RESUMO
Thyroid cancer is the most common type of endocrine cancer. Chemokine-like factor (CKLF)-like MARVEL transmembrane domain containing 6 (CMTM6) is recognized as one of its potential immunotherapy targets. The purpose of this study was to investigate the role and molecular mechanism of CMTM6 in regulating the development of thyroid cancer cells. In this study, expression levels of CMTM6 and the sodium/iodide symporter (NIS) were detected by qRT-PCR. Additionally, colony formation assay and flow cytometry were used to detect cell proliferation and apoptosis, while expression levels of various proteins were assessed using Western blotting. Further, the apoptosis and invasion capacity of cells were investigated by scratch and transwell experiments. Finally, the effect of CMTM6 on the epithelial-mesenchymal transition (EMT) of thyroid cancer cells was determined by immunofluorescence assay, which measured the expression levels of epithelial and mesenchymal phenotypic markers. The results of qRT-PCR experiments showed that CMTM6 was highly expressed in thyroid cancer tissues and cells. In addition, knockdown of CMTM6 expression significantly increased NIS expression. Function experiments demonstrated that small interfering (si)-CMTM6 treatment inhibited the proliferation, migration, invasion, and EMT of thyroid cancer cells, while promoting apoptosis of FTC133 cells. Furthermore, mechanistic studies showed that mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinase (ERK) phosphorylation were inhibited by si-CMTM6, as demonstrated by Western blot experiments. In conclusion, our findings demonstrated the role of CMTM6 in the metastasis of thyroid cancer. Briefly, CMTM6 exerts its tumor-promoting effect through the MAPK signaling pathway and could potentially be used as a valuable biomarker for thyroid cancer diagnosis and prognosis.
Assuntos
Proteínas com Domínio MARVEL , Proteínas da Mielina , Simportadores , Neoplasias da Glândula Tireoide , Humanos , Linhagem Celular Tumoral , Proliferação de Células , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Sistema de Sinalização das MAP Quinases , Simportadores/genética , Simportadores/metabolismo , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/metabolismo , Neoplasias da Glândula Tireoide/patologia , Proteínas com Domínio MARVEL/genética , Proteínas com Domínio MARVEL/metabolismo , Proteínas da Mielina/genética , Proteínas da Mielina/metabolismoRESUMO
BACKGROUND: Poly(A)-binding protein interacting protein 1 (PAIP1) is a translational initiation regulatory factor that has been reported as oncogene in multiple malignant diseases. However, its role in hepatocellular carcinoma (HCC) and the potential mechanisms have not been explored. METHODS: PAIP1 expression level in HCC cell lines were detected by real-time quantitative PCR and western blotting. The proliferation and colony formation of HCC cell lines were detected by MTT and colony formation assay. The apoptosis and cell cycle were detected by flow cytometry. The volume and growth rate of the xenograft tumors were observed. The potential mechanism of PAIP1 was analyzed by miRNA Microarray Analysis and TargetScan analysis. RESULTS: PAIP1 is significantly upregulated in HCC cell lines. PAIP1 knockdown dramatically inhibits cell proliferation and colony formation, induces apoptosis and alters the cell cycle distribution by increasing the G2/M cell percentage. Moreover, PAIP1 knockdown significantly reduces tumorigenesis in a murine transplantation model. Bioinformatics and immunoblotting analysis reveal that PAIP1 knockdown dysregulates cyclin D pathway-related proteins. CONCLUSION: PAIP1 plays an oncogenic role in hepatocellular carcinoma.
RESUMO
Has_circ_0008583 is reported to be involved in the progression of hepatocellular carcinoma (HCC), while its biological role in HCC remains unclear. Here, the qRT-PCR was used to detect the expression of has_circ_0008583. The CCK-8 kit was performed to measure cell proliferation. The cell migration and invasion were evaluated by Transwell. A dual-luciferase reporter assay was performed to confirm the target combination between the genes in has_circ_0008583/miR-1301-3p/METTL3 axis. The in vivo role of has_circ_0008583 was verified by murine xenograft assay. Our data showed that hsa_circ_0008583 was upregulated in HCC tissues and cells. Hsa_circ_0008583 overexpression promoted Hep3B cell proliferation, migration and invasion, but hsa_circ_0008583 silencing had an opposing influence. MiR-1301-3p is directly bound to hsa_circ_0008583 and METTL3. MiR-1301-3p overexpression or METTL3 knockdown could partially counteract hsa_circ_0008583 overexpression-mediated influence on HCC cell behaviors. In addition, hsa_circ_0008583 depletion inhibits HCC tumor growth in vivo. In conclusion, hsa_circ_0008583 promotes HCC progression through the miR-1301-3p/METTL3 axis.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , MicroRNAs , Animais , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Metiltransferases/metabolismo , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Circular/genéticaRESUMO
The long non-coding FGD5-AS1 (LncFGD5-AS1) has been reported to be a novel carcinogenic gene and participant in regulating tumor progression by sponging microRNAs (miRNAs). However, the pattern of expression and the biological role of FGD5-AS1 in hepatocellular carcinoma (HCC) remains largely unknown. The expression level of FGD5-AS1 in tumor tissues and cell lines was measured by RT-qPCR. CCK-8, EdU, flow cytometry, wound healing, and transwell chamber assays were performed to investigate the role of FGD5-AS1 in cell proliferation, apoptosis, migration, and invasion in HCC. Dual luciferase reporter, and RNA pull-down assays were performed to identify the regulatory interactions among FGD5-AS1, miR-873-5p and GTP-binding protein 4 (GTPBP4). We found that the expression of FGD5-AS1 was upregulated in HCC tissues and cell lines. Moreover, the knockdown of FGD5-AS1 suppressed cell proliferation, migration and invasion, and induced apoptosis in HCC cells. Further studies demonstrated that FGD5-AS1 could function as a competitive RNA by sponging miR-873-5p in HCC cells. Moreover, GTPBP4 was identified as direct downstream target of miR-873-5p in HCC cells and FGD5-AS1mediated the effects of GTPBP4 by competitively binding with miR-873-5p. Taken together, this study demonstrated the regulatory role of FGD5-AS1 in the progression of HCC and identified the miR-873-5p/GTPBP4 axis as the direct downstream pathway. It represents a promising novel therapeutic strategy for HCC patients.
Assuntos
Carcinoma Hepatocelular/metabolismo , Proteínas de Ligação ao GTP/biossíntese , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/metabolismo , MicroRNAs/biossíntese , Proteínas de Neoplasias/biossíntese , Proteínas Nucleares/biossíntese , Oncogenes , RNA Longo não Codificante/metabolismo , RNA Neoplásico/metabolismo , Regulação para Cima , Carcinoma Hepatocelular/genética , Proteínas de Ligação ao GTP/genética , Células Hep G2 , Humanos , Neoplasias Hepáticas/genética , MicroRNAs/genética , Proteínas de Neoplasias/genética , Proteínas Nucleares/genética , RNA Longo não Codificante/genética , RNA Neoplásico/genéticaRESUMO
PURPOSE: In recent years, long noncoding RNAs (lncRNAs) have received increasing attention as important regulators of cancer development. As yet, however, a large fraction of them has not been characterized in detail, and the functional role of LINC00346 in hepatocellular carcinoma (HCC) has remained unclear. METHODS: The role of LINC00346 in HCC development was investigated using both in vitro and in vivo assays. Interactions between LINC00346, miR-542-3p and WDR18 were assessed using luciferase reporter, RT-qPCR and Western blotting assays. Loss- and gain-of-function experiments were performed to assess the roles of LINC00346, miR-542-3p and WDR18 in HCC cell viability, proliferation, migration and invasion. RESULTS: We found that LINC00346 was upregulated in primary HCC tissues and HCC-derived cell lines and that LINC00346 may promote HCC cell viability, proliferation, migration and invasion. Furthermore, we found that LINC00346 may regulate WDR18 expression via competitively binding to miR-542-3p. This miRNA was found to be downregulated in primary HCC tissues and to act as a tumor suppressor that can inhibit HCC cell viability, proliferation, migration and invasion. In contrast, WDR18 was found to be upregulated in primary HCC tissues and to act as an oncogene. Additional functional studies indicated that WDR18 can activate the Wnt/ß-catenin signaling pathway and its downstream effectors in HCC cells. We also found that LINC00346, through competitive sponging of miR-542-3p, may enhance the expression of WDR18 and activate the Wnt/ß-catenin signaling pathway in HCC cells. Finally, a positive feedback loop involving LINC00346, ß-catenin and MYC in HCC cells was uncovered. CONCLUSIONS: Our results indicate an oncogenic role of LINC00346 in HCC cells via a positive feedback loop involving LINC00346, ß-catenin and MYC, and they may be instrumental for the design of novel HCC biomarkers and/or therapeutic strategies.
Assuntos
Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , RNA Longo não Codificante/metabolismo , beta Catenina/metabolismo , Animais , Carcinogênese/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/mortalidade , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Sobrevivência Celular/genética , Bases de Dados Genéticas , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/mortalidade , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , MicroRNAs/metabolismo , Pessoa de Meia-Idade , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , RNA Longo não Codificante/genética , Transplante Heterólogo , Via de Sinalização Wnt/genéticaRESUMO
Radioresistance is a major obstacle in hepatocellular carcinoma (HCC) radiotherapy. Aberrant expression of long non-coding RNA (lncRNA) has been postulated to be implicated in the development of HCC radioresistance. We investigated the role of lncRNA nuclear enriched abundant transcript 1_2 (NEAT1_2) in radioresistance of HCC and its molecular mechanism in this study. We found that NEAT1_2 and WEE1 were upregulated, and miR-101-3p was downregulated in HCC tissues, as well as HCC cell lines. Downregulation of WEE1 sensitized the radiosensitivity of HCC cells, as evidenced by decreased survival fractions of Huh7 and PLC5 cells and increased percentage of apoptotic cells. Also, knockdown of NEAT1_2 exerted a reinforcing effect on the radiosensitivity of HCC cells. In addition, WEE1 was confirmed as a direct target of miR-101-3p. Upregulation of miR-101-3p obviously decreased the mRNA and protein levels of WEE1 compared with that in the miR-NC group, while transfection of anta-miR-101-3p presented the opposite effects. In parallel, NEAT1_2 was identified to interact with miR-101-3p, and NEAT1_2 upregulated the expression of WEE1 in Huh7 cells through sponging miR-101-3p. Besides, the reinforcing effect of NEAT1_2 silencing could be attenuated by downregulation of miR-101-3p. To conclude, our results support the concept that downregulation of lncRNA NEAT1_2 radiosensitizes hepatocellular carcinoma cells through regulation of miR-101-3p/WEE1 axis.
Assuntos
Carcinoma Hepatocelular/genética , Proteínas de Ciclo Celular/genética , Regulação para Baixo/genética , Neoplasias Hepáticas/genética , MicroRNAs/genética , Proteínas Nucleares/genética , Proteínas Tirosina Quinases/genética , RNA Longo não Codificante/genética , Tolerância a Radiação/genética , Transdução de Sinais , Sequência de Bases , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Humanos , Proteínas Nucleares/metabolismo , Proteínas Tirosina Quinases/metabolismo , RNA Longo não Codificante/metabolismoRESUMO
O-GlcNAcylation is a monosaccharide modification by a residue of N-acetylglucosamine (GlcNAc) attached to serine or threonine moieties on nuclear and cytoplasmic proteins. O-GlcNAcylation is dynamically regulated by O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). Increasing evidence suggests that O-GlcNAcylation is involved in a variety of human cancers. However, the exact role of O-GlcNAcylation in tumor progression remains unclear. Here, we show that O-GlcNAcylation accelerates oncogenic phenotypes of gastric cancer. First, cell models with increased or decreased O-GlcNAcylation were constructed by OGT overexpression, downregulation of OGA activity with specific inhibitor Thiamet-G, or silence of OGT. MTT assays indicated that O-GlcNAcylation increased proliferation of gastric cancer cells. Soft agar assay and Transwell assays showed that O-GlcNAcylation significantly enhanced cellular colony formation, migration, and invasion in vitro. Akt1 activity was stimulated by upregulation of phosphorylation at Ser473 mediated by elevated O-GlcNAcylation. The enhanced cell invasion by Thiamet-G treatment was suppressed by PI3K inhibitor LY294002. Although the cell invasion induced by Thiamet-G was reduced by Akt1 shRNA, it was still higher in comparison with that to the control (cells with Akt1 shRNA alone). And Akt1 overexpression promoted Thiamet-G-induced cell invasion. These results suggested that O-GlcNAcylation enhanced oncogenic phenotypes possibly partially involving PI3K/Akt signaling pathway.