• International Journal of Oral Biology
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• v.37 no.3
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• pp.146-152
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• 2012

MicroRNAs (miRNAs, miRs) are about 21-25 nucleotides in length and regulate mRNA translation by base pairing to partially complementary sites, predominantly in the 3'-untranslated region (3'-UTR) of the target mRNA. In this study, the expression profile of miRNAs was compared and analyzed for the establishment of miRNA-related odontoblast differentiation using MDPC-23 cells derived from mouse dental papilla cells. To determine the expression profile of miRNAs during the differentiation of MDPC-23 cells, we employed miRNA microarray analysis, quantitative real-time PCR (qRT-PCR) and Alizaline red-S staining. In the miRNA microarray analysis, 11 miRNAs were found to be up- or down-regulated more than 3-fold between day 0 (control) and day 5 of MDPC-23 cell differentiation among the 1,769 miRNAs examined. In qRT-PCR analysis, the expression levels of two of these molecules, miR-194 and miR-126, were increased and decreased in the control MDPC-23 cells compared with the MDPC-23 cells at day 5 of differentiation, respectively. Importantly, the overexpression of miR-194 significantly accelerated mineralization compared with the control cultures during the differentiation of MDPC-23 cells. These results suggest that the miR-194 augments MDPC-23 cell differentiation, and potently accelerates the mineralization process. Moreover, these in vitro results show that different miRNAs are deregulated during the differentiation of MDPC-23 cells, suggesting the involvement of these genes in the differentiation and mineralization of odontoblasts.

• Genomics & Informatics
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• v.3 no.1
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• pp.15-23
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• 2005

Background MicroRNAs (miRNAs) are a class of noncoding RNAs found in various organisms such as plants and mammals. However, most of the mRNAs regulated by miRNAs are unknown. Furthermore, miRNA targets in genomes cannot be identified by standard sequence comparison since their complementarity to the target sequence is imperfect in general. In this paper, we propose a kernel-based method for the efficient prediction of miRNA targets. To help in distinguishing the false positives from potentially valid targets, we elucidate the features common in experimentally confirmed targets. Results The performance of our prediction method was evaluated by five-fold cross-validation. Our method showed 0.64 and 0.98 in sensitivity and in specificity, respectively. Also, the proposed method reduced the number of false positives by half compared with TargetScan. We investigated the effect of feature sets on the classification of miRNA targets. Finally, we predicted miRNA targets for several miRNAs in the Caenorhabditis elegans (C. elegans) 3' untranslated region (3' UTR) database. Condusions The targets predicted by the suggested method will help in validating more miRNA targets and ultimately in revealing the role of small RNAs in the regulation of genomes. Our algorithm for miRNA target site detection will be able to be improved by additional experimental­knowledge. Also, the increase of the number of confirmed targets is expected to reveal general structural features that can be used to improve their detection.

• Proceedings of the Botanical Society of Korea Conference
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• 1999.07a
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• pp.57-62
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• 1999

Light-regulated translation of chloroplast mRNAs requires nuclear-encoded trans-acting factors that interact with the 5' untranslated region (UTR) of these mRNAs. A set of four proteins (60, 55, 47, and 38 kDa) that bind to the 5'-UTR of the psbA mRNA had been identified in C. reinhardtii. 47 kDa protein (RB47) was found to encode a chloroplast poly (A)-binding protein (cPABP) that specifically binds to the 5'-UTR of the psbA mRNA, and essential for translation of this mRNA, cDNA encoding 60 kDa protein (RB60) was isolated, and the amino acid sequence of the encoded protein was highly homologous to plants and mammalian protein disulfide isomerases (PDI), normally found in the endoplasmic reticulum (ER). Immunoblot analysis of C. reinhardtii proteins showed that anti-PDI recognized a distinct protein of 56 kDa in whole cell extract, whereas anti-rRB60 detected a 60 kDa protein. The ER-PDI was not retained on heparin-agarose resin whereas RB60 was retained. In vitro translation products of the RB60 cDNA can be transported into C. reinhardtii chloroplast in vitro. Immunoblot analysis of isolated pea chloroplasts indicated that higher plant also possess a RB60 homolog. In vitro RNA-binding studies showed that RB60 modulates the binding of cPABP to the 5'-UTR of the psbA mRNA by reversibly changing the redox status of cPABP using redox potential or ADP-dependent phosphorylation. Site-directed mutagenesis of -CGHC- catalytic site in thioredoxin-like domain of RB60 is an unique PDI located in the chloroplast of C. reinhardtii, and suggest that the chloroplast PDI may have evolved to utilize the redox-regulated thioredoxin like domain as a mechanism for regulating the light-activated translation of the psbA mRNA.

• Journal of Life Science
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• v.28 no.6
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• pp.639-647
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• 2018

A new heat shock protein 70 was identified in red-spotted grouper (Epinephelus akaara) based on an expression analysis. The cDNA of red-spotted grouper Hsp70 (designated RgHsp70) was cloned by the rapid amplification of cDNA ends (RACE) techniques. The full-length of RgHsp70 cDNA was 2,152 bp, consisting of a 5'-terminal untranslated region (UTR) of 105 bp, a 3'-terminal UTR of 274 bp, and an open reading frame (ORF) of 1,773 bp that encode a polypeptide of 590 amino acids with a theoretical molecular weight of 64.9 kDa and an estimated isoelectric point of 5.2. Multiple alignment and phylogenetic analyses revealed that the RgHsp70 gene shares a high similarity with other Hsp70 fish genes. RgHsp70 contained all three classical Hsp70 family signatures. The results indicated the RgHsp70 is a member of the heat shock protein 70 family. RgHsp70 mRNA was predominately expressed in the liver, with reduced expression noted in the head-kidney tissues. The expression analysis of different water temperatures (21, 18, 15 and $12^{\circ}C$) for sampled livers revealed that expression gradually increased at $12^{\circ}C$ compared to $21^{\circ}C$. In this study, the effects of water temperature lowering on the physiological conditions were investigated, and the results revealed that novel RgHsp70 may be an important molecule involved in stress responses.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.6
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• pp.3499-3502
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• 2013

MicroRNAs (miRNAs) are a class of endogenously expressed small, non-coding, single-stranded RNAs that negatively regulate gene expression, mainly by binding to 3'- untranslated regions (3'UTR) of their target messenger RNAs (mRNAs), which cause blocks of translation and/or mRNA cleavage. Recently, miRNAprofiling studies demonstrated the microRNA-497 (miR-497) level to be down-regulated in all prostate carcinomas compared with BPH samples. The purpose of this study was to investigate the potential role of miR-497 in human prostate cancer. Proliferation, cell cycle and apoptosis assays were conducted to explore the potential function of miR-497 in human prostate cancer cells. Results showed that miR-497 suppressed cellular growth and initiated G0/G1 phase arrest of LNCaP and PC-3 cells. We also observed that miR-497 increased the percentage of apoptotic cells by increasing caspase-3/7 activity. Taken together, our results demonstrated that miR-497 can inhibit growth and induce apoptosis by caspase-3 activation in prostate cancer cells, which suggest its use as a potential therapeutic target in the future.

• BMB Reports
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• v.47 no.6
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• pp.311-317
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• 2014

microRNAs (miRNAs) are a class of small, non-coding RNAs that play critical posttranscriptional regulatory roles typically through targeting of the 3'-untranslated region of messenger RNA (mRNA). Mature miRNAs are known to be involved in global cellular processes, such as differentiation, proliferation, apoptosis, and organogenesis, due to their capacity to target multiple mRNAs. Thus, imbalances in the expression and/or activity of miRNAs are involved in the pathogenesis of numerous diseases, including pulmonary arterial hypertension (PAH). PAH is a progressive disease characterized by vascular remodeling due to excessive proliferation of pulmonary artery endothelial cells (PAECs) and pulmonary artery smooth muscle cells (PASMCs). Recently, studies have evaluated the roles of miRNAs involved in the pathogenesis of PAH in these pulmonary vascular cells. This review provides an overview of recent discoveries on the role of miRNAs in the pathogenesis of PAH and discusses the potential for miRNAs as therapeutic targets and biomarkers of PAH.

• Genomics & Informatics
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• v.3 no.3
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• pp.53-62
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• 2005

MicroRNAs play an important role in regulating gene expression, but their target identification is a difficult task due to their short length and imperfect complementarity. Burge and coworkers developed a program called TargetScan that allowed imperfect complementarity and established a procedure favoring targets with multiple binding sites conserved in multiple organisms. We improved their algorithm in two major aspects - (i) using well-defined UTR (untranslated region) database, (ii) examining the extent of conservation inside the 3' UTR specifically. Average length in our UTR database, based on the ECgene annotation, is more than twice longer than the Ensembl. Then, TargetScan was used to identify putative binding sites. The extent of conservation varies significantly inside the 3' UTR. We used the 'tight' tracks in the UCSC genome browser to select the conserved binding sites in multiple species. By combining the longer 3' UTR data, TargetScan, and tightly conserved blocks of genomic DNA, we identified 107 putative target genes with multiple binding sites conserved in multiple species, of which 85 putative targets are novel.

• Journal of Life Science
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• v.27 no.12
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• pp.1494-1499
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• 2017

Oral infection due to Candida albicans is a widely recognized and frequent cause of superficial infections of the oral mucosa (oral candidiasis). Although oral candidiasis is not a life-threatening fungemia, it can cause severe problems in individuals under certain conditions. MicroRNAs (miRNAs) are noncoding, small RNA molecules, which regulate the expression of other genes by inhibiting the translation of target mRNAs. The present study was designed to identify miRNAs in C. albicans and determine their possible roles in this organism. miRNA-sized small RNAs (msRNAs) were cloned in C. albicans by deep sequencing, and their secondary structures were analyzed. All the cloned msRNAs satisfied conditions required to qualify them as miRNAs. Bioinformatics analysis revealed that two of the most highly expressed C. albicans msRNAs, Ca-363 and Ca-2019, were located in the 3' untranslated region of the corticosteroid-binding protein 1 (CBP1) gene in a reverse orientation. miRNA mimics were transformed into C. albicans to investigate their RNA-inhibitory functions. RNA oligonucleotide-transformed C. albicans was then observed by fluorescent microscopy. Quantitative PCR analysis showed that these msRNAs did not inhibit CBP1 gene expression 4 hr and 8 hr after ectopic miRNA transformation. These results suggest that msRNAs in C. albicans possess an miRNA-triggered RNA interference gene-silencing function, which is distinct from that exhibited by other eukaryotic systems.

• Journal of Microbiology
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• v.41 no.3
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• pp.239-247
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• 2003

The LRV1-4 capsid protein possesses an endoribonuclease activity that is responsible for the single site-specific cleavage in the 5' untranslated region (UTR) of its own viral RNA genome and the formation of a conserved stem-loop structure (stem-loop IV) in the UTR is essential for the accurate RNA cleavage by the capsid protein. To delineate the nucleotide sequences, which are essential for the correct formation of the stem-loop structure for the accurate RNA cleavage by the viral capsid protein, a wildtype minimal RNA transcript (RNA 5' 249-342) and several synthetic RNA transcripts encoding point-mutations in the stem-loop region were generated in an in vitro transcription system, and used as substrates for the RNA cleavage assay and RNase mapping studies. When the RNA 5' 249-342 transcript was subjected to RNase T1 and A mapping studies, the results showed that the predicted RNA secondary structure in the stem-loop region using FOLD analysis only existed in the presence of Mg$\^$2＋/ ions, suggesting that the metal ion stabilizes the stem-loop structure of the substrate RNA in solution. When point-mutated RNA substrates were used in the RNA cleavage assay and RNase T1 mapping study, the specific nucleotide sequences in the stem-loop region were not required for the accurate RNA cleavage by the viral capsid protein, but the formation of a stem-loop like structure in a region (nucleotides from 267 to 287) stabilized by Mg$\^$2＋/ ions was critical for the accurate RNA cleavage. The RNase T1 mapping and EMSA studies revealed that the Ca$\^$2＋/ and Mn$\^$2＋/ ions, among the reagents tested, could change the mobility of the substrate RNA 5' 249-342 on a gel similarly to that of Mg$\^$2＋/ ions, but only Ca$\^$2＋/ ions identically showed the stabilizing effect of Mg$\^$2＋/ ions on the stem-loop structure, suggesting that binding of the metal ions (Mg$\^$2＋/ or Ca$\^$2＋/) onto the RNA substrate in solution causes change and stabilization of the RNA stem-loop structure, and only the substrate RNA with a rigid stem-loop structure in the essential region can be accurately cleaved by the LRV1-4 viral capsid protein.

• IMMUNE NETWORK
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• v.11 no.3
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• pp.135-154
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• 2011

The great discovery of microRNAs (miRNAs) has revolutionized current cell biology and medical science. miRNAs are small conserved non-coding RNA molecules that post-transcriptionally regulate gene expression by targeting the 3' untranslated region of specific messenger RNAs for degradation or translational repression. New members of the miRNA family are being discovered on a daily basis and emerging evidence has demonstrated that miRNAs play a major role in a wide range of developmental process including cell proliferation, cell cycle, cell differentiation, metabolism, apoptosis, developmental timing, neuronal cell fate, neuronal gene expression, brain morphogenesis, muscle differentiation and stem cell division. Moreover, a large number of studies have reported links between alterations of miRNA homeostasis and pathological conditions such as cancer, psychiatric and neurological diseases, cardiovascular disease, and autoimmune disease. Interestingly, in addition, miRNA deficiencies or excesses have been correlated with a number of clinically important diseases ranging from cancer to myocardial infarction. miRNAs can repress the gene translation of hundreds of their targets and are therefore well-positioned to target a multitude of cellular mechanisms. As a consequence of extensive participation in normal functions, it is quite logical to ask the question if abnormalities in miRNAs should have importance in human diseases. Great discoveries and rapid progress in the past few years on miRNAs provide the hope that miRNAs will in the near future have a great potential in the diagnosis and treatment of many diseases. Currently, an explosive literature has focussed on the role of miRNA in human cancer and cardiovascular disease. In this review, I briefly summarize the explosive current studies about involvement of miRNA in various human cancers and cardiovascular disease.