FANCD2 and RAD51 recombinase directly inhibit DNA2 nuclease at stalled replication forks and FANCD2 acts as a novel RAD51 mediator in strand exchange to promote genome stability.

FANCD2 protein, a key coordinator and effector of the interstrand crosslink repair pathway, is also required to prevent excessive nascent strand degradation at hydroxyurea-induced stalled forks. The RAD51 recombinase has also been implicated in regulation of resection at stalled replication forks. The mechanistic contributions of these proteins to fork protection are not well understood. Here, we used purified FANCD2 and RAD51 to study how each protein regulates DNA resection at stalled forks. We characterized three mechanisms of FANCD2-mediated fork protection: (1) The N-terminal domain of FANCD2 inhibits the essential DNA2 nuclease activity by directly binding to DNA2 accounting for over-resection in FANCD2 defective cells. (2) Independent of dimerization with FANCI, FANCD2 itself stabilizes RAD51 filaments to inhibit multiple nucleases, including DNA2, MRE11 and EXO1. (3) Unexpectedly, we uncovered a new FANCD2 function: by stabilizing RAD51 filaments, FANCD2 acts to stimulate the strand exchange activity of RAD51. Our work biochemically explains non-canonical mechanisms by which FANCD2 and RAD51 protect stalled forks. We propose a model in which the strand exchange activity of FANCD2 provides a simple molecular explanation for genetic interactions between FANCD2 and BRCA2 in the FA/BRCA fork protection pathway.

Heat Shock Response and Heat Shock Proteins: Current Understanding and Future Opportunities in Human Diseases.

The heat shock response is an evolutionarily conserved mechanism that protects cells or organisms from the harmful effects of various stressors such as heat, chemicals toxins, UV radiation, and oxidizing agents. The heat shock response triggers the expression of a specific set of genes and proteins known as heat shock genes/proteins or molecular chaperones, including HSP100, HSP90, HSP70, HSP60, and small HSPs. Heat shock proteins (HSPs) play a crucial role in thermotolerance and aiding in protecting cells from harmful insults of stressors. HSPs are involved in essential cellular functions such as protein folding, eliminating misfolded proteins, apoptosis, and modulating cell signaling. The stress response to various environmental insults has been extensively studied in organisms from prokaryotes to higher organisms. The responses of organisms to various environmental stressors rely on the intensity and threshold of the stress stimuli, which vary among organisms and cellular contexts. Studies on heat shock proteins have primarily focused on HSP70, HSP90, HSP60, small HSPs, and ubiquitin, along with their applications in human biology. The current review highlighted a comprehensive mechanism of heat shock response and explores the function of heat shock proteins in stress management, as well as their potential as therapeutic agents and diagnostic markers for various diseases.

Molecular Mechanisms of Neuroprotection by Ketone Bodies and Ketogenic Diet in Cerebral Ischemia and Neurodegenerative Diseases.

Ketone bodies (KBs), such as acetoacetate and ß-hydroxybutyrate, serve as crucial alternative energy sources during glucose deficiency. KBs, generated through ketogenesis in the liver, are metabolized into acetyl-CoA in extrahepatic tissues, entering the tricarboxylic acid cycle and electron transport chain for ATP production. Reduced glucose metabolism and mitochondrial dysfunction correlate with increased neuronal death and brain damage during cerebral ischemia and neurodegeneration. Both KBs and the ketogenic diet (KD) demonstrate neuroprotective effects by orchestrating various cellular processes through metabolic and signaling functions. They enhance mitochondrial function, mitigate oxidative stress and apoptosis, and regulate epigenetic and post-translational modifications of histones and non-histone proteins. Additionally, KBs and KD contribute to reducing neuroinflammation and modulating autophagy, neurotransmission systems, and gut microbiome. This review aims to explore the current understanding of the molecular mechanisms underpinning the neuroprotective effects of KBs and KD against brain damage in cerebral ischemia and neurodegenerative diseases, including Alzheimer's disease and Parkinson's disease.

Beta-naphthoflavone and doxorubicin synergistically enhance apoptosis in human lung cancer cells by inducing doxorubicin accumulation, mitochondrial ROS generation, and JNK pathway signaling.

Doxorubicin is one of the most effective chemotherapeutic agents available for treating various cancers, including lung cancer-the leading cause of cancer death in both men and women. However, its clinical application has been impeded by severe adverse effects, notably cardiotoxicity. Development of cellular resistance to doxorubicin is another major obstacle that must be overcome for broader application of the drug. In the present study, we examined the therapeutic potential of beta-naphthoflavone (BNF), a synthetic derivative of a naturally occurring flavonoid, in combination with doxorubicin for the treatment of lung cancer. Among our novel observations were that BNF enhances the efficacy of doxorubicin by inducing doxorubicin accumulation, mitochondrial ROS generation, and JNK pathway signaling in lung cancer cells. These combined effects were also evident in many other cancer cell types. BNF further exhibited synergistic induction of apoptosis in lung cancer cells when combined with several other cancer drugs, including irinotecan, cisplatin, and 5-fluorouracil. Our results suggest that BNF can be developed as a promising adjuvant agent for enhancing the efficacy of doxorubicin.

Honokiol inhibits hepatoma carcinoma cell migration through downregulated Cyclophilin B expression.

Hepatocellular carcinoma (HCC) is the fifth common types of cancer with poor prognosis in the world. Honokiol (HNK), a natural biphenyl compound derived from the magnolia plant, has been reported to exert anticancer effects, but its mechanism has not been elucidated exactly. In the present study, HNK treatment significantly suppressed the migration ability of HepG2 and Hep3B human hepatocellular carcinoma. The treatment reduced the expression levels of the genes associated with cell migration, such as S100A4, MMP-2, MMP-9 and Vimentin. Interestingly, treatment with HNK significantly reduced the expression level of Cyclophilin B (CypB) which stimulates cancer cell migration. However, overexpressed CypB abolished HNK-mediated suppression of cell migration, and reversed the apoptotic effects of HNK. Altogether, we concluded that the suppression of migration activities by HNK was through down-regulated CypB in HCC. These finding suggest that HNK may be a promising candidate for HCC treatment via regulation of CypB.

2-O-Methylhonokiol Suppresses HCV Replication via TRAF6-Mediated NF-kB Activation.

Hepatitis C virus (HCV) is associated with various liver diseases. Chronic HCV infection is characterized by an abnormal host immune response. Therefore, it is speculated that to suppress HCV, a well-regulated host immune response is necessary. 2-O-methylhonokiol was identified by the screening of anti-HCV compounds using Renilla luciferase assay in Huh 7.5/Con 1 genotype 1b replicon cells. Here, we investigated the mechanism by which 2-O-methylhonokiol treatment inhibits HCV replication using real-time PCR. Our data shows that treatment with 2-O-methylhonokiol activated innate immune responses via nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB) pathway. Additionally, the immunoprecipitation result shows that treatment with 2-O-methylhonokiol augmented tumor necrosis factor receptor (TNFR)-associated factor 6 (TRAF6) by preventing p62 from binding to TRAF6, resulting in reduced autophagy caused by HCV. Finally, we reproduced our data with the conditioned media from 2-O-methylhonokiol-treated cells. These findings strongly suggest that 2-O-methylhonokiol enhances the host immune response and suppresses HCV replication via TRAF6-mediated NF-kB activation.

Correlation between Oxidative Stress and Transforming Growth Factor-Beta in Cancers.

The downregulation of reactive oxygen species (ROS) facilitates precancerous tumor development, even though increasing the level of ROS can promote metastasis. The transforming growth factor-beta (TGF-ß) signaling pathway plays an anti-tumorigenic role in the initial stages of cancer development but a pro-tumorigenic role in later stages that fosters cancer metastasis. TGF-ß can regulate the production of ROS unambiguously or downregulate antioxidant systems. ROS can influence TGF-ß signaling by enhancing its expression and activation. Thus, TGF-ß signaling and ROS might significantly coordinate cellular processes that cancer cells employ to expedite their malignancy. In cancer cells, interplay between oxidative stress and TGF-ß is critical for tumorigenesis and cancer progression. Thus, both TGF-ß and ROS can develop a robust relationship in cancer cells to augment their malignancy. This review focuses on the appropriate interpretation of this crosstalk between TGF-ß and oxidative stress in cancer, exposing new potential approaches in cancer biology.

Anti-obesity Effect of Fermented Persimmon Extracts via Activation of AMP-Activated Protein Kinase.

There is significant cultivation of persimmon (Diospyros kaki) in East Asia, a plant whose fruit has abundant nutrients, including vitamins, polyphenols, and dietary fiber. Persimmon dietary supplements can benefit health by amelioration of diabetes, cardiovascular disease, and obesity. There are also persimmon-based beverages produced via fermentation, such as wines and vinegars, and increasing consumption of these products in East Asia. Although there is great interest in functional foods, the health effects of fermented persimmon extract (FPE) are completely unknown. We examined the effects of FPE on the metabolic parameters of mice fed a high-fat diet (HFD). Our results indicated that FPE supplementation led to an approx. 15% reduction of body weight, reduced abdominal and liver fat, and reduced serum levels of triglycerides, total cholesterol, and glucose. FPE also blocked the differentiation of murine 3T3-L1 pre-adipocyte cells into mature adipocytes. We suggest that gallic acid is a major bioactive component of FPE, and that AMP-activated protein kinase mediates the beneficial effects of FPE and gallic acid.

Jeju Magma-Seawater Inhibits α-MSH-Induced Melanogenesis via CaMKKß-AMPK Signaling Pathways in B16F10 Melanoma Cells.

Melanin protects skin from ultraviolet radiation, toxic drugs, and chemicals. Its synthesis is sophisticatedly regulated by multiple mechanisms, including transcriptional and enzymatic controls. However, uncontrolled excessive production of melanin can cause serious dermatological disorders, such as freckles, melasma, solar lentigo, and cancer. Moreover, melanogenesis disorders are also linked to neurodegenerative diseases. Therefore, there is a huge demand for safer and more potent inhibitors of melanogenesis. In the present study, we report novel inhibitory effects of Jeju magma-seawater (JMS) on melanogenesis induced by α-melanocyte stimulating hormone (α-MSH) in B16F10 melanoma cells. JMS is the abundant underground seawater found in Jeju Island, a volcanic island of Korea. Research into the physiological effects of JMS is rapidly increasing due to its high contents of various minerals that are essential to human health. However, little is known about the effects of JMS on melanogenesis. Here, we demonstrate that JMS safely and effectively inhibits α-MSH-induced melanogenesis via the CaMKKß (calcium/calmodulin-dependent protein kinase ß)-AMPK (5' adenosine monophosphate-activated protein kinase) signaling pathway. We further demonstrate that AMPK inhibits the signaling pathways of protein kinase A and MAPKs (mitogen-activated protein kinase), which are critical for melanogenesis-related gene expression. Our results highlight the potential of JMS as a novel therapeutic agent for ameliorating skin pigmentation-related disorders.

A Small Molecule, 4-Phenylbutyric Acid, Suppresses HCV Replication via Epigenetically Induced Hepatic Hepcidin.

Hepatic hepcidin is a well-known major iron regulator and has been reported to be closely related to hepatitis C virus (HCV) replication. However, pharmacological targeting of the hepcidin in HCV replication has not been reported. A short-chain fatty acid, 4-Phenyl butyrate (4-PBA), is an acid chemical chaperone that acts as a histone deacetylase inhibitor (HDACi) to promote chromosomal histone acetylation. Here, we investigated the therapeutic effect of 4-PBA on hepcidin expression and HCV replication. We used HCV genotype 1b Huh 7.5-Con1 replicon cells and engraftment of NOD/SCID mice as in vitro and in vivo models to test the effect of 4-PBA. It was found that 4-PBA inhibited HCV replication in Huh7.5-Con1 replicon cells in a concentration- and time-dependent manner through the induction of hepcidin expression by epigenetic modification and subsequent upregulation of interferon-α signaling. HCV formed a membranous web composed of double-membrane vesicles and was utilized for RNA replication. Moreover, 4-PBA also disrupted the integrity of the membranous web and interfered with the molecular interactions critical for the assembly of the HCV replication complex. These findings suggest that 4-PBA is a key epigenetic inducer of anti-HCV hepatic hepcidin and might at least in part play a role in targeting host factors related to HCV infection as an attractive complement to current HCV therapies.

Correction to: Licochalcone A induces apoptosis through endoplasmic reticulum stress via a phospholipase Cγ1-, Ca2+-, and reactive oxygen species-dependent pathway in HepG2 human hepatocellular carcinoma cells.

The original version of this article contained mistakes in figures. The western blot data for pro-caspase-3 and cleaved caspase-3 (Fig. 1d), ß-actin (Fig. 1d), PLCγ1 (Fig. 5d), and eIF2α (Fig. 7d) are incorrect. The corrected Figs. 1d, 5d, and 7d are shown below. The corrections do not influence either the validity of the published data or the conclusion described in the article.

Correction to: Apicidin induces endoplasmic reticulum stress- and mitochondrial dysfunction-associated apoptosis via phospholipase Cγ1- and Ca2+-dependent pathway in mouse Neuro-2a neuroblastoma cells.

The original version of this article contained a mistake in the figure. The Ca2 + confocal image for the 2-APB/Apicidin-120 min in Fig. 5d is incorrect. The correction does not influence either the validity of the published data or the conclusion described in the article. The corrected Fig. 5d is given below.

Cyclophilin B induces chemoresistance by degrading wild-type p53 via interaction with MDM2 in colorectal cancer.

Colorectal cancer (CRC) is one of the leading causes of cancer-related deaths worldwide. Chemoresistance is a major problem for effective therapy in CRC. Here, we investigated the mechanism by which peptidylprolyl isomerase B (PPIB; cyclophilin B, CypB) regulates chemoresistance in CRC. We found that CypB is a novel wild-type p53 (p53WT)-inducible gene but a negative regulator of p53WT in response to oxaliplatin treatment. Overexpression of CypB shortens the half-life of p53WT and inhibits oxaliplatin-induced apoptosis in CRC cells, whereas knockdown of CypB lengthens the half-life of p53WT and stimulates p53WT-dependent apoptosis. CypB interacts directly with MDM2, and enhances MDM2-dependent p53WT ubiquitination and degradation. Furthermore, we firmly validated, using bioinformatics analyses, that overexpression of CypB is associated with poor prognosis in CRC progression and chemoresistance. Hence, we suggest a novel mechanism of chemoresistance caused by overexpressed CypB, which may help to develop new anti-cancer drugs. We also propose that CypB may be utilized as a predictive biomarker in CRC patients. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Recombinant human Cyclophilin A stimulates hair follicle cells via Wnt/ß-catenin signaling pathway.

OBJECTIVES: To explore potential effects of recombinant peptidyl-prolyl isomerase human Cyclophilin A (CypA) in the growth of mouse hair and human hair follicle dermal papilla cells (HDP). RESULTS: The hair growth of 8-weeks C57BL/6 mouse was significantly stimulated by recombinant human CypA protein. CypA also promoted Human Dermal Papilla (HDP) cell growth and induced ß-catenin protein through GSK3ß inhibition. CONCLUSIONS: The molecular chaperone CypA stimulates hair follicle and HDP cells through Wnt/ß-catenin signaling pathway, suggesting that it might be a key molecule for hair loss disorder therapy.

Correction to: Release of overexpressed CypB activates ERK signaling through CD147 binding for hepatoma cell resistance to oxidative stress.

The original version of this article contained a mistake. The bands for HA Tag and t-ERK in Figs. 2d, 2h, 3d are incorrect. The author informs that these errors had no influence in the scientific content of the paper. The corrected figures (Figs. 2 and 3) are given below.

Transient activation of AMP-activated protein kinase at G1/S phase transition is required for control of S phase in NIH3T3 cells.

AMP-activated protein kinase (AMPK) functions as a cellular energy sensor by monitoring the cellular AMP:ATP ratio and plays a central role in cellular and whole-body energy homeostasis. Recent studies have suggested that AMPK also contribute to cell cycle regulation, but its role in this field remains almost elusive. In the present study, we report that AMPKα1 was transiently activated during G1/S transition phase in NIH3T3 cells in the absence of any metabolic stress. Inhibition of AMPK activity at G1/S transition phase completely blocked cells from entering S phase; in contrast, persistent activation of AMPK at G1/S transition phase allowed cells to normally enter S phase, but these cells failed to proceed to G2/M phase, stacking at S phase. We further demonstrated that activation of AMPK at G1/S transition phase depends on Ca2+ transients and CaMKKß activity, but not on energy status. Collectively, these data indicate that temporal regulation of AMPK is required for proper control of S phase in NIH3T3 cells.

Xanthohumol ameliorates 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced cellular toxicity in cultured MC3T3-E1 osteoblastic cells.

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is an environmental contaminant. Xanthohumol is a prenylated flavonoid found in hops (Humulus lupulus) and beer. The aim of the current study was to explore the role of xanthohumol in modulating the toxicity of TCDD in MC3T3-E1 osteoblastic cells. In cells treated with TCDD alone, intracellular Ca2+ concentrations, mitochondrial membrane potential disruption, reactive oxygen species production, cardiolipin peroxidation, nitric oxide release and cytochrome P450 1A1 expression were significantly increased. TCDD treatment increased the mRNA levels of extracellular signal-regulated kinase 1 and nuclear factor kappa B, and significantly decreased the level of protein kinase B (AKT) in MC3T3-E1 osteoblastic cells. However, the presence of xanthohumol alleviated the pathological effects of TCDD. In addition, xanthohumol treatment significantly increased the expression of genes associated with osteoblast differentiation (alkaline phosphatase, osteocalcin, osteoprotegerin and osterix). We conclude that xanthohumol has a beneficial influence and may antagonize TCDD toxicity in osteoblastic cells.

Fluoxetine induces apoptosis through endoplasmic reticulum stress via mitogen-activated protein kinase activation and histone hyperacetylation in SK-N-BE(2)-M17 human neuroblastoma cells.

Fluoxetine (FLX) is an antidepressant drug that belongs to the class of selective serotonin reuptake inhibitors. FLX is known to induce apoptosis in multiple types of cancer cells. In this study, the molecular mechanisms underlying the anti-cancer effects of FLX were investigated in SK-N-BE(2)-M17 human neuroblastoma cells. FLX induced apoptotic cell death, activation of caspase-4, -9, and -3, and expression of endoplasmic reticulum (ER) stress-associated proteins, including C/EBP homologous protein (CHOP). Inhibition of ER stress by treatment with the ER stress inhibitors, salubrinal and 4-phenylbutyric acid or CHOP siRNA transfection reduced FLX-induced cell death. FLX induced phosphorylation of mitogen-activated protein kinases (MAPKs) family, p38, JNK, and ERK, and an upstream kinase apoptosis signal kinase 1 (ASK1). Inhibition of MAPKs and ASK1 reduced FLX-induced cell death and CHOP expression. We then showed that FLX reduced mitochondrial membrane potential (MMP) and ER stress inhibitors as well as MAPK inhibitors ameliorated FLX-induced loss of MMP. Interestingly, FLX induced hyperacetylation of histone H3 and H4, upregulation of p300 histone acetyltransferase (HAT), and downregulation of histone deacetylases (HDACs). Treatment with a HAT inhibitor anacardic acid or p300 HAT siRNA transfection blocked FLX-induced apoptosis in SK-N-BE(2)-M17 cells. However, FLX did not induce histone acetylation and anacardic acid had no protective effect on FLX-induced cell death and CHOP expression in MYCN non-amplified SH-SY5Y human neuroblastoma and MYCN knockdowned SK-N-BE(2)-M17 cells. These findings suggest that FLX induces apoptosis in neuroblastoma through ER stress and mitochondrial dysfunction via the ASK1 and MAPK pathways and through histone hyperacetylation in a MYCN-dependent manner.