• BMB Reports
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• v.50 no.6
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• pp.318-322
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• 2017

Brain cytoplasmic 200 RNA (BC200 RNA) is a neuron-specific non-coding RNA, implicated in the inhibition of local synaptodendritic protein synthesis, and is highly expressed in some cancer cells. Although BC200 RNA has been shown to inhibit translation in vitro, the cellular location of this inhibition is unknown. In this study, we used a BC200 RNA-recognizing antibody to identify the cellular locations of BC200 RNA in HeLa cervical carcinoma cells. We observed punctate signals in both the cytoplasm and nucleus, and further discovered that BC200 RNA co-localized with the p-body decapping enzyme, DCP1A, and the heterogeneous nuclear ribonucleoprotein E2 (hnRNP E2). The latter is a known BC200 RNA-binding partner protein and a constituent of p-bodies. This suggests that BC200 RNA is localized to p-bodies via hnRNP E2.

• BMB Reports
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• v.48 no.7
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• pp.367-368
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• 2015

It has long been thought that glucocorticoid receptor (GR) functions as a DNA-binding transcription factor in response to its ligand (a glucocorticoid) and thus regulates various cellular and physiological processes. It is also known that GR can bind not only to DNA but also to mRNA; this observation points to the possible role of GR in mRNA metabolism. Recent data revealed a molecular mechanism by which binding of GR to target mRNA elicits rapid mRNA degradation. GR binds to specific RNA sequences regardless of the presence of a ligand. In the presence of a ligand, however, the mRNA-associated GR can recruit PNRC2 and UPF1, both of which are specific factors involved in nonsense-mediated mRNA decay (NMD). PNRC2 then recruits the decapping complex, consequently promoting mRNA degradation. This mode of mRNA decay is termed "GR-mediated mRNA decay" (GMD). Further research demonstrated that GMD plays a critical role in chemotaxis of immune cells by targeting CCL2 mRNA. All these observations provide molecular insights into a previously unappreciated function of GR in posttranscriptional regulation of gene expression. [BMB Reports 2015; 48(7): 367-368]

• Molecules and Cells
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• v.42 no.4
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• pp.356-362
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• 2019

The binding of MS2 bacteriophage coat protein (MCP) to MS2 binding site (MBS) RNA stem-loop sequences has been widely used to label mRNA for live-cell imaging at single-molecule resolution. However, concerns have been raised recently from studies with budding yeast showing aberrant mRNA metabolism following the MS2-GFP labeling. To investigate the degradation pattern of MS2-GFP-labeled mRNA in mammalian cells and tissues, we used Northern blot analysis of ${\beta}$-actin mRNA extracted from the Actb-MBS knock-in and $MBS{\times}MCP$ hybrid mouse models. In the immortalized mouse embryonic cell lines and various organ tissues derived from the mouse models, we found no noticeable accumulation of decay products of ${\beta}$-actin mRNA compared with the wild-type mice. Our results suggest that accumulation of MBS RNA decay fragments does not always happen depending on the mRNA species and the model organisms used.

• Korean Journal of Microbiology
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• v.54 no.4
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• pp.325-329
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• 2018

Thp1/PCID2, PCI domain-containing protein, is a component of the evolutionally conserved TREX-2 complex linking mRNA transcription and export. In fission yeast, Schizosaccharomyces pombe, the pci2 (SPBC1105.07c) gene encodes a PCI domain-containing protein that is predicted as a fission yeast orthologue of Thp1 (in budding yeast)/PCID2 (in human). Repression of pci2 expression inhibited both growth and mRNA export. And over-expression of pci2 also exhibited growth retardation with slight accumulation of $poly(A)^+$ RNA in the nucleus. Moreover, yeast two-hybrid and co-immunoprecipitation analysis showed that the Pci2 protein physically interacted with Sac3 and Dss1, which are members of TREX-2 complex. These observations support that the S. pombe Pci2 protein, as a component of TREX-2 complex, is implicated in mRNA export.

• Journal of dental hygiene science
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• v.13 no.3
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• pp.321-329
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• 2013

Although it has been reported that lysyl oxidase (LOX) is involved in odontoblastic differentiation, the role of LOX on odontoblastic differentiation by hydrogen peroxide ($H_2O_2$) have not been clarified. In the present study, we investigated whether $H_2O_2$, reactive oxygen species (ROS), is modulated the messenger RNA (mRNA) expression and activity of LOX during odontoblastic differentiation of human dental pulp (HDP) cells. The mRNA expression was quantified by reverse transcriptase polymerase chain reaction (RT-PCR) analysis, and LOX enzyme activity was measured by high sensitive fluorescent assay. Expression of the odontoblastic differentiation marker genes were assessed in the presence and absence of specific small interfering RNAs (siRNAs) of the LOX and LOXL. The $H_2O_2$-induced mRNA expression of LOX family was significant reduction of LOX, LOXL, and LOXL3 mRNA levels in HDP cells. LOX enzyme activity was increased at $H_2O_2$ 0.3 mM for 24 hours. The mRNA expression of alkaline phosphatase (ALP), osteopontin (OPN), and osteocalcin (OCN) was inhibited by LOX- and LOXL-specific siRNAs whereas the mRNA expression of dentin matrix protein1 (DMP1), and dentin sialophosphoprotein (DSPP) was inhibited by LOX-specific siRNA. In LOX enzyme activity, siRNA-induced knockdown of both LOX and LOXL inhibited the total amine oxidase activity in HDP cells, as in the case of mRNA expression. In conclusion, the essential role of $H_2O_2$ on odontoblastic differentiation suggests that its regulation by LOX may have pharmacologic importance in HDP cells.

• Journal of Microbiology
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• v.33 no.2
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• pp.160-164
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• 1995

The splicing activity of T4 phage td intron RNA has been examined with various Mg$^{2+}$ ions such as MGCl$_{2}$, MgS $O_{4}$ and magnesium acetate using various splicing conditions such as different incubation time and temperature. The maximum splicing of td intron RNA occurred at the concentration of 5 mM MgCl$_{2}$. Raising the Mg$^{2+}$ concentration up to 15 mM appeared to promote P2 delection mutant to overcome the loss of some splicing activity. In both wild type and mutant, a complete hydrolysis of RNA occurred at 30 mM MgCI$_{2}$ MgS $O_{4}$ and magnesium acetate exhibited the rate and pattern of RNA splicing identical to MGCI$_{2}$. The optimal splicing conditions involve the incubation of RNA with 5 mM MgCI$_{2}$ at 58 .deg.C for 15 min. The results suggest that Mg$^{2+}$ may play a key role in the catalytic mechanism of td intron RNA.n RNA.

• Research in Plant Disease
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• v.7 no.3
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• pp.155-163
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• 2001

Two attenuated Cucumber mosaic virus (CMV) isolates, Paf-CMV and Rs2-CMV that had been selected from CMV isolates associated with satellite RNA (satRNA) were tested for cross-protection effect in pepper plants. The viruses selected as attenuated strains appeared to be identical serologically and physically to the challenge virus (Mf-CMV), but they were lower in the dilution end-point of infectivity of crude sap than Mf-CMV When symptoms were observed in several indicator plants after inoculation, Paf-CMV and Rs2-CMV were symptomless or showed mild mosaic symptoms while another satRNA isolate Ap-CMV developed severe mosaic symptoms on the leaves as Mf-CMV. The nucleotide sequences of the satRNAs were determined by sequencing full-length cDNA clones. Paf-, Rs2- and Ap-satRNAs were 386, 335, and 347 nucleotides long, respectively, The sequences were then compared with the other known Y-satRNA, revealing that nucleotide sequences of the satRNAs consisted of 5'- and 3'-terminal conserved regions. However variations occurred on the middle regions of the sequences, especially those related to symptom interference, showing significant differences between Paf-satRNA and other isolates. Infectious transcripts of Paf-satRNA and Rs2-satRNA induced mild mosaic symptoms in pepper plants when supported by genomic RNAs of Mf-CMV. Under greenhouse conditions, Paf-CMV and Rs2-CMV were tested for cross-protection effect in pepper and tobacco (Nicotiana tabacum cv, Xanthi nc) plants against Mf-CMV. No symptoms were developed on the plants vaccinated with Paf-CMV until 3 weeks after inoculation with the virulent strain; however another attenuated isolate, Rs2-CMV, showed less effectiveness in cross-protection. Depending on the concentration of the challenged virus, symptoms sometimes appeared later in the upper leaves. However, in plants challenged with low concentrations (below 0.2 mg/ml) of the challenge inoculum, symptoms caused by the virulent strain did not develop on the plants vaccinated with Paf-CMV. In the field experiments, the number of pepper plants with severe mosaic symptoms in the control plots was progressively increased after transplanting and reached approximately 50% after 50 days. On the other hand, the incidence of mosaic disease appeared very low on the plants that had received the protective inoculation with Paf-CMV.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.8
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• pp.3471-3476
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• 2014

Background: Aberrant expression of the microRNA-29 family is associated with tumorigenesis and cancer progression. As transport carriers, tumor-derived exosomes are released into the extracellular space and regulate multiple functions of target cells. Thus, we assessed the possibility that exosomes could transport microRNA-29c as a carrier and correlations between microRNA-29c and apoptosis of bladder cancer cells. Materials and Methods: A total of 28 cancer and adjacent tissues were examined by immunohistochemistry to detect BCL-2 and MCL-1 expression. Disease was Ta-T1 in 12 patients, T2-T4 in 16, grade 1 in 8, 2 in 8 and 3 in 12. The expression of microRNA-29c in cancer tissues was detected by quantitative reverse transcriptase PCR (QRT-PCR). An adenovirus containing microRNA-29c was used to infect the BIU-87 human bladder cancer cell line. MicroRNA-29c in exosomes was measured by QRT-PCR. After BIU-87 cells were induced by exosomes-derived microRNA-29c, QRT-PCR was used to detect the level of microRNA-29c. Apoptosis was examined by flow cytometry and BCL-2 and MCL-1 mRNA expressions were assessed by reverse transcription-polymerase chain reaction. Western blotting was used to determine the protein expression of BCL-2 and MCL-1. Results: The expressions of BCL-2 and MCL-1 protein were remarkably increased in bladder carcinoma (p<0.05), but was found mainly in the basal and suprabasal layers in adjacent tissues. The expression of microRNA-29c in cancer tissues was negatively correlated with the BCL-2 and MCL-1. The expression level of microRNA-29c in exosomes and BIU-87 cells from the experiment group was higher than that in control groups (p<0.05). Exosome-derived microRNA-29c induced apoptosis (p<0.01). Although only BCL-2 was reduced at the mRNA level, both BCL-2 and MCL-1 were reduced at the protein level. Conclusions: Human bladder cancer cells infected by microRNA-29c adenovirus can transport microRNA-29c via exosomes. Moreover, exosome-derived microRNA29c induces apoptosis in bladder cancer cells by down-regulating BCL-2 and MCL-1.

• Genomics & Informatics
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• v.12 no.2
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• pp.71-75
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• 2014

The tRNA structure contains conserved modifications that are responsible for its stability and are involved in the initiation and accuracy of the translation process. tRNA modification enzymes are prevalent in bacteria, archaea, and eukaryotes. tRNA Gm18 methyltransferase (TrmH) and tRNA $m^1G37$ methyltransferase (TrmD) are prevalent and essential enzymes in bacterial populations. TrmH involves itself in methylation process at the 2'-OH group of ribose at the 18th position of guanosine (G) in tRNAs. TrmD methylates the G residue next to the anticodon in selected tRNA subsets. Initially, $m^1G37$ modification was reported to take place on three conserved tRNA subsets ($tRNA^{Arg}$, $tRNA^{Leu}$, $tRNA^{Pro}$); later on, few archaea and eukaryotes organisms revealed that other tRNAs also have the $m^1G37$ modification. The present study reveals Gm18, $m^1G37$ modification, and positions of $m^1G$ that take place next to the anticodon in tRNA sequences. We selected extremophile organisms and attempted to retrieve the $m^1G$ and Gm18 modification bases in tRNA sequences. Results showed that the Gm18 modification G residue occurs in all tRNA subsets except three tRNAs ($tRNA^{Met}$, $tRNA^{Pro}$, $tRNA^{Val}$). Whereas the $m^1G37$ modification base G is formed only on $tRNA^{Arg}$, $tRNA^{Leu}$, $tRNA^{Pro}$, and $tRNA^{His}$, the rest of the tRNAs contain adenine (A) next to the anticodon. Thus, we hypothesize that Gm18 modification and $m^1G$ modification occur irrespective of a G residue in tRNAs.

• Bulletin of the Korean Chemical Society
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• v.26 no.12
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• pp.2033-2037
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• 2005

In vitro selection was used to isolate $Mg^{2+}$-dependent self-cleaving ribozymes with cis-cleavage activity from a pre-tRNA library having 40-mer random sequences attached to 5'-end of E. coli $tRNA^{Phe}$. After 8 rounds of SELEX (Systematic Evolution of Ligands by Exponential Enrichment), RNA molecules which can self-cleave at the high concentration of $Mg^{2+}$ were isolated. The selected ribozymes can carry out the self-cleavage reaction in the presence of 100 mM $Mg^{2+}$ but not in 10 mM $Mg^{2+}$. The cleavage sites of the ribozymes are located at +3 and +4 of $tRNA^{Phe}$, compared with +1 position of 5'-end cleavage site of pre-tRNA by RNase P. New RNA constructs deprived of its D stem-loop, anticodon stem-loop, variable loop and T stem-loop, respectively showed the cleavage specificity identical to a ribozyme having the intact tRNA structure. Also, the new ribozyme fused with both a ribozyme and $tRNA^{Leu}$ showed the cleavage activities at the various sites within its sequences, different from two sites of position +3 and +4 observed in the ribozyme with $tRNA^{Phe}$. Our results suggest that the selected ribozyme is not structural-specific for tRNA.