• Journal of the Korean Magnetic Resonance Society
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• v.25 no.2
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• pp.17-23
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• 2021

High mobility group protein B1 (HMGB1) is a highly conserved, non-histone, chromatin associated nuclear protein encoded by HMGB1 gene. HMGB1 proteins may be general co-factors in Hox-mediated transcriptional activation that facilitate the access of Hox proteins to specific DNA targets. It is unclear that the exact binding interface of Hoxc9DBD and HMGB1. To identify the interface and binding affinity of Hoxc9DBD and HMGB1 A-box, the paramagnetic probe, MTSL was used in NMR titration experiment. It is attached to the N-terminal end of HMGB1 A-box by reaction with thiol groups. The backbone assignment of HMGB1 A-box was achieved with 3D NMR techinques. The ¹⁵N-labeled HMGB1 A-box was titrated with MTSL-labeled Hoxc9DBD respectively. Based on the chemical shift changes we can identify the interacting residues and further map out the binding sites on the protein structure. The NMR titration result showed that the binding interface of HMGB1 A-box is around loop-1 between helix-1 and helix-2. In addition, the additional contacts were found in N- and C-terminus. The N-terminal arm region of Hoxc9DBD is the major binding region and the loop between helix1 and helix2 is the minor binding region.

• Molecules and Cells
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• v.41 no.4
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• pp.362-372
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• 2018

High mobility group box 2 (HMGB2) is an abundant, chromatin-associated, non-histone protein involved in transcription, chromatin remodeling, and recombination. Recently, the HMGB2 gene was found to be significantly downregulated during senescence and shown to regulate the expression of senescent-associated secretory proteins. Here, we demonstrate that HMGB2 transcription is repressed by p21 during radiation-induced senescence through the ATM-p53-p21 DNA damage signaling cascade. The loss of p21 abolished the downregulation of HMGB2 caused by ionizing radiation, and the conditional induction of p21 was sufficient to repress the transcription of HMGB2. We also showed that the p21 protein binds to the HMGB2 promoter region, leading to sequestration of RNA polymerase and transcription factors E2F1, Sp1, and p300. In contrast, NF-Y, a CCAAT box-binding protein complex, is required for the expression of HMGB2, but NF-Y binding to the HMGB2 promoter was unaffected by either radiation or p21 induction. A proximity ligation assay results confirmed that the chromosome binding of E2F1 and Sp1 was inhibited by p21 induction. As HMGB2 have been shown to regulate premature senescence by IR, targeting the p21-mediated repression of HMGB2 could be a strategy to overcome the detrimental effects of radiation-induced senescence.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.10
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• pp.5789-5795
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• 2013

Background: The high mobility group box 1 (HMGB1) protein is a widespread nuclear protein present in most cell types. It typically locates in the nucleus and functions as a nuclear cofactor in transcription regulation. However, HMGB1 can also localize in the cytoplasm and be released into extracellular matrix, where it plays critical roles in carcinogenesis and inflammation. However, it remains elusive whether HMGB1 is relocated to cytoplasm in clear cell renal cell carcinoma (ccRCC). Methods: Nuclear and cytoplasmic proteins were extracted by different protocols from 20 ccRCC samples and corresponding adjacent renal tissues. Western blotting and immunohistochemistry were used to identify the expression of HMGB1 in ccRCC. To elucidate the potential mechanism of HMGB1 cytoplasmic translocation, HMGB1 proteins were enriched by immunoprecipitation and analyzed by mass spectrometry (MS). Results: The HMGB1 protein was overexpressed and partially localized in cytoplasm in ccRCC samples (12/20, 60%, p<0.05). Immunohistochemistry results indicated that ccRCC of high nuclear grade possess more HMGB1 relocation than those with low grade (p<0.05). Methylation of HMGB1 at lysine 112 in ccRCC was detected by MS. Bioinformatics analysis showed that post-translational modification might affect the binding ability to DNA and mediate its translocation. Conclusion: Relocation of HMGB1 to cytoplasm was confirmed in ccRCC. Methylation of HMGB1 at lysine 112 might the redistribution of this cofactor protein.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.12
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• pp.4865-4869
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• 2014

Objective: This study evaluated the expression level of high mobility group box-B1 (HMGB-1) and matrix metalloproteinase-9 (MMP-9) in non-small cell lung cancer (NSCLC) inmorder to reveal any relation with development and prognosis. Methods: NSCLC and normal tissues were selected from 30 patients at age of 30-73, and used for RT-PCR and Western blot analyses of HMGB-1. A total of 100 paraffin embedded NSCLC tissues were also isolated from patients through surgical resection, and used for detection of HMGB-1 by immunohistochemistry. In addition, 50 samples were also applied for MMP-9 detection, and 30 normal tissues were considered as controls. Correlation analysis of HMGB-1 and MMP-9 was carried out by Pearsons correlation coefficient. Results: The average expression level of HMGB-1 in NSCLC patients was significantly higher than in normal lung tissues. In addition, patients in III-IV period exhibit significantly higher positive rate of HMGB-1 when compared with I-II period cases. Furthermore, a positive correlation with HMGB-1 was found in the expression of MPP-9. Conclusion: HMGB-1 was highly expressed in NSCLC, which may become a prognostic and predictive marker for NSCLC. Besides, MPP-9 was positively correlated with HMGB-1.

• Korean Journal of Veterinary Research
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• v.51 no.3
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• pp.239-241
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• 2011

In this study, we investigated the kinetics of tumor necrosis factor (TNF)-${\alpha}$, interleukin (IL)-6 and high mobility group box 1 (HMGB1) concentrations in a 48-h model of canine endotoxemia by lipopolysaccharide (LPS) injection. Four healthy beagles were slowly administered 1 mg/kg of LPS diluted in normal saline, while two others were administered normal saline as controls. Blood collection was performed at 0 h (baseline), 1 h and 3 h (for TNF-${\alpha}$), 6 h, 12 h, 24 h and 48 h of the experiment, and cytokine levels were determined using the sandwich ELISA method. Early increments of TNF-${\alpha}$ and IL-6 were observed (< 3 h), but HMGB1 levels increased the most at 12 h of the experiment and gradually decreased until 48 h. During the whole experiment, IL-6 and HMGB1 were sustained over 12 h of LPS injection, whereas TNF-${\alpha}$ decreased within 6 h of LPS injection. Taken together, canine HMGB1 levels increase relatively late (< 12 h) and sustained longer than TNF-${\alpha}$ and IL-6 in response to endotoxin. This is the first study to evaluate canine HMGB1 cytokine from endotoxemia in dogs.

• BMB Reports
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• v.49 no.2
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• pp.99-104
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• 2016

The nuclear non-histone protein high mobility group box (HMGB) 1 is known to having an inhibitory effect on the repair of DNA damaged by the antitumor drug cisplatin in vitro. To investigate the role of HMGB1 in living cells, we studied the DNA repair of cisplatin damages in mouse fibroblast cell line, NIH-3T3. We evaluated the effect of the post-synthetic acetylation and C-terminal domain of the protein by overexpression of the parental and mutant GFP fused forms of HMGB1. The results revealed that HMGB1 had also an inhibitory effect on the repair of cisplatin damaged DNA in vivo. The silencing of HMGB1 in NIH-3T3 cells increased the cellular DNA repair potential. The increased levels of repair synthesis could be "rescued" and returned to less than normal levels if the knockdown cells were transfected with plasmids encoding HMGB1 and HMGB1 K2A. In this case, the truncated form of HMGB1 also exhibited a slight inhibitory effect.

• Journal of Life Science
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• v.29 no.11
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• pp.1179-1191
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• 2019

Cancer cells undergo the epithelial-mesenchymal transition (EMT) and show unique oncogenic metabolic phenotypes such as the glycolytic switch (Warburg effect) which are important for tumor development and progression. The EMT is a critical process for tumor invasion and metastasis. High-mobility group box 1 (HMGB1) is a chromatin-associated nuclear protein, but it acts as a damage-associated molecular pattern molecule when released from dying cells and immune cells. HMGB1 induces the EMT, as well as invasion and metastasis, thereby contributing to tumor progression. Here, we show that HMGB1 induced the EMT by activating Snail. In addition, the HMGB1/Snail cascade was found induce a glycolytic switch. HMGB1 also suppressed mitochondrial respiration and cytochrome c oxidase (COX) activity by a Snail-dependent reduction in the expression of the COX subunits COXVIIa and COXVIIc. HMGB1 also upregulated the expression of several key glycolytic enzymes, including hexokinase 2 (HK2), phosphofructokinase-2/fructose-2,6-bisphosphatase 2 (PFKFB2), and phosphoglycerate mutase 1 (PGAM1), in a Snail-dependent manner. However, HMGB1 was found to regulate some other glycolytic enzymes including lactate dehydrogenases A and B (LDHA and LDHB), glucose transporter 1 (GLUT1), and monocarboxylate transporters 1 and 4 (MCT1 and 4) in a Snail-independent manner. Transfection with short hairpin RNAs against HK2, PFKFB2, and PGAM1 prevented the HMGB1-induced EMT, indicating that glycolysis is associated with HMGB1-induced EMT. These findings demonstrate that HMGB1 signaling induces the EMT, glycolytic switch, and mitochondrial repression via Snail activation.

• Tuberculosis and Respiratory Diseases
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• v.80 no.2
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• pp.153-158
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• 2017

Background: Recently, increased levels of high-mobility group box 1 protein (HMGB1) have been identified in various inflammatory conditions and infections. However, no studies have evaluated the HMGB1 level in nontuberculous mycobacterial (NTM) lung disease, and compared it to mycobacterial lung disease. Methods: A total of 60 patients newly diagnosed with NTM lung disease, 44 culture-positive pulmonary tuberculosis (TB) patients, and 34 healthy controls, were included in this study. The serum HMGB1 concentrations were quantified using HMGB1 enzyme-linked immunosorbent assay kits. Results: Serum HMGB1 level in patients with pulmonary TB or NTM lung disease, was significantly lower than that of the healthy controls. In addition, the serum HMGB1 level in TB patients was significantly lower than patients with NTM lung disease. However, the levels in NTM patient subgroups did not differ according to the causative species, disease progression, and disease phenotype. Conclusion: Although low levels of serum HMGB1 has the potential to be a marker of mycobacterial lung disease, these levels were unable to differentiate disease progression and disease phenotype in NTM lung diseases.

• Bulletin of the Korean Chemical Society
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• v.35 no.10
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• pp.2955-2962
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• 2014

Asian ginseng is used as a treatment for cardiovascular diseases, ischemia, and cancers. High mobility group box 1 (HMGB1) protein acts as a late mediator of severe vascular inflammatory conditions. However, the effect of ginsenosides from Asian ginseng on HMGB1-induced inflammatory responses has not been studied. We addressed this question by monitoring the effects of ginsenoside treatment on lipopolysaccharide (LPS) and cecal ligation and puncture (CLP)-mediated release of HMGB1, and HMGB1-mediated regulation of proinflammatory responses. Ginsenoside treatment suppressed LPS-mediated release of HMGB1 and HMGB1-mediated cytoskeletal rearrangements. Ginsenosides also inhibited HMGB1-mediated inflammatory responses. In addition, ginsenosides inhibited the production of tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$) and activation of protein kinase B (Akt), nuclear factor-${\kappa}B$ (NF-${\kappa}B$), and extracellular-regulated kinases (ERK) 1/2 by HMGB1. Ginsenosides also decreased CLP-induced release of HMGB1, production of interleukin (IL) $1{\beta}/6$, and mortality. These results suggested that ginsenosides may be potential therapeutic agents for treatment of vascular inflammatory diseases through inhibition of the HMGB1 signaling pathway.

• IMMUNE NETWORK
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• v.12 no.4
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• pp.148-154
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• 2012

Previously, we have reported that high mobility group box 1 (HMGB1), a proinflammatory mediator in sepsis, is released via the IFN-${\beta}$-mediated JAK/STAT pathway. However, detailed mechanisms are still unclear. In this study, we dissected upstream signaling pathways of HMGB1 release using various molecular biology methods. Here, we found that calcium/calmodulin-dependent protein kinase (CaM kinase, CaMK) is involved in HMGB1 release by regulating IFN-${\beta}$ production. CaMK inhibitor, STO609, treatment inhibits LPS-induced IFN-${\beta}$ production, which is correlated with the phosphorylation of interferon regulatory factor 3 (IRF3). Additionally, we show that CaMK-I plays a major role in IFN-${\beta}$ production although other CaMK members also seem to contribute to this event. Furthermore, the CaMK inhibitor treatment reduced IFN-${\beta}$ production in a murine endotoxemia. Our results suggest CaMKs contribute to HMGB1 release by enhancing IFN-${\beta}$ production in sepsis.