• BMB Reports
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• v.32 no.6
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• pp.594-598
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• 1999

The Jun and Fos families of eukaryotic transcription factors form heterodimers capable of binding to their cognate DNA enhancer elements. We are interested in searching for inhibitors or antagonists of the binding of the Jun-Fos heterodimer to the activator protein-1 (AP-1) site. The basic-region leucine zipper (bZIP) domain of c-Fos was expressed as a fusion protein with glutathione S-transferase, and allowed to form a heterodimer with the bZIP domain of c-Jun. The heterodimer was bound to glutathione-agarose, to which were added radiolabeled AP-1 nucleotides. After thorough washing, the gel-bound radioactivity was counted. The assay is faster than the coventional electrophoretic mobility shift assay because the gel electrophoresis step and the autoradiography step are eliminated. Moreover, the assay is very sensitive, allowing the detection of picomolar quantities of nucleotides, and is not affected by up to 50% dimethylsulfoxide, a solvent for hydrophobic inhibitors. Curcumin and dihydroguaiaretic acid, recently known inhibitors of Jun-Fos-DNA complex formation, were applied to this Jun-GST-fused Fos system and revealed to decrease the dimer-DNA binding.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.12
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• pp.4871-4876
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• 2014

Background: We sought to evaluate the role of tumor associated macrophages (TAMs) on the promotion of coal tar pitch extract (CTPE)-induced tumorigenesis of human bronchial epithelial cells (BEAS-2B) and tumor metastasis in nude mice, and related mechanisms. Materials and Methods: BEAS-2B cells were first treated with 2.4 mg/mL CTPE for 72 hours. After removal of CTPE, the cells were continuously cultured and passaged using trypsin-EDTA. THP-1 cells were used as macrophage-like cells. BEAS-2B cells under different conditions (n=6/group) were injected into the back necks of nude mice, and alterations of tumor xenograft growth, indicative of tumorigenicity, and tumor metastasis were determined. Pathological changes (tumor nests and microvascular lesions) of HE-stained tumor tissues were also evaluated. The expression of AP-1(c-Jun) in xenografts and metastatic tumors was determined using immunohistochemistry. Results: Tumor size and weight in nude mice transplanted with the mixture of CTPE-induced passage 30 BEAS-2B and THP-1 cells (2:1) were increased compared to those from the CTPE-treated BEAS-2B cells at passage 30 alone at different observation time points. Tumor metastasis to lymph nodes and liver was only detected after transplantation of a mixture the two kinds of cells. The numbers of tumor nests and microvascular lesions, and the expression levels of AP-1 (c-Jun) in tumors from the mixture of two kinds of cells were increased apparently in contrast to those in tumor from the CTPE-treated BEAS-2B cells of passage 30 alone. In addition, there was positive correlation between AP-1 (c-Jun) expression level and the number of microvascular lesions, or between AP-1 (c-Jun) expression level and tumor metastasis in these two groups. Conclusions: TAMs not only facilitate tumorigenesis transformation of CTPE-induced BEAS-2B cells, but also promote tumor growth, angiogenesis and metastasis in nude mice in vivo, which may be mediated by AP-1.

• Journal of Nutrition and Health
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• v.35 no.1
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• pp.53-59
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• 2002

Green tea has been recognized as a favorite beverage for centuries in Easter and Westers cultures. Recently, anti-tumor effects of green tea constituents have received increasing attention. However, the mechanism of catechin-mediated cytotoxicity against tumor cells remains to be elusive. To elucidate the mechanical insights of anti-tumor effects, (-)epigallocatechin-gallate(EGCG) of catechin was applied to human lung cancer A549 cells. (-)EGCG induced the death of A549 cells, which was revealed as apoptosis in DNA fragmentation assay. (-)EGCG induced the activation of caspase family cysteine proteases including capase-3, -8 and -9 proteases in A549 cells. Furthermore, (-)EGCG increased the phosphotransferase activity of c-Jun N-terminal kinase 1JNK 1), which further induced tole transcriptional activation of activating protein-1(AP-1) in A549 cells. We suggest that (-)EGCG-induced apotosis of A549 cells is mediated by signaling pathway involving caspase family cysteine protease, JNK1 and transcription factor, AP-1.

• Molecules and Cells
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• v.28 no.6
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• pp.545-551
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• 2009

Mitogen-activated protein kinases (MAPK) affect the activation of activator protein-1 (AP-1), which plays an important role in regulating a range of cellular processes. However, the roles of these signaling factors on hydrogen peroxide ($H_2O_2$)-induced cell death are unclear. This study examined the effects of $H_2O_2$ on the activation of MAPK and AP-1 by exposing the cells to $H_2O_2$ generated by either glucose oxidase or a bolus addition. Exposing BJAB or Jurkat cells to $H_2O_2$ affected the activities of MAPK differently according to the method of $H_2O_2$ exposure. $H_2O_2$ increased the AP-1-DNA binding activity in these cells, where continuously generated $H_2O_2$ led to an increase in mainly the c-Fos, FosB and c-Jun proteins. The c-Jun-$NH_2$-terminal kinase (JNK)-mediated activation of c-Jun was shown to be related to the $H_2O_2$-induced cell death. However, the suppression of $H_2O_2$-induced oxidative stress by either JNK inhibitor or c-Jun specific antisense transfection was temporary in the cells exposed to glucose oxidase but not to a bolus $H_2O_2$. This was associated with the disruption of death signaling according to the severe and prolonged depletion of reduced glutathione. Overall, these results suggest that $H_2O_2$ may decide differently the mode of cell death by affecting the intracellular redox state of thiol-containing antioxidants, and this depends more closely on the duration exposed to $H_2O_2$ than the concentration of this agent.

• BMB Reports
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• v.52 no.6
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• pp.385-390
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• 2019

Leptin, an adipokine regulating energy metabolism, appears to be associated with breast cancer progression. Insulin-like growth factor-1 (IGF-1) mediates the pathogenesis of breast cancer. The regulation of IGF-1 expression by leptin in breast cancer cells is unclear. Here, we found that leptin upregulates IGF-1 expression at the transcriptional level in breast cancer cells. Activating protein-1 (AP-1)-binding element within the proximal region of IGF-1 was necessary for leptin-induced IGF-1 promoter activation. Forced expression of AP-1 components, c-FOS or c-JUN, enhanced leptin-induced IGF-1 expression, while knockdown of c-FOS or c-JUN abrogated leptin responsiveness. All three MAPKs (ERK1/2, JNK1/2, and p38 MAPK) mediated leptin-induced IGF-1 expression. These results suggest that leptin contributes to breast cancer progression through the transcriptional upregulation of leptin via the MAPK pathway.

• Molecules and Cells
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• v.46 no.6
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• pp.387-398
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• 2023

Microtubule acetylation has been proposed as a marker of highly heterogeneous and aggressive triple-negative breast cancer (TNBC). The novel microtubule acetylation inhibitors GM-90257 and GM-90631 (GM compounds) cause TNBC cancer cell death but the underlying mechanisms are currently unknown. In this study, we demonstrated that GM compounds function as anti-TNBC agents through activation of the JNK/AP-1 pathway. RNA-seq and biochemical analyses of GM compound-treated cells revealed that c-Jun N-terminal kinase (JNK) and members of its downstream signaling pathway are potential targets for GM compounds. Mechanistically, JNK activation by GM compounds induced an increase in c-Jun phosphorylation and c-Fos protein levels, thereby activating the activator protein-1 (AP-1) transcription factor. Notably, direct suppression of JNK with a pharmacological inhibitor alleviated Bcl2 reduction and cell death caused by GM compounds. TNBC cell death and mitotic arrest were induced by GM compounds through AP-1 activation in vitro. These results were reproduced in vivo, validating the significance of microtubule acetylation/JNK/AP-1 axis activation in the anti-cancer activity of GM compounds. Moreover, GM compounds significantly attenuated tumor growth, metastasis, and cancer-related death in mice, demonstrating strong potential as therapeutic agents for TNBC.

• Microbiology and Biotechnology Letters
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• v.48 no.3
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• pp.310-315
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• 2020

Orostachys japonicus (O. japonicus) is known as a medicinal plant for the treatment of various symptoms. This study investigated the anti-inflammatory effect of the hexane fraction from O. japonicus (OJH) on the LPS-stimulated response in RAW 264.7 macrophage cells. This study was conducted to confirm the effect of cell cytotoxicity and production of reactive oxygen species (ROS) in OJH-treated macrophage cells. Additionally, pro-inflammatory cytokines and transcription factors were determined using RT-PCR and western blotting assay. OJH showed no change in lactate dehydrogenase (LDH) levels and exhibited reduced ROS levels in LPS-induced inflammatory cells. Moreover, OJH significantly suppressed the mRNA levels of proinflammatory cytokines, including IL-1β, IL-2, IL-6, TNF-α, and IP-10. Furthermore, OJH effectively inhibited the protein levels of AP-1 (p-c-Jun and p-c-Fos) and p-IRF3 in a dose-dependent manner. In conclusion, our results demonstrate that OJH exhibits strong anti-inflammatory activities via regulation of inflammatory factors.

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

• Bulletin of the Korean Chemical Society
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• v.25 no.12
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• pp.1769-1774
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• 2004

Jun/Fos, a crucial factor in transmitting the tumor-promoting signal from the extracellular environment to the nuclear transcription machinery, has a dimerization interface possessing several coiled structural properties. Jun and Fos can interact with the DNA regulatory region, AP-1 (Activator Protein-1), which is composed of 5'-TGAC/GTCA-3'.$^1$ Curcumin is a well-known anticancer and anti-inflammatory compound.$^{2,3}$ It also acts as an inhibitor of the Jun-Fos function. c-Fos and c-Jun with a bZIP region are overexpressed in BL21 E. coli and purified with an $Ni^{2+}$ affinity column. The inhibitors of Fos-Jun-AP-1 complex formation were searched through the EMSA (electrophoresis mobility shift assay) experiment, and new curcuminoids were synthesized and investigated as to their inhibitory effect on the same system. Two curcuminoids showed a stronger inhibitory effect than curcumin. This inhibitory activity was quantified with EMSA. 1,7-bis(4-methyl)-1,6-heptadiene-3,5-dione (BJC003) and 1,7-bis(4-hydroxy-5-methoxy-3-nitrophenyl)-1,6-heptadiene-3,5-dione (BJC005) showed remarkably high inhibitory activities. $IC_{50}$ of 1,7-bis(4-methyl)-1,6-heptadiene-3,5-dione (BJC003) and 1,7-bis(4-hydroxy-5-methoxy-3-nitrophenyl)-1,6-heptadiene-3,5-dione (BJC005) are 8.98 ${\mu}M$ and 5.40 ${\mu}M$, respectively. However, 1,7-bis(4-methyl-3-nitrophenyl)-1,6-heptadiene-3,5-dione (BJC004) did not show inhibitory activity.

• Bulletin of the Korean Chemical Society
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• v.27 no.4
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• pp.535-538
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• 2006

In our previous studies, we have observed that curcumin and momordin I isolated from Ampelopsis radix inhibit the formation of Fos-Jun-activation protein-1 (AP-1) DNA complex. We have screened more effective compounds which have a 5-membered ring framework like momordin I and have modified disaccharide or carboxylic acid portions in momordin I. We synthesized momordin I derivatives according to the published method with slight modification. Synthetic momordin I derivatives showed remarkable inhibitory activities on Fos-Jun-AP-1 DNA complex formation results in in vitro assays. The $IC_{50}$ values of momordin I derivatives were about 4.0 $\mu$M in an electrophoretic mobility shift assay (EMSA). This value is about 125 times higher than that of curcumin and about 12 times higher than that for curcumin derivative C1, and moreover about 30 times higher than that for momordin I. We found momordin I derivatives (a) and (b) are the strongest inhibitory compound for Fos-Jun-AP-1 DNA complex formation.