RESUMO
INTRODUCTION: Circular RNAs (circRNAs) are related to the pathogenesis and progression of bladder cancer (BC). This research aimed to investigate the role and mechanism of hsa_circ_0008035 (circ_0008035) in BC progression. METHODS: Circ_0008035, microRNA (miR)-1,184, and Ras-related protein 2B (RAP2B) levels were examined in BC via quantitative real-time polymerase chain reaction and Western blotting. Cell Counting Kit-8, colony formation, 5-ethynyl-2'-deoxyuridine staining, flow cytometry, caspase-3 assay kit, transwell, and tube formation assays were conducted to estimate the effects of circ_0008035 on the malignant phenotypes of BC tumors. The interaction between RNAs and genes was evaluated via a dual-luciferase reporter and RNA immunoprecipitation assays. A xenograft model of BC in nude mice was established to estimate the effect of circ_0008035 in BC in vivo. RESULTS: Circ_0008035 and RAP2B levels were upregulated, while miR-1184 abundance was downregulated in BC tissues and cells. Circ_0008035 knockdown constrained cell proliferation, migration, invasion and angiogenesis but promoted apoptosis in vitro. And circ_0008035 silencing curbed xenograft tumor growth in vivo. Circ_0008035 acted as a miRNA sponge for miR-1184. Circ_0008035 increased RAP2B expression by sponging miR-1184. MiR-1184 downregulation relieved the effects of circ_0008035 knockdown on BC progression. And RAP2B knockdown partly reversed the effects of miR-1184 overexpression on BC progression. CONCLUSION: Circ_0008035-mediated BC progression via regulating the miR-1184/RAP2B axis, providing a potential target for BC treatment.
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MicroRNAs , Neoplasias da Bexiga Urinária , Humanos , Animais , Camundongos , Camundongos Nus , Neoplasias da Bexiga Urinária/genética , Bexiga Urinária , Apoptose , Bandagens , Proliferação de Células , MicroRNAs/genética , Proteínas rap de Ligação ao GTPRESUMO
BACKGROUND: Transmembrane protein 43 (TMEM43), a member of the transmembrane protein subfamily, plays a critical role in the initiation and development of cancers. However, little is known concerning the biological function and molecular mechanisms of TMEM43 in pancreatic cancer. METHODS: In this study, TMEM43 expression levels were analyzed in pancreatic cancer samples compared with control samples. The relationship of TMEM43 expression and disease-free survival (DFS) and overall survival (OS) were assessed in pancreatic cancer patients. In vitro and in vivo assays were performed to explore the function and role of TMEM43 in pancreatic cancer. Coimmunoprecipitation (co-IP) followed by protein mass spectrometry was applied to analyze the molecular mechanisms of TMEM43 in pancreatic cancer. RESULTS: We demonstrated that TMEM43 expression level is elevated in pancreatic cancer samples compared with control group, and is correlated with poor DFS and OS in pancreatic cancer patients. Knockdown of TMEM43 inhibited pancreatic cancer progression in vitro, decreased the percentage of S phase, and inhibited the tumorigenicity of pancreatic cancer in vivo. Moreover, we demonstrated that TMEM43 promoted pancreatic cancer progression by stabilizing PRPF3 and regulating the RAP2B/ERK axis. CONCLUSIONS: The present study suggests that TMEM43 contributes to pancreatic cancer progression through the PRPF3/RAP2B/ERK axis, and might be a novel therapeutic target for pancreatic cancer.
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Neoplasias Pancreáticas , Linhagem Celular Tumoral , Proliferação de Células , MAP Quinases Reguladas por Sinal Extracelular/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteínas Nucleares/metabolismo , Neoplasias Pancreáticas/genética , Ribonucleoproteína Nuclear Pequena U4-U6/metabolismo , Proteínas rap de Ligação ao GTP/genética , Proteínas rap de Ligação ao GTP/metabolismoRESUMO
Cutaneous squamous cell carcinoma (CSCC) is an epidermal skin cancer that evolves from normal epidermis along several pre-malignant stages. Previously we found specific miRNAs alterations in each step along these stages. miR-199a-3p expression decreases at the transition to later stages. A crucial step for epithelial carcinoma cells to acquire invasive capacity is the disruption of cell-cell contacts and the gain of mesenchymal motile phenotype, a process known as epithelial-to-mesenchymal transition (EMT). This study aims to study the role of decreased expression of miR-199a-3p in keratinocytes' EMT towards carcinogenesis. First, we measured miR-199a-3p in different stages of epidermal carcinogenesis. Then, we applied Photoactivatable Ribonucleoside-Enhanced Crosslinking and Immunoprecipitation (PAR-CLIP) assay to search for possible biochemical targets of miR-199a-3p and verified that Ras-associated protein B2 (RAP2B) is a bona-fide target of miR-199a-3p. Next, we analyzed RAP2B expression, in CSCC biopsies. Last, we evaluated possible mechanisms leading to decreased miR-199a-3p expression. miR-199a-3p induces a mesenchymal to epithelial transition (MET) in CSSC cells. Many of the under-expressed genes in CSCC overexpressing miR-199a-3p, are possible targets of miR-199a-3p and play roles in EMT. RAP2B is a biochemical target of miR-199a-3p. Overexpression of miR-199a-3p in CSCC results in decreased phosphorylated focal adhesion kinase (FAK). In addition, inhibiting FAK phosphorylation inhibits EMT marker genes' expression. In addition, we proved that DNA methylation is part of the mechanism by which miR-199a-3p expression is inhibited. However, it is not by the methylation of miR-199a putative promoter. These findings suggest that miR-199a-3p inhibits the EMT process by targeting RAP2B. Inhibitors of RAP2B or FAK may be effective therapeutic agents for CSCC.
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Carcinoma de Células Escamosas , MicroRNAs , Neoplasias Cutâneas , Humanos , Carcinoma de Células Escamosas/patologia , Regulação Neoplásica da Expressão Gênica , Proteínas ras/metabolismo , Linhagem Celular Tumoral , Neoplasias Cutâneas/patologia , MicroRNAs/genética , MicroRNAs/metabolismo , Transição Epitelial-Mesenquimal/genética , Proliferação de Células , Proteínas rap de Ligação ao GTP/genética , Proteínas rap de Ligação ao GTP/metabolismoRESUMO
Rap2B, belonging to the Ras superfamily of small guanosine triphosphate-binding proteins, is upregulated and contributes to the progression of several tumors by acting as an oncogene, including hepatocellular carcinoma (HCC). However, the mechanism underlying the functional roles of Rap2B in HCC remains unclear. In this study, the evaluation of Rap2B expression in HCC cells and tissues was achieved by qRT-PCR and western blot assays. The effects of Rap2B on the malignant biological behaviors in HCC were explored by means of MTT assay, flow cytometry analysis, and Transwell invasion assay, respectively. Protein levels of Ki67, matrix metalloproteinase (MMP)-2, MMP-9, and cleaved caspase-3, together with the alternations of the ERK1/2 and PTEN/PI3K/Akt pathways were qualified by western blot assay. Further verification of the Rap2B function on HCC tumorigenesis was attained by performing in vivo assays. We found that Rap2B levels were upregulated in HCC tissues and cells. Rap2B silencing led to a reduction of cell-proliferative and invasive abilities, and an increase of apoptosis in HCC cells. In addition, xenograft tumor assay demonstrated that Rap2B silencing repressed HCC xenograft tumor growth in vivo. In addition, we found that Rap2B knockdown significantly inhibited the ERK1/2 and PTEN/PI3K/Akt cascades in HCC cells and xenograft tumor tissues. Together, Rap2B knockdown inhibited HCC-malignant progression, which was involved in inhibiting the ERK1/2 and PTEN/PI3K/Akt pathways. Our findings contribute to understanding of the molecular mechanism of Rap2B in HCC progression.
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Carcinoma Hepatocelular/metabolismo , Técnicas de Silenciamento de Genes , Neoplasias Hepáticas/metabolismo , Sistema de Sinalização das MAP Quinases , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , PTEN Fosfo-Hidrolase/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteína rab2 de Ligação ao GTP/metabolismo , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Células Hep G2 , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Proteína Quinase 1 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/genética , PTEN Fosfo-Hidrolase/genética , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt , Proteína rab2 de Ligação ao GTP/genéticaRESUMO
PURPOSE: Rap2B, a member of the GTP-binding proteins, is generally up-regulated in numerous types of tumors. Nevertheless, the influence and regulatory mechanisms of Rap2B in gliomas are still not corroborated. Therefore, we analyzed the expression of Rap2B in glioma tissues and cells, and researched its significance in adhesion, proliferation, migration and invasion of the glioma cell line. METHODS: We analyzed the expression of Rap2B in different pathologic grades of glioma tissues by tissue microarray and immunohistochemistry. We assessed the expression of Rap2B in glioma tissue and non-tumor tissue by Western blot. And the expression of Rap2b protein in glioma cells and normal human astrocytes (NHA) was detected by Western blot. In addition, we disclosed the effect of Rap2B knockdown on cell adhesion, proliferation, migration and invasion by using cell attachment assay, CCK-8 assay, cell migration assay and Wound Healing assay, cell invasion assay, respectively. Western blot was used to detect the changes of expression level of NF-kB, MMP-2 and MMP-9 protein when downregulated the expression of Rap2B. RESULTS: The tissue microarray immunohistochemical results of glioma showed that the expression of Rap2B had no significant correlations between Rap2B expression and the clinicopathologic variables, including patient age (P = 0.352), gender (P = 0.858), WHO Grade (P = 0.693) and histology type (P = 0.877). Western blot analysis showed that the glioma tissue had a dramatically increase of Rap2B expression compared with the non-tumor tissues (P < 0.01). And the expression of Rap2B was markedly up-regulated in all 5 glioma cell lines compared with that in normal human astrocytes (NHA) (P < 0.01). We found that the ability of adhesion, proliferation, migration and invasion of glioma cells were significantly decreased after downregulated Rap2B expression compared with the control group (P < 0.05). In addition, Western blot results showed that the expression levels of NF-kB, MMP-2 and MMP-9 in the interference group were significantly lower than those in the negative control group (P < 0.05). CONCLUSIONS: Rap2B expression is up-regulated in glioma tissues and glioma cell lines. Knockdown of Rap2B inhibits glioma cells' adhesion and proliferation in vitro. Knockdown of Rap2B inhibits glioma cells' migration in vitro. Knockdown of Rap2B inhibits glioma cells' invasion and MMPs activity through NF-kB pathway.
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Biomarcadores Tumorais/metabolismo , Adesão Celular , Movimento Celular , Proliferação de Células , Glioma/patologia , Proteínas rap de Ligação ao GTP/metabolismo , Apoptose , Biomarcadores Tumorais/genética , Feminino , Seguimentos , Regulação Neoplásica da Expressão Gênica , Glioma/genética , Glioma/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , NF-kappa B/genética , NF-kappa B/metabolismo , Invasividade Neoplásica , Prognóstico , RNA Interferente Pequeno/genética , Transdução de Sinais , Células Tumorais Cultivadas , Proteínas rap de Ligação ao GTP/antagonistas & inibidores , Proteínas rap de Ligação ao GTP/genéticaRESUMO
BACKGROUND: MicroRNAs (miRNAs), a class of small-regulatory RNA molecules, were closely involved in the pathogenesis of a broad-spectrum of colorectal cancer (CRC). But role of miR-147b in CRC still remains unclear. METHODS: Real-time RT-PCR or Western blotting was utilized to detect the expressions of miR-147b and RAP2B in CRC cell lines and tissues. Luciferase reporter assays were conducted to detect the associations between miR-147b and 3'UTRs of RAP2B. A series of assays were performed to evaluate the effect of miR-147b on proliferation, migration, and invasion of CRC in vitro and in vivo. RESULTS: We found that the level of miR-147b was significantly lower in CRC tissues than in normal tissues (p = 0.0006). Enforced expression of miR-147b led to suppression of CRC cell proliferation in vitro and tumor growth in vivo. Specifically, miR-147b promoted proliferation by arresting CRC cells in the G1/G0 phase. Mechanically, RAP2B was identified as a direct target gene of miR-147b and RAP2B rescued the suppression of proliferation and invasion reduced by miR-147b in CRC cells. CONCLUSIONS: miR-147b not only plays important roles in the regulation of cell proliferation and tumor growth in CRC, which might be a potential prognostic marker or therapeutic target for CRC.
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Proliferação de Células , Neoplasias Colorretais/genética , MicroRNAs/genética , Invasividade Neoplásica , Proteínas rap de Ligação ao GTP/genética , Animais , Linhagem Celular Tumoral , Movimento Celular , Neoplasias Colorretais/patologia , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Experimentais/genética , Neoplasias Experimentais/patologiaRESUMO
Human renal cell carcinoma which is a highly vascular tumor is the leading cause of death from urologic cancers. Angiogenesis has a pivotal role in oncogenesis and in the viability and expansion of renal cell carcinoma. Rap2B, as a small guanosine triphosphate-binding protein of the Ras family, was first discovered in the early 1990s during the screening of a platelet complementary DNA library. Previous studies have shown that Rap2B aberrantly expressed in human carcinogenesis and promoted the development of tumors via multiple signaling pathways. However, the function of Rap2B in tumor angiogenesis that is necessary for tumor growth and metastasis remains unknown. In this study, we examined the role of Rap2B in angiogenesis in renal cell carcinoma by Western blot, quantitative polymerase chain reaction, enzyme-linked immunosorbent assay, human umbilical vascular endothelial cells growth assay, and endothelial cell tube formation assay. We found that Rap2B promoted angiogenesis in vitro and in vivo. Moreover, our data illustrated that phosphoinositide 3-kinase/AKT signaling pathway is involved in Rap2B-mediated upregulation of vascular endothelial growth factor and renal cell carcinoma angiogenesis. Taken together, these results revealed that Rap2B promotes renal cell carcinoma angiogenesis via phosphoinositide 3-kinase/AKT/vascular endothelial growth factor signaling pathway, which suggests that Rap2B is a novel therapeutic target for renal cell carcinoma anti-angiogenesis therapy.
Assuntos
Carcinoma de Células Renais/genética , Proliferação de Células/genética , Neovascularização Patológica/genética , Proteínas rap de Ligação ao GTP/genética , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Regulação Neoplásica da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana , Humanos , Terapia de Alvo Molecular , Metástase Neoplásica , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/patologia , Proteína Oncogênica v-akt/genética , Fosfatidilinositol 3-Quinases/genética , Transdução de Sinais/genética , Fator A de Crescimento do Endotélio Vascular/genética , Proteínas rap de Ligação ao GTP/biossínteseRESUMO
Rap2b, a member of the guanosine triphosphate-binding proteins, is widely up-regulated in many types of tumors. However, the functional role of Rap2b in tumorigenesis of lung cancer remains to be fully elucidated. In this study, we investigated the effect of Rap2b on the lung cancer malignant phenotype, such as cell proliferation and metastasis. We found that Rap2b could promote the abilities of lung cancer cell wound healing, migration, and invasion via increasing matrix metalloproteinase-2 enzyme activity. Furthermore, Rap2b overexpression could increase the phosphorylation level of extracellular signal-regulated protein kinases 1/2. In conclusion, our results suggested that Rap2b may be a potential therapeutic target for lung cancer.
Assuntos
Movimento Celular/genética , Proliferação de Células/genética , Neoplasias Pulmonares/genética , Proteínas rap de Ligação ao GTP/biossíntese , Células A549 , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Neoplasias Pulmonares/patologia , Metaloproteinase 2 da Matriz/biossíntese , Invasividade Neoplásica/genética , Metástase Neoplásica , Proteínas rap de Ligação ao GTP/genéticaRESUMO
Rap2B GTPase, a member of Ras-related protein superfamily, was first discovered from a platelet cDNA library in the early 1990s. Since then, it has been reported to play an important role in regulating cellular processes including cytoskeletal organization, cell growth, and proliferation. It can be stimulated and suppressed by a wide range of external and internal inducers, circulating between GTP-bound active state and GDP-bound inactive state. Increasing focus on Ras signaling pathway reveals critical effects of Rap2B on tumorigenesis. In particular, Rap2B behaves in a p53-dependent manner in regulation of apoptosis and migration. Apart from being an oncogenic activator, Rap2B has been found to participate in many other physiological events via diverse downstream effectors. In this review, we present recent studies on the structure, regulation, and multiple biological functions of Rap2B, shedding light on its potential status in treatment of cancer as well as other diseases.
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Regulação Neoplásica da Expressão Gênica , Neoplasias/patologia , Proteínas rap de Ligação ao GTP/química , Proteínas rap de Ligação ao GTP/metabolismo , Animais , Humanos , Neoplasias/genética , Neoplasias/metabolismo , Transdução de Sinais , Proteínas rap de Ligação ao GTP/genéticaRESUMO
BACKGROUND: The relationship between lung cancer and smoking has been demonstrated. The Rap2B gene is usually overexpressed in lung cancers. This study was aimed to investigate the Rap2B gene expression and its promoter methylation in human bronchial epithelial cells (16HBE) treated by cigarette smoke condensate (CSC). METHODS: 16HBE cells were treated with CSC (1/8 IC50). Soft ager assay, tumorigenicity test, chromosome aberrations analysis were used to identify the transformed cells. The expression level of mRNA and protein of Rap2B was detected using real time PCR and Western blotting, respectively. The genome DNA methylation level was detected using combined bisulfite restriction analysis (COBRA) and the methylation status of the target fragment in Rap2B gene promoter was determined by bisulfite sequencing PCR (BSP). RESULTS: The 16HBE cells were successfully malignant transformed after the chronic exposure to CSC. The expression of Rap2B gradually increased in the process of malignant transformation. Meanwhile, global DNA was hypomethylated. However, no obvious change was observed in the methylation level of Rap2B gene promoter in transformed 16HBE cells. CONCLUSIONS: Rap2B gene may play an important role in the process of lung cancer and global DNA hypomethylation might be an early event in tumorigenesis.
Assuntos
Metilação de DNA/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Nicotiana , Fumaça/efeitos adversos , Proteínas rap de Ligação ao GTP/genética , Brônquios/citologia , Linhagem Celular , Transformação Celular Neoplásica , Células Epiteliais/metabolismo , Humanos , Regiões Promotoras Genéticas , Proteínas rap de Ligação ao GTP/metabolismoRESUMO
Glioma is one of the most common primary brain tumors and has a poor prognosis. Rap2B, a member of the Ras family of oncogenes, is highly expressed and promotes the progression of several tumors, including glioma. However, the mechanism underlying the role of Rap2B in glioma is not fully understood. In the present study, after transfection, Rap2B expression was detected by reverse transcription PCR and western blot analysis. Cell proliferation and cell migration assays were performed to determine the effects of Rap2B on the malignant biological behaviors of glioma cells. The changes of ERK pathway-associated proteins were examined by western blot analysis. Enzyme-linked immunosorbent assay (ELISA) and western blot analysis were utilized to detect the protein levels of matrix metalloproteinase (MMP)2 and MMP9. Then, The Cancer Genome Atlas database was used to determine the association between Rap2B expression and clinical parameters in patients with glioblastoma multiforme and low-grade glioma (LGG). Results revealed that Rap2B was highly expressed in human glioma compared with that in adjacent normal tissues and normal human astrocytes, and that silenced Rap2B led to a reduction of cell proliferation and migration ability in glioma cells. Conversely, overexpressed Rap2B in both U87 and U251 cells significantly enhanced these malignant activities. In addition, ELISA assay and western blotting showed that Rap2B increased MMP2 and MMP9 expression. The western blot assay revealed that Rap2B induced the phosphorylation of ERK in glioma cells. Furthermore, silencing the ERK pathway by SCH772984 led to the inhibition of Rap2B-mediated proliferation, migration and the reduction of MMP2 and MMP9 expression. Kaplan-Meier analysis revealed that increased Rap2B expression was associated with poorer survival of patients with LGG. These results demonstrated that Rap2B may participate in the processes of glioma cell proliferation and migration through enhancing MMP2 and MMP9 expression via the ERK pathway. Thus, Rap2B could potentially be used as a promising therapeutic target and prognostic biomarker in glioma.
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The long non-coding RNA colon cancer-associated transcript 1 (CCAT1) has been investigated to involve in the progression of non-small cell lung cancer (NSCLC). Thus, this study aims to explore the detailed molecular mechanisms of CCAT1 in NSCLC. The expression of CCAT1, miR-216a-5p, RAP2B, Bax, Bcl-2, and cleaved caspase 3 was detected by qRT-PCR or Western blot. Cell proliferation, apoptosis, migration, and invasion were analyzed using cell counting kit-8, flow cytometry or Transwell assays, respectively. The interaction between miR-216a-5p and CCAT1 or RAP2B was analyzed by luciferase reporter, RNA immunoprecipitation, and pull-down assays. The expression of CCAT1 was elevated in NSCLC, and CCAT1 deletion could inhibit NSCLC cell proliferation, migration, and invasion but induce apoptosis in vitro as well as imped tumor growth in vivo. MiR-216a-5p was confirmed to be a target of CCAT1, and silencing miR-216a-5p could reverse CCAT1 depletion-mediated inhibitory effects on cell tumorigenesis in NSCLC. Besides that, miR-216a-5p was decreased in NSCLC, and miR-216a-5p restoration inhibited cell tumorigenesis by regulating RAP2B, which was verified to be a target of miR-216a-5p. Additionally, co-expression analysis suggested that CCAT1 indirectly regulated RAP2B level by targeting miR-216a-5p in NSCLC cells. Taken together, CCAT1 deletion could inhibit cell progression in NSCLC through miR-216a-5p/RAP2B axis, indicating a novel pathway underlying NSCLC cell progression and providing new potential targets for NSCLC treatment.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Progressão da Doença , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , MicroRNAs/metabolismo , RNA Longo não Codificante/metabolismo , Proteínas rap de Ligação ao GTP/metabolismo , Animais , Sequência de Bases , Carcinogênese/genética , Carcinogênese/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , Modelos Biológicos , Fenótipo , RNA Longo não Codificante/genética , Transdução de SinaisRESUMO
Background: To explore the biological effects and potential molecular mechanisms of long non-coding RNA (lncRNA) gastric carcinoma proliferation enhancing transcript 1 (GHET1) in acute myeloid leukemia (AML). Methods: Fluorescence in situ hybridization was performed to determine the location of GHET1. Quantitative polymerase chain reaction (qPCR) was performed to verify RNA expression. GHET1 overexpression and knockdown were achieved by transfection of the expression vector or short hairpin RNA. Western blotting, qPCR, Cell Counting Kit-8 assay, JC-1 staining, and flow cytometry were performed to measure GHET1 function. The dual luciferase reporter assay was performed to confirm the relationship between microRNA 105 (mir-105) and Ras-related protein Rap-2B (RAP2B). Results: GHET1 was localized in the nucleus of NB4 cell lines. GHET1 expression was elevated in AML cell lines compared with normal bone marrow mononuclear cells. GHET1 knockdown led to inhibition of proliferation and promoted the differentiation and apoptosis of AML cell lines. Furthermore, GHET1 directly bound to miR-105 and downregulated miR-105 expression. MiR-105 overexpression suppressed proliferation and induced the differentiation and apoptosis of AML cell lines. In addition, RAP2B was confirmed to be a target gene of miR-105 and an inverse correlation was shown between their expression levels in AML cell lines; when miR-105 increased, Rap-2B level decreased and vice versa. Conclusion: This study demonstrated that the GHET1/miR-105/Rap2B axis may be a critical signaling pathway involved in AML progression.
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Numerous studies have reported that the long non-coding RNA (lncRNA) small nucleolar RNA host gene 6 (SNHG6; ENSG00000245910) participates in the development of malignant tumors. However, the underlying mechanism of SNHG6 in the development of melanoma remains unknown. Thus, the present study aimed to investigate the biological role of SNHG6 in the progression of melanoma. SNHG6 expression in melanoma tissues and cells was assessed using a bioinformatics approach and reverse transcription-quantitative PCR analysis. Cell viability was determined using the Cell Counting Kit-8 and colony formation assays. The correlation between microRNA (miR)-101-3p, SNHG6 and RAP2B expression levels was assessed using Pearson's correlation analysis. Bioinformatic analysis and luciferase reporter assay were utilized to confirm the interaction between miR-101-3p and SNHG6 or RAP2B. The Transwell assay was conducted to examine the migratory and invasive activities of melanoma cells. In the present study, SNHG6 expression was upregulated in melanoma tissues and cell lines, and SNHG6 silencing suppressed melanoma cell viability, migration and invasion. SNHG6 was directly bound to miR-101-3p, which interacted with RAP2B. In addition, miR-101-3p expression was negatively correlated with SNHG6 or RAP2B expression. miR-101-3p silencing partially abrogated the suppressive effect of SNHG6-knockdown on RAP2B expression. Moreover, the data demonstrated that RAP2B overexpression reversed the inhibitory effects on melanoma cell proliferation, migration and invasion induced by SNHG6 silencing. In conclusion, the present study identified that SNHG6 accelerated melanoma progression via regulating the miR-101-3p/RAP2B axis. Thus, the SNHG6/miR-101-3p/RAP2B signaling pathway may be a novel therapeutic target for melanoma.
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Rap2b is a novel p53 target we have identified recently. Knockdown of Rap2b sensitizes HCT116 cells to adriamycin-induced apoptosis, indicating that Rap2b promotes adriamycin resistance in cancer cells. In the present study, we designed a nanostructure-based drug/gene delivery system to evaluate the potential of Rap2b siRNA as a therapeutic agent against human cancers. Specifically, after co-incubated with HCT116 cells, adriamycin- and Rap2b siRNA-loaded gold nanoshells were internalized. Subsequent laser irradiation promoted release of adriamycin and Rap2b siRNA from the nanoparticles. The laser-induced release of Rap2b siRNA decreased cellular expression of Rap2b and significantly enhanced the anticancer therapeutic efficacy of adriamycin in vitro and in vivo. In addition, laser irradiation of the nanoparticles might exert an additional thermal killing effect on cancer cells and further improved the anticancer efficacy of adriamycin. In summary, Rap2b siRNA is a potential enhancing agent for adriamycin-based anticancer therapeutics and the gold nanoshell-based drug/gene delivery system carrying both adriamycin and Rap2b siRNA provides a promising anticancer therapeutic strategy.
Assuntos
Doxorrubicina/administração & dosagem , Técnicas de Transferência de Genes , Terapia Genética/métodos , Neoplasias Experimentais/terapia , RNA Interferente Pequeno/administração & dosagem , Proteínas rap de Ligação ao GTP/antagonistas & inibidores , Animais , Western Blotting , Sistemas de Liberação de Medicamentos/métodos , Ouro , Células HCT116 , Humanos , Camundongos , Camundongos Nus , Microscopia Confocal , Nanoconchas , Nanotecnologia , Reação em Cadeia da Polimerase , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The tumor suppressor p53 plays a pivotal role in numerous cellular responses as it regulates cell proliferation, metabolism, cellular growth, and autophagy. In order to identify novel p53 target genes, we utilized an unbiased microarray approach and identified Rap2B as a robust candidate, which belongs to the Ras-related GTP-binding protein superfamily and exhibits increased expression in various human cancers. We demonstrated that p53 increases the intracellular IP3 and Ca2+ levels and decreases the LC3 protein levels through its target gene Rap2B, suggesting that p53 can inhibit the autophagic response triggered by starvation via upregulation of the Rap2B-PLCε-IP3-Ca2+ pathway. As a confirmed target gene of p53, we believe that further investigating potential functions of Rap2B in autophagy and tumorigenesis will provide a novel strategy for cancer therapy.
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Rap2B, a member of the Ras family of small GTP-binding proteins, reportedly presents a high level of expression in various human tumors and plays a significant role in the development of tumor. However, the function of Rap2B in prostate cancer (PCa) remains unclear. We elucidated the stimulative role of Rap2B in PCa cell proliferation, migration and invasion by means of the CCK-8 cell proliferation assay, cell cycle analysis and transwell migration assay. Western blot analysis uncovered that elevated Rap2B leads to increased phosphorylation levels of FAK, suggesting that FAK-dependent pathway might be responsible for the effect of Rap2B on PCa cells migration and invasion. Inversely, FAK-specific inhibitor (PF-573228) can abort Rap2B-induced FAK phosphorylation. In vivo experiment confirmed that Rap2B positively regulated PCa growth and metastasis, as well as the expression of phosphorylated FAK. Collectively, these findings shed light on Rap2B as a potential therapeutic target for PCa.
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Neoplasias da Próstata/enzimologia , Neoplasias da Próstata/patologia , Proteínas rap de Ligação ao GTP/metabolismo , Animais , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Quinase 1 de Adesão Focal/metabolismo , Xenoenxertos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Invasividade Neoplásica , Metástase Neoplásica , FosforilaçãoRESUMO
The Ras family small guanosine 5'-triphosphate (GTP)-binding protein Rap2B is is a member of the Ras oncogene family and a novel target of p53 that regulates the p53-mediated pro-survival function of cells. The Rap2B protein shares ~90% homology with Rap2A, and its sequence is 70% identical to other members of the Rap family such as RaplA and RaplB. As a result, Rap2B has been theorized to have similar signaling effectors to the GTPase-binding protein Rap, which mediates various biological functions, including the regulation of sterile 20/mitogen-activated proteins. Since its identification in the early 1990s, Rap2B has elicited a considerable interest. Numerous studies indicate that Rap2B exerts specific biological functions, including binding and stimulating phospholipase C-ε and interferon-γ. In addition, downregulation of Rap2B affects the growth of melanoma cells. The present review summarizes the possible effectors and biological functions of Rap2B. Increasing evidence clearly supports the association between Rap2B function and tumor development. Therefore, it is conceivable that anticancer drugs targeting Rap2B may be generated as novel therapies against cancer.
RESUMO
Bladder cancer is the 7th most common cancer type in the world, and microRNAs (miRNAs) play important roles in cancer progression. In the present study, we investigated the roles and molecular mechanisms of miR-194 in bladder cancer. The results demonstrated that the expression level of miR-194 is significantly down-regulated in bladder cancer cell lines and clinical tissues. Overexpression of miR-194 inhibited cell proliferation and invasion in J82 and T24 cells. Further mechanistic study showed that overexpression of miR1-94 induced G0/G1 phase arrest as well as apoptosis in J82 and T24 cells. In addition, by using bioinformatics tool (Targetscan), RAP2B is found to be a target of miR-194, and miR-194 down-regulates the expression level of RAP2B via directly targeting its 3'UTR. Knockdown of RAP2B also inhibited cell proliferation and invasion in J28 cells. More importantly, restoration of RAP2B activity rescued the inhibitory effects of miR-194 on cell proliferation and invasion in J82 cells. Further analysis of bladder cancer clinical samples showed that miR-194 is inversely correlated with RAP2B. Collectively, our study may implicate that miR-194 plays an important role in the regulation of bladder cancer progression. In summary, our study may implicate that miR-194 acts as a tumor suppressor and plays an important role in the regulation of bladder cancer progression.
Assuntos
MicroRNAs/metabolismo , Proteínas Repressoras/metabolismo , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/patologia , Proteínas rap de Ligação ao GTP/genética , Apoptose/genética , Sequência de Bases , Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células , Regulação para Baixo/genética , Regulação Neoplásica da Expressão Gênica , Humanos , MicroRNAs/genética , Invasividade Neoplásica , Proteínas rap de Ligação ao GTP/metabolismoRESUMO
Metals are a threat to human health by increasing disease risk. Experimental data have linked altered miRNA expression with exposure to some metals. MiRNAs comprise a large family of non-coding single-stranded molecules that primarily function to negatively regulate gene expression post-transcriptionally. Although several human populations are exposed to low concentrations of As, Cd and Pb as a mixture, most toxicology research focuses on the individual effects that these metals exert. Thus, this study aims to evaluate global miRNA and mRNA expression changes induced by a metal mixture containing NaAsO2, CdCl2, Pb(C2H3O2)2·3H2O and to predict possible metal-associated disease development under these conditions. Our results show that this metal mixture results in a miRNA expression profile that may be responsible for the mRNA expression changes observed under experimental conditions in which coding proteins are involved in cellular processes, including cell death, growth and proliferation related to the metal-associated inflammatory response and cancer.