• BMB Reports
• /
• v.43 no.6
• /
• pp.438-444
• /
• 2010

Dnd (dead end) gene encodes an RNA binding protein and is specifically expressed in primordial germ cells (PGCs) as a vertebrate-specific component of the germ plasma throughout embryogenesis. By utilizing a technique of specific nucleic acids associated with proteins (SNAAP), 13 potential target mRNAs of zebrafish Dnd (ZDnd) protein were identified from 8-cell embryo, and 8 target mRNAs have been confirmed using an RT-PCR analysis. Of the target mRNAs, the present study is focused on the regulation of geminin, which is an inhibitor of DNA replication. Using electrophoretic mobility shift assay (EMSA), we demonstrated that ZDND protein bound the 67-nucleotide region from 864 to 931 in the 3'UTR of geminin mRNA, a sequence containing 60.29% of uridine. Results from a dual-luciferase assay in HEK293 cells showed that ZDND increases the translation of geminin. Taken together, the identification of target mRNA for ZDnd will be helpful to further explore the biological function of Dnd in zebrafish germ-line development as well as in cancer cells.

• Journal of Gastric Cancer
• /
• v.23 no.2
• /
• pp.340-354
• /
• 2023

Purpose: Gastric cancer (GC) is the second most lethal cancer globally and is associated with poor prognosis. Fatty acid-binding proteins (FABPs) can regulate biological properties of carcinoma cells. FABP5 is overexpressed in many types of cancers; however, the role and mechanisms of action of FABP5 in GC remain unclear. In this study, we aimed to evaluate the clinical and biological functions of FABP5 in GC. Materials and Methods: We assessed FABP5 expression using immunohistochemical analysis in 79 patients with GC and evaluated its biological functions following in vitro and in vivo ectopic expression. FABP5 targets relevant to GC progression were determined using RNA sequencing (RNA-seq). Results: Elevated FABP5 expression was closely associated with poor outcomes, and ectopic expression of FABP5 promoted proliferation, invasion, migration, and carcinogenicity of GC cells, thus suggesting its potential tumor-promoting role in GC. Additionally, RNA-seq analysis indicated that FABP5 activates immune-related pathways, including cytokine-cytokine receptor interaction pathways, interleukin-17 signaling, and tumor necrosis factor signaling, suggesting an important rationale for the possible development of therapies that combine FABP5-targeted drugs with immunotherapeutics. Conclusions: These findings highlight the biological mechanisms and clinical implications of FABP5 in GC and suggest its potential as an adverse prognostic factor and/or therapeutic target.

• The Korean Journal of Physiology and Pharmacology
• /
• v.6 no.2
• /
• pp.101-107
• /
• 2002

In the present study, the postnatal developmental changes in the expressional levels of cardiac sarcoplasmic reticulum (SR) $Ca^{2+}$ regulatory proteins, i.e. $Ca^{2+}-ATPase,$ phospholamban, and $Ca^{2+}$ release channel, were investigated. Both SR $Ca^{2+}-ATPase$ and phospholamban mRNA levels were about 35% of adult levels at birth and gradually increased to adult levels. Protein levels of both SR $Ca^{2+}-ATPase$ and phospholamban, which were measured by quantitative immunoblotting, were closely correlated with the mRNA levels. The initial rates of $Ca^{2+}$ uptake at birth were about 40% of adult rates and also increased gradually during the myocardial development. Consequently, the relative phospholamban/$Ca^{2+}-ATPase$ ratio was 1 in developmental hearts. $Ca^{2+}$ release channel (ryanodine receptor) mRNA was about $50{\sim}60%$ at birth and increased gradually to adult level throughout the postnatal rat heart development. $^3[H]ryanodine$ binding increased gradually during postnatal myocardial development, which was closely correlated with ryanodine mRNA expression levels during the development except the ryanodine mRNA level at birth. These findings indicate that cardiac SR $Ca^{2+}-ATPase,$ phospholamban, and $Ca^{2+}$ release channel are expressed coordinately, which may be necessary for intracellular $Ca^{2+}$ regulation during the rat heart development.

• Molecules and Cells
• /
• v.40 no.1
• /
• pp.54-65
• /
• 2017

Although cancer/testis antigen DDX53 confers anti-cancer drug-resistance, the effect of DDX53 on cancer stem cell-like properties and autophagy remains unknown. MDA-MB-231 ($CD133^+$) cells showed higher expression of DDX53, SOX-2, NANOG and MDR1 than MDA-MB-231 ($CD133^-$). DDX53 increased in vitro self-renewal activity of MCF-7 while decreasing expression of DDX53 by siRNA lowered in vitro self-renewal activity of MDA-MB-231. DDX53 showed an interaction with EGFR and binding to the promoter sequences of EGFR. DDX53 induced resistance to anti-cancer drugs in MCF-7 cells while decreased expression of DDX53 by siRNA increased the sensitivity of MDA-MB-231 to anti-cancer drugs. Negative regulators of DDX53, such as miR-200b and miR-217, increased the sensitivity of MDA-MB-231 to anti-cancer drugs. MDA-MB-231 showed higher expression of autophagy marker proteins such as ATG-5, $pBeclin1^{Ser15}$ and LC-3I/II compared with MCF-7. DDX53 regulated the expression of marker proteins of autophagy in MCF-7 and MDA-MB-231 cells. miR-200b and miR-217 negatively regulated the expression of autophagy marker proteins. Chromatin immunoprecipitation assays showed the direct regulation of ATG-5. The decreased expression of ATG-5 by siRNA increased the sensitivity to anti-cancer drugs in MDA-MB-231 cells. In conclusion, DDX53 promotes stem cell-like properties, autophagy, and confers resistance to anti-cancer drugs in breast cancer cells.

• Environmental Analysis Health and Toxicology
• /
• v.19 no.3
• /
• pp.261-269
• /
• 2004

Although the biological functions of metallothioneins (MTs) are still being investigated, they have been suggested to be involved in detoxification of heavy metals, scavenging of free radicals, and protection against alkylating agents. MTs have been reported to be induced in most of animal tissues by heavy metals such as zinc, copper, mercury and cadmium, and the proteins have binding affinities to the metals. However, the presence or induction of MTs was reported not to be clear in leydig cells, which produce testosterone for the maturation of spermatozoa in male testes. In this study, we investigated the inducibility of metallothionein isomers by cadmium in cultured mouse leydig cells. Total RNA was extracted from the near confluent grown leydig cells and RT-PCR was Performed using the Primers which were synthesized on the basis of MT-1, 2, 3 and 4 cDNA from GenBank database. As results, MT-1 and MT-2 mRNA were found to be expressed in cadmium non-treated control cells and MT 1 mRNA expression was dose-dependent when leydig cells were treated with cadmium chloride. But MT-3 which is known to be brain specific and MT-4 which is another isoform of metallothionein, were not expressed. Other genes induced or depressed in cadmium treated leydig cells were also identified by microarray techniques.

• Genomics & Informatics
• /
• v.19 no.3
• /
• pp.29.1-29.10
• /
• 2021

In our previous studies, we have demonstrated the association of certain variants of the thyroid-stimulating hormone receptor (TSHR), thyroid peroxidase (TPO), and thyroglobulin (TG) genes with congenital hypothyroidism. Herein, we explored the mechanistic basis for this association using different in silico tools. The mRNA 3'-untranslated region (3'-UTR) plays key roles in gene expression at the post-transcriptional level. In TSHR variants (rs2268477, rs7144481, and rs17630128), the binding affinity of microRNAs (miRs) (hsa-miR-154-5p, hsa-miR-376a-2-5p, hsa-miR-3935, hsa-miR-4280, and hsa-miR-6858-3p) to the 3'-UTR is disrupted, affecting post-transcriptional gene regulation. TPO and TG are the two key proteins necessary for the biosynthesis of thyroid hormones in the presence of iodide and H₂O₂. Reduced stability of these proteins leads to aberrant biosynthesis of thyroid hormones. Compared to the wild-type TPO protein, the p.S398T variant was found to exhibit less stability and significant rearrangements of intra-atomic bonds affecting the stoichiometry and substrate binding (binding energies, ΔG of wild-type vs. mutant: -15 vs. -13.8 kcal/mol; and dissociation constant, K_d of wild-type vs. mutant: 7.2E^-12 vs. 7.0E^-11 M). The missense mutations p.G653D and p.R1999W on the TG protein showed altered ΔG(0.24 kcal/mol and 0.79 kcal/mol, respectively). In conclusion, an in silico analysis of TSHR genetic variants in the 3'-UTR showed that they alter the binding affinities of different miRs. The TPO protein structure and mutant protein complex (p.S398T) are less stable, with potentially deleterious effects. A structural and energy analysis showed that TG mutations (p.G653D and p.R1999W) reduce the stability of the TG protein and affect its structure-functional relationship.

• BMB Reports
• /
• v.47 no.4
• /
• pp.233-238
• /
• 2014

Polypyrimidine tract-binding protein 1 (PTBP1) and its brain-specific homologue, PTBP2, are associated with pre-mRNAs and influence pre-mRNA processing, as well as mRNA metabolism and transport. They play important roles in neural differentiation and glioma development. In our study, we detected the expression of the two proteins in glioma cells and predicted that they may be sumoylated using SUMOplot analyses. We confirmed that PTBP1 and PTBP2 can be modified by SUMO1 with co-immunoprecipitation experiments using 293ET cells transiently co-expressing SUMO1 and either PTBP1 or PTBP2. We also found that SUMO1 modification of PTBP2 was enhanced by Ubc9 (E2). The mutation of the sumoylation site (Lys137) of PTBP2 markedly inhibited its modification by SUMO1. Interestingly, in T98G glioma cells, the level of sumoylated PTBP2 was reduced compared to that of normal brain cells. Overall, this study shows that PTBP2 is posttranslationally modified by SUMO1.

• IMMUNE NETWORK
• /
• v.2 no.3
• /
• pp.150-157
• /
• 2002

Background: Although Tat plays a role as a potent transactivator in the viral gene expression from the Human Immunodeficiency Virus type 1 long terminal repeat (HIV-1 LTR), it does not function efficiently in rodent cells implying the absence of a human specific factor essential for Tat-medicated transactivation in rodent cells. In previous experiments, we demonstrated that one of chimeric forms of TAR (transacting responsive element) of HIV-1 LTR compensated the restriction in rodent cells. Methods: To characterize the nature of the compensation, we tested the effects of several upstream binding factors of HIV-1 LTR by simple substitution, and also examined the role of the configuration of the upstream binding factor(s) indirectly by constructing spacing mutants that contained insertions between Sp1 and TATA box on Tat-mediated transactivation. Results: Human Sp1 had no effect whereas its associated factors displayed differential effects in human and rodent cells. In addition, none of the spacing mutants tested overcame the restriction in rodent cells. Rather, when the secondary structure of the chimeric HIV-1 TAR construct was destroyed, the compensation in rodent cells was disappeared. Interestingly, the proper interaction between Sp1 and TATA box binding proteins, which is essential for Tat-dependent transcription, was dispensable in rodent cells. Conclusion: This result suggests that the human-specific Tat cofactor acts to allow Tat to interact effectively in a ribonucleoprotein complex that includes Tat, cellular factors, and TAR RNA, rather than be associated with the HIV-1 LTR upstream DNA binding factors.

• Journal of Animal Science and Technology
• /
• v.46 no.6
• /
• pp.909-916
• /
• 2004

CCAAT/enhancer binding proteins(C/EBP) are a group of transcription factors expressed during preadipocyte differentiation. In the C/EBPs, C/EBPa plays an important role in lipid deposition and adipocyte differentiation. In this studies, we report the identification, characterization, and expression of a Hanwoo CIEBP$\alpha$ The Hanwoo C/EBP$\alpha$DNA includes a 1059 bp open reading frame encoding a protein of 353 amino acids. The CIEBPa amino acid sequences of the Hanwoo show strong conservation with the corresponding sequences reported in other species. The distribution of C/EBP$\alpha$ mRNA in various tissues of Hanwoo aged 12 months were investigated using Northern blotting analysis. The highest expression was detected in adipose tissue and more lower expression was detected in colon and lung. We also identified expression of C/EBPa mRNA in Hanwoo sirloin and adipose tissue aged 12, 26, and 30 months by real-time RT-PCR. The higest expression were detected at 26 months in the sirloin and at 12 and 26 months in the adipose tissue.

• Proceedings of the Korean Biophysical Society Conference
• /
• 1999.06a
• /
• pp.43-43
• /
• 1999

Signal peptides of secretary proteins interact with various membranes and non-membrane components during the translocation. We investigated the interaction of signal peptides of ribose binding protein (RBP) with Escherichia coli (E.coli) signal recognition particle (SRP), SecA and lipid bilayer. Previous studies showed that the functional signal peptides inhibit the GTPase activity of E.coli SRP which consisted of F로 and 4.5S RNA.(omitted)