Association between polymorphism in the promoter region of lncRNA GAS5 and the risk of colorectal cancer

AbstractThe growth arrest special 5 (GAS5), as a research hotspot of long noncoding RNAs (lncRNAs), has been reported to be associated with colorectal cancer (CRC). However, the association between polymorphisms in GAS5 and the risk of CRC was not clear. In the present study, a case–control study in 1078 CRC patients and 1175 matched healthy controls was performed to evaluate the association between the potential functional genetic variants in GAS5 and the risk of CRC. PCR-TaqMan, qPCR, dual-luciferase assay, electrophoretic mobility shift assay (EMSA), flow cytometry, migration and invasion assays were performed to evaluate the function of polymorphism. Results showed that subjects carrying the rs55829688 CT/TT genotypes had a significantly higher risk of CRC when compared with the CC genotype. Further qPCR assay confirmed that the CRC tissues with rs55829688 CT/TT genotypes had a higher GAS5 mRNA expression level. The dual-luciferase assay, qPCR and EMSA assay revealed that rs55829688 T>C polymorphism could decrease the expression level of GAS5 by impacting the binding ability of the transcription factor Yin Yang-1 (YY1) to the GAS5 promoter region. The expression of apoptosis-related proteins were detected by Western blot. Further, flow cytometry, migration, and invasion experiments showed that GAS5 repressed apoptosis and increased invasion and migration capability of CRC cells. Taken together, our findings provided evidence that the rs55829688 variant in the GAS5 promoter was associated with the risk of CRC and decreased expression of GAS5 by affecting the binding affinity of the transcription factors YY1 to GAS5.

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miR-375 inhibits the proliferation, migration and invasion of esophageal squamous cell carcinoma by targeting XPR1

Current Gene Therapy ◽

10.2174/1566523220666201229155833 ◽

2020 ◽

Vol 20 ◽

Author(s):

Wenbin Wu ◽

Yangmei Zhang ◽

Xiaowu Li ◽

Xiang Wang ◽

Yuan Yuan

Keyword(s):

Squamous Cell Carcinoma ◽

Esophageal Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Squamous Cell ◽

Luciferase Assay ◽

Migration And Invasion ◽

Promoting Effect ◽

Dual Luciferase Assay ◽

Invasion And Migration ◽

And Migration

Objective: The purpose of this study was to explore the mechanism of the miR-375/XPR1 axis in esophageal squamous cell carcinoma (ESCC) and provide a new idea for targeted therapy of ESCC. Methods: Differentially expressed genes in GEO and TCGA databases were analyzed by bioinformatics. The expression levels of miR-375 and XPR1 mRNA were detected by qRT-PCR. Protein expression of XPR1 was detected by western blot. Bioinformatics analysis and dual luciferase assay were conducted to confirm the targeting relationship between miR-375 and XPR1. The viability, proliferation, migration and invasion of cells in each treatment group were detected by CCK-8, colony formation, wound healing and Transwell assays. Results: Significantly down-regulated miR-375 and remarkably up-regulated XPR1 were observed in ESCC tissue and cells. Overexpression of miR-375 inhibited proliferation, invasion and migration of ESCC cells, and greatly reduced the promoting effect of XPR1 overexpression on cell proliferation, migration and invasion. Dual luciferase assay confirmed that miR-375 targeted and inhibited XPR1 expression in ESCC. Conclusion: These results demonstrate the regulatory role of the miR-375/XPR1 axis in ESCC cells and provide a new potential target for the precise treatment of patients with ESCC.

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MicroRNA-490-3p inhibits migration and chemoresistance of colorectal cancer cells via targeting TNKS2

World Journal of Surgical Oncology ◽

10.1186/s12957-021-02226-1 ◽

2021 ◽

Vol 19 (1) ◽

Author(s):

Jing Li ◽

Rubing Mo ◽

Linmei Zheng

Keyword(s):

Colorectal Cancer ◽

Cancer Cells ◽

Target Genes ◽

Chemotherapy Resistance ◽

Luciferase Assay ◽

Migration And Invasion ◽

Invasion And Migration ◽

Colorectal Cancer Cells ◽

And Migration ◽

Common Malignancy

Abstract Objective Colorectal cancer is one of the most common malignancy in the world. The oncogenesis of colorectal cancer is still not fully elucidated. It was reported that microRNA-490-3p (miR-490-3p) was closely related to the regulation of cancers. However, if miR-490-3p could also affect colorectal cancer and the specific mechanism remains unclear. Methods qRT-PCR was conducted to examine the expression of miR-490-3p. DIANA, miRDB, and TargetScan databases were used to identify target genes. LOVO and SW480 cells were transfected by miR-490-3p mimics and inhibitors. Transwell assay was used to measure cell invasion and migration. Cisplatin and fluorouracil were administered to investigate chemotherapy resistance. Western blot was used to measure TNKS2 protein expression. Binding sites were verified using the double luciferase assay. Results miR-490-3p expression was low in the colorectal cancer cells. The level of miR-490-3p was negatively correlated with cell migration and invasion of cancer cells. miR-490-3p could bind to TNKS2 mRNA 3′UTR directly. miR-490-3p can suppress cell viability and resistance to chemotherapy in colorectal cancer cells through targeting TNKS2. Conclusions miR-490-3p could affect colorectal cancer by targeting TNKS2. This study may provide a potential therapeutic target for colorectal cancer.

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LncRNA AFAP1-AS1 Promotes the Progression of Colorectal Cancer through miR-195-5p and WISP1

Journal of Oncology ◽

10.1155/2021/6242798 ◽

2021 ◽

Vol 2021 ◽

pp. 1-10

Author(s):

Yongsheng Li ◽

Zhongpeng Zhu ◽

Xiaokun Hou ◽

Yongjun Sun

Keyword(s):

Colorectal Cancer ◽

Noncoding Rna ◽

Molecular Mechanisms ◽

Luciferase Assay ◽

Migration And Invasion ◽

Dual Luciferase Assay ◽

Inhibition Effect ◽

Common Cancer ◽

Relationship Of ◽

Malignant Behavior

Objective. Colorectal cancer (CRC) is the most common cancer. But, the molecular mechanisms of CRC progression are not fully understood. This study was conducted to explore how the long noncoding RNA actin filament-associated protein 1-antisense RNA1 (lncRNA AFAP1-AS1) participates in CRC progression through the regulation of microRNA-195-5p (miR-195-5p) and wingless-type inducible signaling pathway protein-1 (WISP1). Methods. The expressions of AFAP1-AS1, miR-195-5p, and WISP1 were detected by RT-qPCR or western blot. A dual-luciferase assay confirmed the target relationship of AFAP1-AS1, miR-195-5p, and WISP1. Colony formation, wound-healing, and Transwell assays were used to detect the growth, migration, and invasion abilities of cells, respectively. Results. AFAP1-AS1 and WISP1 expressions were notably increased, and miR-195-5p expression was markedly reduced in CRC. The dual-luciferase assay verified that AFAP1-AS1 could bind to miR-195-5p. AFAP1-AS1 knockdown could inhibit the malignant behavior of CRC cells. miR-195-5p could target and regulate WISP1 expression. Overexpression of WISP1 or miR-195-5p inhibition reversed the inhibition effect of AFAP1-AS1 knockdown on the biological activity of CRC cells. Conclusions. AFAP1-AS1 knockdown may inhibit the proliferation, migration, and invasion of CRC cells through the miR-195-5p/WISP1 axis.

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Decreased level of miR-1301 promotes colorectal cancer progression via activation of STAT3 pathway

Biological Chemistry ◽

10.1515/hsz-2020-0301 ◽

2020 ◽

Vol 0 (0) ◽

Author(s):

Fangfang Yang ◽

Hua Wang ◽

Bianbian Yan ◽

Tong Li ◽

Lulu Min ◽

...

Keyword(s):

Colorectal Cancer ◽

Cell Migration ◽

Cancer Progression ◽

Luciferase Assay ◽

Migration And Invasion ◽

Loss Of Function ◽

Aberrant Expression ◽

Cell Migration And Invasion ◽

Lovo Cells ◽

Colorectal Cancer Progression

Abstract The molecular pathogenesis of colorectal cancer (CRC) has been widely investigated in recent years. Accumulating evidence has indicated that microRNA (miRNA) dysregulation participates in the processes of driving CRC initiation and progression. Aberrant expression of miR-1301 has been found in various tumor types. However, its role in CRC remains to be elucidated. In the present study, we identified miR-1301 was enriched in normal colorectal tissues and significantly down-regulated in CRC. Decreased level of miR-1301 strongly correlated with aggressive pathological characteristics, including advanced stage and metastasis. Bioinformatics and dual luciferase assay demonstrated that STAT3 is a direct target of miR-1301. Gain and loss-of-function assays showed that miR-1301 had no effect on cell proliferation. Overexpression of miR-1301 suppressed cell migration and invasion capacity of pSTA3-positive LoVo cells, but not pSTAT3-negative SW480 cells, while inhibition of miR-1301 consistently promoted cell migration and invasion in both cell lines. Additionally, miR-1301 inhibition restored the suppressed migration and invasion of STAT3- knockdown LoVo cells. MiR-1301 functioned as a tumor suppressor to modulate the IL6/STAT3 signaling pathway. In summary, this study highlights the significant role of miR- 1301/STAT3 axis in CRC metastasis.

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

Cancer Cell International ◽

10.1186/s12935-020-01706-w ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Jinjun Ye ◽

Jidong Liu ◽

Tao Tang ◽

Le Xin ◽

Xing Bao ◽

...

Keyword(s):

Colorectal Cancer ◽

Tumor Growth ◽

Cell Cycle Progression ◽

Bioinformatics Analysis ◽

Scratch Test ◽

Luciferase Assay ◽

Migration And Invasion ◽

Cycle Progression ◽

Ki67 Expression ◽

And Function

Abstract 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.

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Hypermethylated miR-424 in Colorectal Cancer Subsequently Upregulates VEGF

10.21203/rs.3.rs-239421/v1 ◽

2021 ◽

Author(s):

Zahra Nouri Ghonbalani ◽

Shiva Shahmohamadnejad ◽

Parvin Pasalar ◽

Ehsan Khalili

Keyword(s):

Colorectal Cancer ◽

Cancer Cells ◽

Cancer Progression ◽

Promoter Region ◽

Hct116 Cell ◽

Methylation Status ◽

Expression Level ◽

Cancer Tissue ◽

Vegf Expression ◽

Cancer Tissues

Abstract PurposeColorectal cancer (CRC) is the second leading cause of death from cancer in adults. Recent advances have shown that cancer cells can have some epigenetic changes involved in all stages of cancer. It has also been shown that miR-424 acts as gene expression regulators in many biological processes, including angiogenesis with mediators such as VEGF. In the current study, to identify the potential role of miR-424 in colorectal cancer progression, methylation status of miR-424 promoter region and its expression level have been evaluated. Besides, the correlation between VEGF level and miR-424 expression level has been assessed.MethodsMethylation status miR-424 promoter was assessed using methylation-specific polymerase chain reaction (MSP). The expression level of miR-424 in human colorectal cancer tissue was analyzed by quantitative PCR. HCT116 cell line was selected to evaluate the correlation between the miR-424 expression level and the promoter's methylation status. VEGF expression, one out of mir-424 targets involved in angiogenesis and cancer progression, was measured by western blot analysis in the pairs of cancer tissues and their adjacent tissues.ResultsOur results have revealed that the promoter region of miR-424 is methylated in cancer cells compared to normal cells, leading to down-regulation of miR-424 in the colorectal cancer tissues compared to the normal tissues. Also, we found that the expression protein's level of VEGF in the tumor cells increased compared with normal tissues.ConclusionThe present study suggests that hypermethylation downregulates miR-424. VEGF expression is upregulated with decreased miR-424 in colorectal cancer, which results in cancer progression.

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The transcription factor USF1 promotes glioma cell invasion and migration by activating lncRNA HAS2-AS1

Bioscience Reports ◽

10.1042/bsr20200487 ◽

2020 ◽

Vol 40 (8) ◽

Author(s):

Juntong Wang ◽

Jingshun Gu ◽

Aiwu You ◽

Jun Li ◽

Yuyan Zhang ◽

...

Keyword(s):

Transcription Factor ◽

Cell Invasion ◽

Promoter Region ◽

Glioma Cell ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Mesenchymal Transition ◽

Invasion And Migration ◽

Starting Point ◽

And Migration

Abstract Objective: The role of lncRNAs in tumor has been widely concerned. The present study took HAS2-AS1 (the antisense RNA 1 of HAS2) as a starting point to explore its expression in glioma and its role in the process of migration and invasion, providing a strong theoretical basis for mining potential therapeutic targets of glioma. Methods: Clinical data of glioma were obtained from The Cancer Genome Atlas (TCGA) database and differentially expressed lncRNAs were analyzed by edgeR. The hTFtarget database was used to predict the upstream transcription factors of HAS2-AS1 and the JASPAR website was used to predict the binding sites of human upstream transcription factor 1 (USF1) and HAS2-AS1. qRT-PCR was used to detect the expressions of HAS2-AS1 and USF1 in glioma tissues and cell lines. The effects of silencing HAS2-AS1 on the migration and invasion of cancer cells were verified by wound healing and Transwell invasion assays. The chromatin immunoprecipitation (ChIP) and dual luciferase reporter assays were applied to demonstrate the binding of USF1 and HAS2-AS1 promoter region. Western blot was used to detect the expressions of epithelial–mesenchymal transition (EMT)-related proteins. Results: HAS2-AS1 was highly expressed in glioma tissues and cells, and was significantly associated with poor prognosis. Silencing HAS2-AS1 expression inhibited glioma cell migration, invasion and EMT. USF1 was highly expressed in glioma and positively correlated with HAS2-AS1. The transcription of HAS2-AS1 was activated by USF1 via binding to HAS2-AS1 promoter region, consequently potentiating the invasion and migration abilities of glioma cells. Conclusion: These results suggested that the transcription factor USF1 induced up-regulation of lncRNA HAS2-AS1 and promoted glioma cell invasion and migration.

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miR-17-5p promotes the invasion and migration of colorectal cancer by regulating HSPB2

10.21203/rs.3.rs-63587/v2 ◽

2020 ◽

Author(s):

Wei fang Yu ◽

Jia Wang ◽

Chao Li ◽

Mingda Xuan ◽

Shuangshuang Han ◽

...

Keyword(s):

Colorectal Cancer ◽

Cell Proliferation ◽

Lymph Node ◽

Target Genes ◽

Migration And Invasion ◽

Invasion And Migration ◽

Normal Tissues ◽

Node Metastasis ◽

Rescue Experiment ◽

And Migration

Abstract Background: MicroRNA (miRNA) can affect tumor progression by regulating cell proliferation, apoptosis and metastasis. After miRNA microarray chip analysis of colorectal cancer (CRC) tissues and adjacent normal tissues, a significant upregulation of miR-17-5p expression was found in CRC tissues. However, the underlying mechanism of miR-17-5p in CRC is still unclear.Methods: The levels of miR-17-5p in 47 paired CRC and adjacent normal tissue samples were determined by quantitative real-time PCR (qRT-PCR). CCK-8, colony formation, flow cytometry and transwell assays were used to explore the biological effects of miR-17-5p on CRC cells. In addition, the transcriptome sequencing and miRNA target prediction software were employed to identify targets of miR-17-5p. Luciferase reporter detection was used to demonstrate the direct binding of target genes by miR-17-5p. The rescue experiment was conducted to investigate the biological function of target genes and regulatory mechanism of miR-17-5p on target genes.Results: The expression of miR-17-5p was significantly higher in CRC tissues than in adjacent normal tissues. In CRC group, the expression of miR-17-5p in cancer tissues with lymph node metastasis was higher compared with those without lymph node metastasis. Overexpression of miR-17-5p inhibited CRC cell apoptosis, as well as promoting proliferation, migration and invasion. We hypothesized that HSPB2 might be a target gene of miR-17-5p and validated for the first time that miR-17-5p binds directly to the 3’-UTR of HSPB2. In the rescue experiment, the tumor suppressive effect of HSPB2 was detected and miR-17-5p could promote cell proliferation, migration and invasion by targeting HSPB2.Conclusion: MiR-17-5p promotes invasion and migration by inhibiting HSPB2 in CRC, thereby implicating its potential as a novel diagnostic biomarker and therapeutic target for CRC.

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A Functional SNP in the Promoter of LBX1 Is Associated With the Development of Adolescent Idiopathic Scoliosis Through Involvement in the Myogenesis of Paraspinal Muscles

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.777890 ◽

2021 ◽

Vol 9 ◽

Author(s):

Leilei Xu ◽

Zhenhua Feng ◽

Zhicheng Dai ◽

Wayne Y. W. Lee ◽

Zhichong Wu ◽

...

Keyword(s):

Adolescent Idiopathic Scoliosis ◽

Idiopathic Scoliosis ◽

Muscle Development ◽

Luciferase Assay ◽

Paraspinal Muscles ◽

Mobility Shift ◽

Putative Promoter Region ◽

Mobility Shift Assay ◽

First Time ◽

Functional Snp

Previous studies have shown that LBX1 is associated with adolescent idiopathic scoliosis (AIS) in multiple populations. For the first time, rs1322330 located in the putative promoter region of LBX1 was found significantly associated with AIS in the Chinese population [p = 6.08 × 10–14, odds ratio (OR) = 1.42, 95% confidence interval of 1.03–1.55]. Moreover, the luciferase assay and electrophoretic mobility shift assay supported that the allele A of rs1322330 could down-regulate the expression of LBX1 in the paraspinal muscles of AIS. In addition, silencing LBX1 in the myosatellite cells resulted in significantly inhibited cell viability and myotube formation, which supported an essential role of LBX1 in muscle development of AIS. To summarize, rs1322330 may be a novel functional SNP regulating the expression of LBX1, which was involved in the etiology of AIS possibly via regulation of myogenesis in the paraspinal muscles.

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miR-224 targets BTRC and promotes cell migration and invasion in colorectal cancer

3 Biotech ◽

10.1007/s13205-020-02477-x ◽

2020 ◽

Vol 10 (11) ◽

Author(s):

Qi Zheng ◽

Jane J. Yu ◽

Chenggang Li ◽

Jiali Li ◽

Jiping Wang ◽

...

Keyword(s):

Colorectal Cancer ◽

Cell Migration ◽

Protein Expression ◽

Molecular Mechanisms ◽

Early Stage ◽

Luciferase Assay ◽

Migration And Invasion ◽

Cell Migration And Invasion ◽

Normal Tissues ◽

The Impact

AbstractOur study aims to investigate the impact of miR-224 on cell migration and invasion in colorectal cancer (CRC) as well as its molecular mechanisms. The results showed that miR-224 was significantly upregulated in CRC compared to normal tissues via the TCGA database. Overexpression of miR-224 promoted CRC cell migration and invasion, while inhibition of miR-224 demonstrated the opposite result via transwell assays. In addition, we found that BTRC was a target gene of miR-224 through the miRecords database and dual-luciferase assay, while western blot together with RT-qPCR showed that inhibition of miR-224 led to elevated BTRC expression in protein level but not in mRNA level, and also decreased the expression of β-catenin. In reference to the Human Protein Atlas, BTRC protein expression was higher in normal tissues than in CRC tissues. In conclusion, miR-224 regulates its target BTRC protein expression and its related Wnt/β-catenin pathway. Its impact on cell migration and invasion in CRC cells suggested that miR-224 could be a prospective therapeutic target for early-stage non-metastatic CRC.

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