• Animal Bioscience
• /
• v.34 no.2
• /
• pp.172-184
• /
• 2021

Objective: Vanin1 (VNN1) is a pantetheinase that can catalyze the hydrolysis of pantetheine to produce pantothenic acid and cysteamine. Our previous studies showed that VNN1 is specifically expressed in chicken liver. In this study, we aimed to investigate the roles of peroxisome proliferators activated receptor α (PPARα) and miRNA-181a-5p in regulating VNN1 gene expression in chicken liver. Methods: 5'-RACE was performed to identify the transcription start site of chicken VNN1. JASPAR and TFSEARCH were used to analyze the potential transcription factor binding sites in the promoter region of chicken VNN1 and miRanda was used to search miRNA binding sites in 3' untranslated region (3'UTR) of chicken VNN1. We used a knock-down strategy to manipulate PPARα (or miRNA-181a-5p) expression levels in vitro to further investigate its effect on VNN1 gene transcription. Luciferase reporter assays were used to explore the specific regions of VNN1 targeted by PPARα and miRNA-181a-5p. Results: Sequence analysis of the VNN1 promoter region revealed several transcription factor-binding sites, including hepatocyte nuclear factor 1α (HNF1α), PPARα, and CCAAT/enhancer binding protein α. GW7647 (a specific agonist of PPARα) increased the expression level of VNN1 mRNA in chicken primary hepatocytes, whereas knockdown of PPARα with siRNA increased VNN1 mRNA expression. Moreover, the predicted PPARα-binding site was confirmed to be necessary for PPARα regulation of VNN1 gene expression. In addition, the VNN1 3'UTR contains a sequence that is completely complementary to nucleotides 1 to 7 of miRNA-181a-5p. Overexpression of miR-181a-5p significantly decreased the expression level of VNN1 mRNA. Conclusion: This study demonstrates that PPARα is an important transcriptional activator of VNN1 gene expression and that miRNA-181a-5p acts as a negative regulator of VNN1 expression in chicken hepatocytes.

• Korean Journal of Plant Tissue Culture
• /
• v.24 no.1
• /
• pp.1-35
• /
• 1997

Fertilized egg, by successive cell divisions, differentiates into different tissues and organs with various structures and functions. Different cells and tissues contain different proteins, products of selective gene expression. Not all the genes in any genomes are equally active, temporal and spatial gene expression being the general rule. Present paper attempts to review the tanscriptional mechanisms or the initiations of transcription from several angles. In some of the organisms the genes in the process of transcription or the genes in the inactive state can be seen under the light microscope. Some bands of Drosophila polytene chromosomes may exhibit a swollen or puff appearance under certain conditions. A puff, unfolded or decondensed form of chromomere, represents sets of intense transcriptional activity or RNA synthesis. The heterochromatic X chromosome whose genes remain inactive in the female mammals can be visualized as a dark staining structure called Barr body, Configuration of chromatin differs between transcribed and nontranscribed chromatin. Modification to the chromatin facilitates RNA synthesis. The movement of large polymerase molecule along the DNA would probably be facilitated if some modifications of the chromatin configuration is effected. Methylation of cytosines in CG sequences is associated with inactive genes. Methylation can play a role in determination of mammalian cells during embryogenesis. Demethylation is necessary for the gene to be expressed during development A histone modification that is also known to be correlated with transcriptional capacity of chromatin is acetylation of the lysine residues of the core histones. Chromatin containing a high level of histone acetylation is very sensitive to DNase 1. For the transcription to occur TBP must first bind to the TATA box. Another TF, TF IIB, then binds to the promoter-TBP complex, facilitating the access of RNA polymerase to the transcription initiation site. As recently as eight years ago researchers assumed that histones were irrelevant to the regulation of gene expression. Histones combine with the DNA to form nucleosome of the chromatin. Histones are vital participant in gene regulation. Histone and basal factors compete for access to TATA box. When DNA is exposed to basal factors before histones are introduced, the basal factors assemble on TATA boxes preventing the access of histones, allowing transcription to occur, for transcription to begin, activator protein at the upstream activation sequence or enhancer must interact with the tail of histone H4 at TATA box and cause the histone role particle to dissociate from the TATA box leading to partial breakup of the histone core particle and allowing the basal factors to bind to the TATA box. New concept of genomic flux in contrast to the old concept of static genome has been developed based on the powerful new molecular techniques. Genomic changes such as repetitive DNAs and transposable elements, it is assumed but not yet proved, may affect some of the developmental patterns that characterize particular cells, tissues, organs, and organisms. In the last decade or so remarkable achievement have been made in the researches of the structures and functions of TFs and the specific target sequences located in promoters or enhancers where these TFs bind. TFs have independent domains that bind DNA and that activate transcription. DNA binding domain of TFs serves to bring the protein into the right location. There are many types of DNA binding domains. Common types of motifs can be found that are responsible for binding to DNA. The motifs are usually quite short and comprise only a small part of the protein structure. Steroid receptors have domains for hormone binding, DNA binding, and activating transcription. The zinc finger motif comprises a DNA binding domain. Leucine zipper consist of a stretch of amino acids with a leucine residue in every seventh position Two proteins form a dimer because they interact by means of leucine zippers on similar α-helical domain. This positions their DNA binding basic domains for interaction with the two halves of a DNA sequence with dyad symmetry of TGACTCA, ACTGAGT.

• BMB Reports
• /
• v.45 no.3
• /
• pp.171-176
• /
• 2012

Receptor activator of NF-${\kappa}B$ ligand (RANKL) triggers the differentiation of bone marrow-derived monocyte/macrophage precursor cells (BMMs) of hematopoietic origin into osteoclasts through the activation of mitogen-activated protein (MAP) kinases and transcription factors. Recently, reactive oxygen species (ROS) and antioxidant enzymes were shown to be closely associated with RANKL-mediated osteoclast differentiation. Although glutaredoxin2 (Glrx2) plays a role in cellular redox homeostasis, its role in RANKL-mediated osteoclastogenesis is unclear. We found that Glrx2 isoform b (Glrx2b) expression is induced during RANKLmediated osteoclastogenesis. Over-expression of Glrx2b strongly enhanced RANKL- mediated osteoclastogenesis. In addition, Glrx2b-transduced BMMs enhanced the expression of key transcription factors c-Fos and NFATc1, but pre-treatment with SB203580, a p38-specific inhibitor, completely blocked this enhancement. Conversely, down-regulation of Glrx2b decreased RANKL- mediated osteoclastogenesis and the expression of c-Fos and NFATc1 proteins. Also, Glrx2b down-regulation attenuated the RANKL-induced activation of p38. Taken together, these results suggest that Glrx2b enhances RANKL-induced osteoclastogenesis via p38 activation.

• Journal of dental hygiene science
• /
• v.17 no.1
• /
• pp.81-86
• /
• 2017

Bacterial infection and smoking are an important risk factors involved in the development and progression of periodontitis. However, the signaling mechanism underlying the host immune response is not fully understood in periodontal lesions. In this study, we determined the expression of janus kinase (JAK)/signal transducer and activator of transcription (STAT) on Porphyromonas gingivalis lipopolysaccharide (LPS)- and nicotine-induced cytotoxicity and the production of inflammatory mediators, using osteoblasts. The cells were cultured with 5 mM nicotine in the presence of $1{\mu}g/ml$ LPS. Cell viability was determined using MTT assay. The role of JAK on inflammatory mediator expression and production, and the regulatory mechanisms involved were assessed via enzyme-linked immunosorbent assay, reverse transcription-polymerase chain reaction, and Western blot analysis. LPS- and nicotine synergistically induced the production of cyclooxgenase-2 (COX-2) and prostaglandin $E_2$ ($PGE_2$) and increased the protein expression of JAK/STAT. Treatment with an JAK inhibitor blocked the production of COX-2 and $PGE_2$ as well as the expression of pro-inflammatory cytokines, such as tumor necrosis factor-${\alpha}$, interleukin-$1{\beta}$ ($IL-1{\beta}$), and IL-6 in LPS- and nicotine-stimulated osteoblasts. These results suggest that JAK/STAT is closely related to the LPS- and nicotine-induced inflammatory effects and is likely to regulate the immune response in periodontal disease associated with dental plaque and smoking.

• Molecules and Cells
• /
• v.23 no.1
• /
• pp.23-29
• /
• 2007

Cyclic AMP-responsive element binding protein (CREB) is known to be associated with angiogenesis. In the present study we investigated the possible role of CREB in the expression of vascular endothelial growth factor (VEGF) by mouse macrophages. Over-expression of CREB increased VEGF secretion by cells of the RAW264.7 mouse macrophage cell line. It also increased the promoter activity of a mouse reporter driven by the VEGF promoter, while a dominant negative CREB (DN-CREB) abrogated the activity, suggesting that CREB mediates VEGF transcription. Forskolin, an adenylyl cyclase activator, stimulated VEGF transcription, and the PKA inhibitor H89 abolished this effect. IFN-${\gamma}$, a potent cytokine, stimulated VEGF expression only in part through the PKA-CREB pathway. These results indicate that PKA phosphorylates CREB and so induces VEGF gene expression. An analysis of mutant promoters revealed that one of the putative CREB responsive elements (CREs), at -399 ~ -388 in the promoter, is critical for CREB-mediated VEGF promoter activity, and the significance of this CRE was confirmed by chromatin immunoprecipitation assays.

• Journal of Pharmacopuncture
• /
• v.22 no.4
• /
• pp.253-259
• /
• 2019

Objectives: It is reported that tumor-associated macrophages (TAMs) contribute to cancer progression by promoting tumor growth and metastasis. The purpose of this study is to investigate the effect of different fractions of Adenophora triphylla var. japonica (AT) on the polarization of macrophages into the M2 phenotype, a major phenotype of TAMs. Methods: We isolated hexane, ethyl acetate, and butanol fractions from crude ethanol extract of AT. The cytotoxicity of AT in RAW264.7 cells was examined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. RAW264.7 cells were polarized into the M2 phenotype by treatment with interleukin (IL)-4 and IL-13. The expression of M2 macrophage marker genes was detected by reverse transcription polymerase chain reaction (RT-PCR). The phosphorylation level of signal transducer and activator of transcription 6 (STAT6) was investigated by western blot analysis. The migration of Lewis lung carcinoma (LLC) cells was examined by transwell migration assay using conditioned media (CM) collected from RAW264.7 cells as a chemoattractant. Results: Among various fractions of AT, the ethyl acetate fraction of AT (EAT) showed the most significant suppressive effect on the mRNA expression of M2 macrophage markers, including arginase-1, interleukin (IL)-10 and mannose receptor C type 1 (MRC-1), up-regulated by treatment of IL-4 and IL-13. In addition, EAT suppressed the phosphorylation of STAT6, a critical regulator of IL-4 and IL-13-induced M2 macrophage polarization. Finally, the increased migration of Lewis lung carcinoma (LLC) cells by CM from M2-polarized RAW264.7 cells was reduced by CM from RAW264.7 cells co-treated with EAT and M2 polarization inducers. Conclusion: We demonstrated that EAT attenuated cancer cell migration through suppression of macrophage polarization toward the M2 phenotype. Additional preclinical or clinical researches are needed to evaluate its regulatory effects on macrophage polarization and anti-cancer activities.

• Rapid Communication in Photoscience
• /
• v.1 no.1
• /
• pp.21-24
• /
• 2012

Influence of gamma rays on the cyanobacterium Synechocystis sp. PCC 6803 cells was investigated in terms of a bidirectional hydrogenase, which is encoded by hoxEFUYH genes and responsible for biohydrogen production. Irradiated cells revealed a substantial change in stoichiometry of photosystems at one day after gamma irradiation at different doses. However, as evaluated by the maximal rate of photosynthetic oxygen evolution, maximal photochemical efficiency of photosystem II, and chlorophyll content, net photosynthesis or photosynthetic capacity was not significantly different between the control and irradiated cells. Instead, transcription of hoxE, hoxH, or lexA, which encodes a subunit of bidirectional hydrogenase or the only transcriptional activator, LexA, for hox genes, was commonly enhanced in the irradiated cells. This transcriptional enhancement was more conspicuously observed immediately after gamma irradiation. In contrast, hydrogenase activities were found to somewhat lower in the irradiated cells. Therefore, we propose that transcription of hox genes should be enhanced by gamma irradiation in a LexA-mediated and possibly photosynthesis-independent manner and that this enhancement might not induce a subsequent increase in hydrogenase activities, probably due to the presence of post-transcriptional and/or post-translational regulatory mechanisms.

• Asian Pacific Journal of Cancer Prevention
• /
• v.16 no.4
• /
• pp.1375-1383
• /
• 2015

Background: Objective Myocardin-related transcription factor (MRTF)-A is a Rho signaling-responsive co-activator of serum response factor (SRF). The purpose of this study is to investigate the role of MRTF-A and AQP1 (aquaporin 1) in pathological vascular remodeling. Materials and Methods: MRTF-A, AQP1 and neointima expression was detected both in the wire injured femoral arteries of wild-type mice and the atherosclerotic aortic tissues of $ApoE^{-/-}$ mice. Expression of ICAM-1, matrix metallopeptidase 9 (MMP-9) and integrin ${\beta}1$ were also assayed. The intercourse relationship between the molecules were investigated by interfering RNA and inhibitor assay. Results: MRTF-A and AQP1 expression were significantly higher in the wire injured femoral arteries of wild-type mice and in the atherosclerotic aortic tissues of $ApoE^{-/-}$ mice than in healthy control tissues. Both in wire-injured femoral arteries in MRTF-A knockout ($Mkl1^{-/-}$) mice and atherosclerotic lesions in $Mkl1^{-/-}$; $ApoE^{-/-}$ mice, neointima formation were significantly attenuated and the expression of AQP1 were significantly decreased. Expression of ICAM-1, matrix metallopeptidase 9 (MMP-9) and integrin ${\beta}1$, three SRF targets and key regulators of cell migration, and AQP1 in injured arteries was significantly weaker in $Mkl1^{-/-}$ mice than in wild-type mice. In cultured vascular smooth muscle cells (VSMCs), knocking down MRTF-A reduced expression of these genes and significantly impaired cell migration. Underlying the increased MRTF-A expression in dedifferentiated VSMCs were the down-regulation of microRNA-300. Moreover, the MRTF-A inhibitor CCG1423 significantly reduced neointima formation following wire injury in mice. Conclusions: MRTF-A could be a novel therapeutic target for the treatment of vascular diseases.

• The Korean Journal of Physiology and Pharmacology
• /
• v.22 no.5
• /
• pp.467-479
• /
• 2018

The aging process induces a plethora of changes in the body including alterations in hormonal regulation and metabolism in various organs including the heart. Aging is associated with marked increase in the vulnerability of the heart to ischemia-reperfusion injury. Furthermore, it significantly hampers the development of adaptive response to various forms of conditioning stimuli (pre/post/remote conditioning). Aging significantly impairs the activation of signaling pathways that mediate preconditioning-induced cardioprotection. It possibly impairs the uptake and release of adenosine, decreases the number of adenosine transporter sites and down-regulates the transcription of adenosine receptors in the myocardium to attenuate adenosine-mediated cardioprotection. Furthermore, aging decreases the expression of peroxisome proliferator-activated receptor gamma co-activator 1-alpha ($PGC-1{\alpha}$) and subsequent transcription of catalase enzyme which subsequently increases the oxidative stress and decreases the responsiveness to preconditioning stimuli in the senescent diabetic hearts. In addition, in the aged rat hearts, the conditioning stimulus fails to phosphorylate Akt kinase that is required for mediating cardioprotective signaling in the heart. Moreover, aging increases the concentration of $Na^+$ and $K^+$, connexin expression and caveolin abundance in the myocardium and increases the susceptibility to ischemia-reperfusion injury. In addition, aging also reduces the responsiveness to conditioning stimuli possibly due to reduced kinase signaling and reduced STAT-3 phosphorylation. However, aging is associated with an increase in MKP-1 phosphorylation, which dephosphorylates (deactivates) mitogen activated protein kinase that is involved in cardioprotective signaling. The present review describes aging as one of the major confounding factors in attenuating remote ischemic preconditioning-induced cardioprotection along with the possible mechanisms.

• Biomolecules & Therapeutics
• /
• v.20 no.1
• /
• pp.57-61
• /
• 2012

The matrix metalloproteinase (MMP) family is involved in the breakdown of the extracellular matrix during normal physiological processes such as embryonic development, reproduction, and tissue remodeling, as well as in disease processes such as pathological aging, arthritis, and metastasis. Oxidative conditions generate reactive oxygen species (ROS) (e.g., hydrogen peroxide [$H_2O_2$]) in cells, which subsequently induce the synthesis of matrix metalloproteinase-1 (MMP-1). MMP-1, an interstitial collagenase, in turn stimulates an aging phenomenon. In this study, baicalein (5,6,7-trihydroxyfl avone) was investigated for its in vitro activity against $H_2O_2$-induced damage using a human skin keratinocyte model. Baicalein pretreatment signifi cantly inhibited $H_2O_2$-induced up-regulation of MMP-1 mRNA, MMP-1 protein expression and MMP-1 activity in cultured HaCaT keratinocytes. In addition, baicalein decreased the transcriptional activity of activator protein-1 (AP-1) and the expression of c-Fos and c-Jun, both components of the heterodimeric AP-1 transcription factor. Furthermore, baicalein reduced phosphorylation of extracellular signal-regulated kinase (ERK) and c-Jun-N-terminal kinase (JNK), which are upstream of the AP-1 transcription factor. The results of this study suggest that baicalein is involved in the inhibition of oxidative stress-induced expression of MMP-1 via inactivation of the ERK/JNK/AP-1 signaling pathway.