Although 53BP1 has been established well as a mediator in DNA damage response, its function in mitosis is not clearly understood. We found that 53BP1 is a mitotic-binding partner of the kinases Plk1 and AuroraA, and that the binding with Plk1 increases the stability of 53BP1 by accelerating its interaction with the deubiquitinase USP7. Depletion of 53BP1 induces mitotic defects such as chromosomal missegregation, misorientation of spindle poles and the generation of extra centrosomes, which is similar phenotype to USP7-knockdown cells. In addition, 53BP1 depletion reduces the levels of p53 and centromere protein F (CENPF), interacting proteins of 53BP1. These phenotypes induced by 53BP1 depletion were rescued by expression of wild-type or phosphomimic mutant 53BP1 but not by expression of a dephosphomimic mutant. We propose that phosphorylation of 53BP1 at S380 accelerates complex formation with USP7 and CENPF to regulate their stability, thus having a crucial role in proper centrosome positioning, chromosomal alignment, and centrosome number.

Molecular inflammation is a pivotal process in various degenerative immune diseases, including asthma and atopic dermatitis. In this study, we examined the effects of Helianthus annuus seed (HAS) aqueous extract on an in vivo anti-asthmatic model. Ovalbumin-induced mice were orally administered the aqueous extract of Helianthus annuus seeds, and their lungs were assessed by hematoxylin and eosin staining. Moreover, the expression levels of IL-4/IL-13 cytokines and IgE were determined. HAS extract induced a decrease in CD4 + cell number, IL-4/IL-13 expression, and IgE secretion levels in the lungs. Our findings collectively suggest that the HAS extract has considerable potential in reducing the asthma-like symptoms induced by a mouse ovalbumin challenge model. However, further isolation and purification of the extract is required to determine the specific factor(s) responsible for its anti-asthmatic activity.Abbreviations: HAS, Helianthus annuus seeds; DPPH, 1,1diphenyl-2-picrylhydrazyl; Fe(III)-TPTZ, ferric tripyridyltriazine; EDTA, ethylenediamine tetraacetic acid; FRAP, ferric ion-reducing antioxidant power; MMP, matrix metalloproteinase; IL, interleukin

Polo-like kinase 1 (Plk1), a conserved Ser/Thr mitotic kinase, has been identified as a promising target for anticancer drug development because its overexpression is correlated with malignancy. Here, we found that genistein, an isoflavone, inhibits Plk1 kinase activity directly. Previously the mitotic disturbance phenomenon induced by treatment with genistein was not fully explained by its inhibitory effect on EGFR. In kinase profiling assays, it showed selectivity relative to a panel of kinases, including EGFR. Treatment with genistein induced cell death in a concentration-dependent manner in cancer cells from diverse tissue origins, but not in non-transformed cells such as hTERT-RPE or MCF10A cells. We also observed that genistein tended to be more selective against cancer cells with mutations in the TP53 gene. TP53-depeleted LNCaP and NCI-H460 cells using shRNA targeting human TP53 were more sensitive to cell death by treatment of genistein. Furthermore, genistein induced mitotic arrest by inhibiting Plk1 activity and, consequently, led to mitotic catastrophe and apoptosis. These data suggest that genistein may be a promising anticancer drug candidate due to its inhibitory activity against Plk1 as well as EGFR and effectiveness toward cancer cells, especially those with p53-mutation. J. Cell. Physiol. 232: 2818-2828, 2017. © 2016 Wiley Periodicals, Inc.

Backgrounds: Despite the clinical success of taxanes, they still have limitations, such as chemoresistance. To overcome the limitations of paclitaxel, genetic alterations and targeting effects of altered genes were observed in paclitaxel-resistant cancer. Because paclitaxel-resistant cancer shows high levels of Plk1, a promising target in chemotherapy, the effectiveness of Plk1 inhibitors in paclitaxel-resistant cancer cells has been investigated. Methods: Paclitaxel-resistant cancer cells were developed by exposure of stepwise escalating levels of paclitaxel. Genetic alterations were detected by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and immunoblotting. Using a cell viability assay, combined targeting effects for Plk1 and androgen receptor (AR) were determined. Clinical data were analyzed to understand the relationship between Plk1 and AR in prostate cancer patients. Results: Treatment with Plk1 inhibitors markedly reduced the expression of MDR1, MRP1, and Plk1 in the paclitaxel-resistant cancer. Among Plk1 inhibitors, genistein, recently found as a direct Plk1 inhibitor, tended to be more effective in the paclitaxel-resistant prostate cancer than the parental cancer cells, which was related to the suppression of the AR, as well as inhibition of Plk1 activity. A combination of Plk1 inhibitors and AR antagonist bicalutamide exhibited a synergistic effect in LNCaP TXR , as well as LNCaP cells, by inhibiting Plk1 and AR. Analysis of clinical data provides evidence for the relevance between Plk1 and AR in prostate cancer patients, showing that Plk1 and AR are strong predictors of poor survival rates. Conclusions: We suggest that cotargeting Plk1 and AR would be effective in advanced chemoresistant prostate cancer cells to overcome the limitations associated with paclitaxel.

Abstract. Gastrodia elata Blume (GEB) is an important medicinal plant in Korea. In order to confirm the anti-tumor activities of GEB extracts, we carried out various in vitro antitumor assays, including a wound assay and an invasion assay using an ethyl ether extract of GEB. The results showed that the GEB extract exhibits potent anti-tumor activity in vitro in a dose-dependent manner. The expression of CD44, cdc42, Timp-2 or RhoA mRNA did not change by GEB treatment, compared to that of the control. GTP-Ras, an active form of a G-coupled protein family, however, is associated with the anti-tumor activity of GEB extracts. We examined various molecular markers related to metastasis by reverse transcriptase-polymerase chain reaction with the extract of GEB-treated B16 cells. There was an increase in GTP-Ras expression by the Gastrodia elata Blume extract. Together, these results suggest that the Gastrodia elata Blume extract could have potential in alleviating tumorigenesis, by a GTPRas-dependent pathway; although the precise molecular mechanisms are still being examined. IntroductionGastrodia elata Blume (GEB) is a traditional herb that has been used in East Asian for centuries. It has been used as an anticonvulsant, analgesic and sedative to combat vertigo, hypertension, general paralysis and tetanus. Vanillyl alcohol and gastrodin, derived from GEB are known to have anticonvulsive actions (1). Recently, it was reported that compounds found in GEB inhibited glutamate-induced apoptosis in neuronal cells (2). In addition, after pentylenetetrazole-induced seizure activity, the ether fraction of GEB has been shown to attenuate a decrease in γ-aminobutyric acid (GABA) and an increase in glutamate content, as well as having anticonvulsant effects (3).GEB is also used as sub-material for food or food-related industry. In 2001, the Korea Food and Drug Administration approved that GEB extracts as food ingredients; however, an in vivo toxicological study is still needed to determine its safety in food. We have already reported that GEB powersupplemented (0.5-1.0%) dough had a membrane-like structure that was more developed than that of the control, resulting in increased bread volumes (4). These results suggest that bread baked with 0.5-1.0% GEB exhibited an increase in loaf volume due to the more complete development of a gluten matrix.In our study, we determined that GEB protected cell damage by ß-amyloid in neuroblastoma cells (5). The ethyl ether fraction of GEB has potent activity toward ß-amyloidinduced cell damage via reducing caspase-3 activity (6,7; data not shown). In the course of a mechanistic study of the methanolic extract of GEB, we found that it has potent antitumor activity in vitro. The major finding of this report is that GEB exhibits anti-metastatic activities which were confirmed by wound and invasion assay in B16 melanoma cells. We further demonstrated that GEB increased anti-tumor activity via a GTP-Ras-dependent pathway. Materials and methodsCell culture. A murine melanoma cell line B16-F1 (B16; Ca...