang ◽

Chen Zou ◽

Lei Pan ◽

Ying Xu ◽

Weidong Qi ◽

...

Keyword(s):

Colorectal Cancer ◽

Cell Proliferation ◽

Cell Lines ◽

Molecular Mechanisms ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Integrated Analysis ◽

Loss Of Function ◽

Invasion And Migration ◽

Tumorigenesis And Progression

Background: microRNAs (miRNAs) are small non-coding RNAs and have been shown to play a crucial role in the colorectal cancer (CRC) tumorigenesis and progression. The aim of this study was to investigate the clinical significance and prognostic value of miR-140-5p in CRC. The exact functions and the underlying molecular mechanisms of miR-140-5p in CRC was further determined. Methods: miR-140-5p expression was detected in CRC samples, their adjacent nontumor tissues as well as CRC cell lines by RT-qPCR. Cell proliferation was detected using CCK-8, and cell invasion and migration were evaluated using Transwell assay. The direct regulation of VEGFA by miR-140-5p was identified using luciferase reporter assay. Results: miR-140-5p was significantly dowregulated in CRC tissues and cell lines. Downregulation of miR-140-5p was significantly correlated with advanced CRC stage and poorer overall survival. Both gain-of-function and loss of function studies demonstrated that miR-140-5p acted as a tumor suppressor by inhibiting cell proliferation, migration and invasion. Integrated analysis identified VEGFA as a direct and functional target gene of miR-140-5p. Silencing VEGFA by small interfering RNA (siRNA) resembled the phenotype resulting from ectopic miR-140-5p expression, while overexpression of VEGFA attenuated the effect of miR-140-5p on CRC cells. Conclusions: Our results suggested a tumor suppressive role of miR-140-5p in CRC tumorigenesis and progression by targeting VEGFA.

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