• Asian Pacific Journal of Cancer Prevention
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• v.14 no.5
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• pp.2983-2986
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• 2013

MicroRNAs (miRNAs) are small, non-coding RNAs (18-25 nucleotides) that post-transcriptionally modulate gene expression by negatively regulating the stability or translational efficiency of their target mRNAs. In this context, the present study aimed to evaluate the in vitro effects of miR-153 inhibition in the breast carcinoma cell line MDA-MB-231. Forty-eight hours after MDA-MB-231 cells were transfected with the miR-153 inhibitor, an MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay was utilized to determine the effects of miR-153 on cell viability. Flow cytometry analysis and assessment of caspase 3/7 activity were adopted to determine whether miR-153 affects the proliferation rates and apoptosis levels of MDA-MB-231 cells. Our results showed that silencing of miR-153 significantly inhibited growth when compared to controls at 48 hours, reducing proliferation by 37.6%, and inducing apoptosis. Further studies are necessary to corroborate our findings and examine the potential use of this microRNA in future diagnostic and therapeutic interventions.

• Journal of Microbiology
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• v.34 no.1
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• pp.30-36
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• 1996

We have exmained the re-establishment of HIMRE mediated silencing function on the transcriptional activity of yeast heast shock gene HSP82. To test whether the onset of SIR repression can occur in growing cells in the rpesence of a potent inhibitor of DNA replication, HMRa/HSP82 strains with SIR4- and SIR4S$^{+}$ genetic backgrounds were arrested in S phase by incubation of a culture in 200 mM hydroxyurea for 120 min. It was clear that following a 20 minute heat shock, silencing of the HMRa/HSP82 allele in cells pretreated with hydroxyurea does occur in a SIR4-dependen fashion, even though the kinetics of repression appears to be substantially delayed. We also have tested whether re- establishement of silencing at the HMR/hsp82 locus can occur in G1-arrested cells. Cell cycle arrest at G1 phase was achieved by treatment of early log a cell cultures with .alpha.-factor mating pheromone, which induces G1 arrest. The result suggests that passage through S phase (and therefore DNA replication) is nor required for re-establishing silencer-mediated repression at the HMNRa/HSP82 locus. Finally, to test whether de nono protein synthesis is required for re-establishment of silencer-mediated repression, cells were pretreated with cycloheximide (500 /.mu.g/ml) 120 min. It was apparent that inhibiting protein synthesis delays, but does not prevent, re-establishment of silencer-mediated repression. Altogether, these results indicate that re-establishment of silencer-mediated repression is not dependent on the DNA replication and has no requirement for protein synthesis.s.

• BMB Reports
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• v.53 no.12
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• pp.658-663
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• 2020

The N-myc downstream regulated gene (NDRG) family members are dysregulated in several tumors. Functionally, NDRGs play an important role in the malignant progression of cancer cells. However, little is known about the potential implications of NDRG4 in pancreatic ductal adenocarcinoma (PDAC). The aim of the current study was to elucidate the expression pattern of NDRG4 in PDAC and evaluate its potential cellular biological effects. Here, we firstly report that epigenetic-mediated silencing of NDRG4 promotes PDAC by regulating mitochondrial function. Data mining demonstrated that NDRG4 was significantly down-regulated in PDAC tissues and cells. PDAC patients with low NDRG4 expression showed poor prognosis. Epigenetic regulation by DNA methylation was closely associated with NDRG4 down-regulation. NDRG4 overexpression dramatically suppressed PDAC cell growth and metastasis. Further functional analysis demonstrated that up-regulated NDRG4 in SW1990 and Canpan1 cells resulted in attenuated mitochondrial function, including reduced ATP production, decreased mitochondrial membrane potential, and increased fragmented mitochondria. However, opposite results were obtained for HPNE cells with NDRG4 knockdown. These results indicate that hypermethylation-driven silencing of NDRG4 can promote PDAC by regulating mitochondrial function and that NDRG4 could be as a potential biomarker for PDAC patients.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.11
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• pp.5619-5625
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• 2012

Gastric carcinoma is a leading cause of cancer death in the world and multi-drug resistance (MDR) is an essential aspect of gastric carcinoma chemotherapy failure. Recent studies have shown that integrin-linked kinase (ILK) is involved in metastasis of human tumors, expression silencing of ILK inhibiting the metastasis of several types of cultured human cancer cells. However, the role and potential mechanism of ILK to reverse the multi-drug resistance in human gastric carcinoma is not fully clear. In this report, we focused on roles of expression silencing of ILK in multi-drug resistance reversal of human gastric carcinoma SGC7901/DDP cells, including increased drug sensitivity to cisplatin, cell apoptosis rates, and intracellular accumulation of Rhodamine-123, and decreased mRNA and protein expression of multi-drug resistance gene (MDR1), multi-drug resistance-associated protein (MRP1), excision repair cross-complementing gene 1 (ERCC1), glutathione S-transferase -${\pi}$ (GST-${\pi}$) and RhoE, and transcriptional activation of AP-1 and NF-${\kappa}B$ in ILK silenced SGC7901/DDP cells. We also found that there was a decreased level of p-Akt and p-ERK. The results indicated that ILK might be used as a potential therapeutic strategy to combat multi-drug resistance through blocking PI3K-Akt and MAPK-ERK pathways in human gastric carcinoma.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.1
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• pp.127-131
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• 2013

Trastuzumab is the first molecular targeting drug to increase the overall survival rate in advanced gastric cancer. However, it has also been found that a high intrinsic or primary trastuzumab resistance exists in some proportion of gastric cancer patients. In order to explore the mechanism of resistance to trastuzumab, firstly we investigated the expression of MUC1 (membrane-type mucin 1) in gastric cancer cells and its relationship with drug-resistance. Then using gene-silencing, we transfected a siRNA of MUC1 into drug-resistant cells. The results showed the MKN45 gastric cell line to be resistant to trastuzumab, mRNA and protein expression of MUC1 being significantly upregulated. After transfection of MUC1 siRNA, protein expression of MUC1 in MKN45cells was significantly reduced. Compared with the junk transfection and blank control groups, the sensitivity to trastuzumab under MUC1 siRNA conditions was significantly increased. These results imply that HER2-positive gastric cancer cell MKN45 is resistant to trastuzumab and this resistance can be cancelled by silencing expression of the MUC1 gene.

• Journal of Crop Science and Biotechnology
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• v.10 no.2
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• pp.73-78
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• 2007

The green revolution has significantly helped in increasing the food production. So far, various breeding methods have been exploited, besides them recombination DNA technology provides another approach for increasing the food production. By means of this technology the losses in food production incurred by various biotic and abiotic stresses can be effectively controlled. In most of the transgenic studies scientists have used antibiotic resistant genes as markers for easy selection of transformants but there are risks involved in use of GM foods. To make such foods safer and environment friendly we have discussed a novel strategy i.e. Cre-lox which involves site specific recombination. By means of Cre-lox the marker genes can be specifically removed once the selection of transformants is over. In addition, this strategy can be used to module the hybrid chromosomes, avoid gene silencing and incorporate single copy of a transgene for its higher expression.

• Molecules and Cells
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• v.42 no.2
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• pp.161-165
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• 2019

Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related deaths worldwide and has high rates of metastasis. Transforming growth factor beta-inducible protein (TGFBI) is an extracellular matrix component involved in tumour growth and metastasis. However, the exact role of TGFBI in NSCLC remains controversial. Gene silencing via DNA methylation of the promoter region is common in lung tumorigenesis and could thus be used for the development of molecular biomarkers. We analysed the methylation status of the TGFBI promoter in 138 NSCLC specimens via methylation-specific PCR and evaluated the correlation between TGFBI methylation and patient survival. TGFBI promoter methylation was detected in 25 (18.1%) of the tumours and was demonstrated to be associated with gene silencing. We observed no statistical correlation between TGFBI methylation and clinicopathological characteristics. Univariate and multivariate analyses showed that TGFBI methylation is significantly associated with poor survival outcomes in adenocarcinoma cases (adjusted hazard ratio = 2.88, 95% confidence interval = 1.19-6.99, P = 0.019), but not in squamous cell cases. Our findings suggest that methylation in the TGFBI promoter may be associated with pathogenesis of NSCLC and can be used as a predictive marker for lung adenocarcinoma prognosis. Further large-scale studies are needed to confirm these findings.

• The Plant Pathology Journal
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• v.21 no.2
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• pp.158-163
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• 2005

Virus-induced gene silencing (VIGS) is a recently developed gene transcript suppression technique for characterizing the function of plant genes. However, efficient VIGS has only been studied in a few plant species. In order to extend the application of VIGS, we examined whether a VIGS vector based on TRV would produce recognizable phenotypes in soybean. Here, we report that VIGS using the Tobacco rattle virus (TRV) viral vector can be used in several soybean cultivars employing various agro-inoculation methods including leaf infiltration, spray inoculation, and agrodrench. cDNA fragments of the soybean phytoene desaturase(PDS) was inserted into TRV RNA-2 vector. By agrodrench, we successfully silenced the expression of PDS encoding gene in soybean. The silenced phenotype of PDS was invariably obvious 3 weeks after inoculation with the TRV-based vector. Real-time RT-PCR analyses showed that the endogenous level of GmPDS transcripts was dramatically reduced in the silenced leaf tissues. These observations confirm that the silenced phenotype is closely correlated with the pattern of tissue expression. The TRV-based VIGS using agrodrench can be applied to functional genomics in a soybean plants to study genes involved in a wide range of biological processes. To our knowledge, this is the first high frequency VIGS method in soybean plants.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.23
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• pp.10299-10306
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• 2015

The esophageal squamous cell carcinoma (ESCC) is thought to develop through a multi-stage process. Epigenetic gene silencing constitutes an alternative or complementary mechanism to mutational events in tumorigenesis. Posttranscriptional regulation of human leukocyte antigen class I (HLA-I) and antigen processing machinery (APM) proteins expression may be associated with novel epigenetic modifications in cancer development. In the present study, we determined the expression levels of HLA-I antigen and APM components by immunohistochemistry. Then by a bisulfite-sequencing PCR (BSP) approach, we identified target CpG islands methylated at the gene promoter region of APM family genes in a ESCC cell line (ECa109), and further quantitative analysis of CpG site specific methylation of these genes in cases of Kazakh primary ESCCs with corresponding non-cancerous esophageal tissues using the Sequenom MassARRAY platform. Here we showed that the development of ESCCs was accompanied by partial or total loss of protein expression of HLA-B, TAP2, LMP7, tapasin and ERp57. The results demonstrated that although no statistical significance was found of global target CpG fragment methylation level sof HLA-B, TAP2, tapasin and ERp57 genes between ESCC and corresponding non-cancerous esophageal tissues, there was significant differences in the methylation level of several single sites between the two groups. Of thesse only the global methylation level of LMP7 gene target fragments was statistically higher ($0.0517{\pm}0.0357$) in Kazakh esophageal cancer than in neighboring normal tissues ($0.0380{\pm}0.0214$, p<0.05). Our results suggest that multiple CpG sites, but not methylation of every site leads to down regulation or deletion of gene expression. Only some of them result in genetic transcription, and silencing of HLA-B, ERp57, and LMP7 expression through hypermethylation of the promoters or other mechanisms may contribute to mechanisms of tumor escape from immune surveillance in Kazakh esophageal carcinogenesis.

• BMB Reports
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• v.48 no.12
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• pp.643-644
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• 2015

In eukaryotes, small RNAs play important roles in both gene regulation and resistance to viral infection. Argonaute proteins have been identified as a key component of the effector complexes of various RNA-silencing pathways, but the mechanistic roles of Argonaute proteins in these pathways are not clearly understood. To address this question, we performed single-molecule fluorescence experiments using an RNA-induced silencing complex (core-RISC) composed of a small RNA and human Argonaute 2. We found that target binding of core-RISC starts at the seed region of the guide RNA. After target binding, four distinct reactions followed: target cleavage, transient binding, stable binding, and Argonaute unloading. Target cleavage required extensive sequence complementarity and accelerated core-RISC dissociation for recycling. In contrast, the stable binding of core-RISC to target RNAs required seed-match only, suggesting a potential explanation for the seed-match rule of microRNA (miRNA) target selection.