MicroRNAs (miRNAs) are small noncoding RNAs that play fundamental roles in diverse biological and pathological processes by targeting the expression of specific genes. Here, we identified 38 methylation-associated miRNAs, the expression of which could be epigenetically restored by cotreatment with 5-aza-2 0 -deoxycytidine and trichostatin A. Among these 38 miRNAs, we further analyzed miR-34b, miR-127-3p, miR-129-3p and miR-409 because CpG islands are predicted adjacent to them. The methylation-silenced expression of these miRNAs could be reactivated in gastric cancer cells by treatment with demethylating drugs in a time-dependent manner. Analysis of the methylation status of these miRNAs showed that the upstream CpG-rich regions of mir-34b and mir-129-2 are frequently methylated in gastric cancer tissues compared to adjacent normal tissues, and their methylation status correlated inversely with their expression patterns. The expression of miR-34b and miR-129-3p was downregulated by DNA hypermethylation in primary gastric cancers, and the low expression was associated with poor clinicopathological features. In summary, our study shows that tumor-specific methylation silences miR-34b and miR-129 in gastric cancer cells.

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MicroRNAs (miRNAs) are endogenous non-protein-coding RNAs of approximately 22 nucleotides. Thousands of miRNA genes have been identified (computationally and/or experimentally) in a variety of organisms, which suggests that miRNA genes have been widely shared and distributed among species. Here, we used unique miRNA sequence patterns to scan the genome sequences of 56 bilaterian animal species for locating candidate miRNAs first. The regions centered surrounding these candidate miRNAs were then extracted for folding and calculating the features of their secondary structure. Using a support vector machine (SVM) as a classifier combined with these features, we identified an additional 13,091 orthologous or paralogous candidate pre-miRNAs, as well as their corresponding candidate mature miRNAs. Stem-loop RT-PCR and deep sequencing methods were used to experimentally validate the prediction results in human, medaka and rabbit. Our prediction pipeline allows the rapid and effective discovery of homologous miRNAs in a large number of genomes.

MicroRNAs (miRNAs) are short noncoding RNAs that play important roles in cellular processes and disease pathogenesis via the control of specific targeted gene expression. The miR-196s miRNA is encoded at three paralogous loci in three HOX clusters and acts as an oncogenic miRNA in cancer progression. Recent studies have demonstrated that the expression of miR-196b increases cell proliferation and survival in leukemic cells. Here, we used a sequential methylation analysis to reveal that the methylation status correlated well with miR-196b expression in different cell lines. Treatment with the demethylating drug 5-Aza-dC reactivated miR-196b transcription in methylation-silenced cells. Using in vitro methylation approach, we further provide evidences that promoter hypermethylation represses miR-196b transcriptional activation tightly in human cancer cell lines. We also demonstrate that the expression of miR-196b is significantly elevated in gastric cancer and that hypomethylation status of miR-196b CpG islands frequently is observed in primary gastric tumors. Our results provide important information on miR-196s regulation and demonstrate that abnormal DNA hypomethylation induces overexpression of miR-196b in gastric cancer.

MicroRNAs (miRNAs) are short noncoding RNAs (~22 nt) that play important roles in the pathogenesis of human diseases by negatively regulating gene expression. Here, we examined the relationship between miR-196a and gastric cancer.By the analysis of 72 gastric cancer samples, we found that the expression level of miR-196a microRNA significantly increased in primary gastric cancer tissues versus adjacent normal tissues. In addition, extracellular miR-196a detected in conditioned medium was strongly correlated with its cellular expression status and increased circulating miR-196a in patient serum was associated with gastric cancer disease status and relapse. Furthermore, ectopic expression of miR-196a microRNA promoted the epithelial-mesenchymal transition and migration/invasion capabilities of transfected cells, suggesting its oncogenic potential in gastric cancer progression. Altogether, our data demonstrate that miR-196a exerts an oncogenic role in gastric cancer and miR-196a may be a novel biomarker for detecting gastric cancer and for monitoring disease recurrence.

E26 transformation-specific sequence (ETS)-2 is a transcriptional modulator located on chromosome 21, alterations in its expression have been implicated with a reduced incidence of solid tumors in Down syndrome patients. MicroRNAs (miRNAs) are thought to participate in diverse biological functions; however, the regulation of miRNAs is not well characterized. Recently, we reported that miR-196b is highly expressed in gastric cancers. Herein, we demonstrate that miR-196b expression was significantly repressed by ETS2 during gastric cancer oncogenesis. We demonstrate that knockdown of endogenous ETS2 expression increases miR-196b expression. A genomic region between −751 and −824 bp upstream of the miR-196b transcriptional start site was found to be critical for the repression activity. This putative regulatory promoter region contains three potential ETS2-binding motifs. Mutations within the ETS2 binding sites blocked the repression activity of ETS2. Furthermore, knockdown of ETS2 or overexpression of miR-196b significantly induced migration and invasion in gastric cancer cells. In addition, alterations in ETS2 and miR-196b expression in gastric cancer cell lines affected the expression of epithelial–mesenchymal transition-related genes. The levels of vimentin, matrix metalloproteinase (MMP)-2 and MMP9 were drastically induced, but levels of E-cadherin were decreased in shETS2- or miR-196b-transfected cells. Our data indicate that ETS2 plays a key role in controlling the expression of miR-196b, and miR-196b may mediate the tumor suppressor effects of ETS2. We demonstrated that miR-196b was transcriptionally regulated by ETS2 and there was an inverse expression profile between miR-196b and ETS2 in clinical samples. This finding could be beneficial for the development of effective cancer diagnostic and alternative therapeutic strategies.

Metastatic spread of cancer cells portends a poor prognosis and mortality for lung cancer patients. Hypoxia-inducible factor-1α (HIF-1α) enhances tumor cell motility by activating the epithelial-to-mesenchymal transition (EMT), which is considered a prerequisite for metastasis. Recent studies of microRNA involvement in cancer invasion and metastasis have highlighted the role of such RNAs in tumor development. However, little work has been done to identify tumor suppressor microRNAs that target HIF-1α to down-modulate the EMT and thereby counteract the aggressiveness and metastasis of lung cancer cells. Here, we identified the 3′-untranslated region of HIF-1α mRNA as a target of miR-622 and established that miR-622-mediated down-modulation of HIF-1α correlates with decreased levels of mesenchymal proteins, including Snail, β-catenin, and vimentin. Functional analyses revealed that increased miR-622 expression inhibited lung cancer cell migration and invasion in vitro. miR-622 also inhibited the genesis of metastatic lung nodules as demonstrated in a lung cancer xenograft model in which nude mice were transplanted with A549 cells expressing miR-622. Mechanistic analyses showed that overexpression of EGF decreased the miR-622 level in A549 cells, and this reduction could be rescued by administrating U0126, an inhibitor of ERK. Moreover, miR-622 overexpression mediated by the transcription factor FOXO3a decreased the invasiveness of lung tumor cells by inhibiting HIF-1α via inactivation of ERK signaling in U0126-treated A549 cells. These findings highlight the pivotal role of the FOXO3a/miR-622 axis in inhibiting HIF-1α to interfere with tumor metastasis, and this information may contribute to development of novel therapeutic strategies for treating aggressive lung cancer.

Background MicroRNAs (miRNAs) are short noncoding RNAs (approximately 22 nucleotides in length) that play important roles in breast cancer progression by downregulating gene expression. The detailed mechanisms and biological functions of miRNA molecules in breast carcinogenesis have yet to be fully elucidated. This study used bioinformatics and experimental approaches to conduct detailed analysis of the dysregulated miRNAs, arm selection preferences, 3' end modifications, and position shifts in isoforms of miRNAs (isomiRs) in breast cancer. Methods Next-generation sequencing (NGS) data on breast cancer was obtained from the NCBI Sequence Read Archive (SRA). The miRNA expression profiles and isomiRs in normal breast and breast tumor tissues were determined by mapping the clean reads back to human miRNAs. Differences in miRNA expression and pre-miRNA 5p/3p arm usage between normal and breast tumor tissues were further investigated using stem-loop reverse transcription and real-time polymerase chain reaction. Results The analysis identified and confirmed the aberrant expression of 22 miRNAs in breast cancer. Results from pathway enrichment analysis further indicated that the aberrantly expressed miRNAs play important roles in breast carcinogenesis by regulating the mitogen-activated protein kinase (MAPK) signaling pathway. Data also indicated that the position shifts in isomiRs and 3' end modifications were consistent in breast tumor and adjacent normal tissues, and that 5p/3p arm usage of some miRNAs displayed significant preferences in breast cancer. Conclusions Expression pattern and arm selection of miRNAs are significantly varied in breast cancers through analyzing NGS data and experimental approach. These miRNA candidates have high potential to play critical roles in the progression of breast cancer and could potentially provide as targets for future therapy.