EZH2-mediated upregulation of ROS1 oncogene promotes oral cancer metastasis.

Current anti-epidermal growth factor receptor (EGFR) therapy for oral cancer does not provide satisfactory efficacy due to drug resistance or reduced EGFR level. As an alternative candidate target for therapy, here we identified an oncogene, ROS1, as an important driver for oral squamous cell carcinoma (OSCC) metastasis. Among tumors from 188 oral cancer patients, upregulated ROS1 expression strongly correlated with metastasis to lung and lymph nodes. Mechanistic studies uncover that the activated ROS1 results from highly expressed ROS1 gene instead of gene rearrangement, a phenomenon distinct from other cancers. Our data further reveal a novel mechanism that reduced histone methyltransferase EZH2 leads to a lower trimethylation of histone H3 lysine 27 suppressive modification, relaxes chromatin, and promotes the accessibility of the transcription factor STAT1 to the enhancer and the intron regions of ROS1 target genes, CXCL1 and GLI1, for upregulating their expressions. Down-regulation of ROS1 in highly invasive OSCC cells, nevertheless, reduces cell proliferation and inhibits metastasis to lung in the tail-vein injection and the oral cavity xenograft models. Our findings highlight ROS1 as a candidate biomarker and therapeutic target for OSCC. Finally, we demonstrate that co-targeting of ROS1 and EGFR could potentially offer an effective oral cancer therapy.

miRNA-34c-5p inhibits amphiregulin-induced ovarian cancer stemness and drug resistance via downregulation of the AREG-EGFR-ERK pathway.

Epithelial ovarian cancer is the most lethal gynecological cancer mainly due to late diagnosis, easy spreading and rapid development of chemoresistance. Cancer stem cells are considered to be one of the main mechanisms for chemoresistance, as well as metastasis and recurrent disease. To explore the stemness characteristics of ovarian cancer stem cells, we successfully enriched ovarian cancer stem-like cells from an established ovarian cancer cell line (SKOV-I6) and a fresh ovarian tumor-derived cell line (OVS1). These ovarian cancer stem-like cells possess important cancer stemness characteristics including sphere-forming and self-renewing abilities, expressing important ovarian cancer stem cell and epithelial-mesenchymal transition markers, as well as increased drug resistance and potent tumorigenicity. Microarray analysis of OVS1-derived sphere cells revealed increased expression of amphiregulin (AREG) and decreased expression of its conserved regulatory microRNA, miR-34c-5p, when compared with the OVS1 parental cells. Overexpression of AREG and decreased miR-34c-5p expression in SKOV-I6 and OVS1 sphere cells were confirmed by quantitative real-time PCR analysis. Luciferase reporter assay and mutant analysis confirmed that AREG is a direct target of miR-34c-5p. Furthermore, AREG-mediated increase of sphere formation, drug resistance toward docetaxel and carboplatin, as well as tumorigenicity of SKOV-I6 and OVS1 cells could be abrogated by miR-34c-5p. We further demonstrated that miR-34c-5p inhibited ovarian cancer stemness through downregulation of the AREG-EGFR-ERK pathway. Overexpression of AREG was found to be correlated with advanced ovarian cancer stages and poor prognosis. Taken together, our data suggest that AREG promotes ovarian cancer stemness and drug resistance via the AREG-EGFR-ERK pathway and this is inhibited by miR-34c-5p. Targeting AREG, miR-34c-5p could be a potential strategy for anti-cancer-stem cell therapy in ovarian cancer.

MicroRNA-149 targets GIT1 to suppress integrin signaling and breast cancer metastasis.

Metastasis is the predominant cause of death in breast cancer patients. Several lines of evidence have shown that microRNAs (miRs) can have an important role in cancer metastasis. Using isogenic pairs of low and high metastatic lines derived from a human breast cancer line, we have identified miR-149 to be a suppressor of breast cancer cell invasion and metastasis. We also identified GIT1 (G-protein-coupled receptor kinase-interacting protein 1) as a direct target of miR-149. Knockdown of GIT1 reduced migration/invasion and metastasis of highly invasive cells. Re-expression of GIT1 significantly rescued miR-149-mediated inhibition of cell migration/invasion and metastasis. Expression of miR-149 impaired fibronectin-induced focal adhesion formation and reduced phosphorylation of focal adhesion kinase and paxillin, which could be restored by re-expression of GIT1. Inhibition of GIT1 led to enhanced protein degradation of paxillin and α5ß1 integrin via proteasome and lysosome pathways, respectively. Moreover, we found that GIT1 depletion in metastatic breast cancer cells greatly reduced α5ß1-integrin-mediated cell adhesion to fibronectin and collagen. Low level of miR-149 and high level of GIT1 was significantly associated with advanced stages of breast cancer, as well as with lymph node metastasis. We conclude that miR-149 suppresses breast cancer cell migration/invasion and metastasis by targeting GIT1, suggesting potential applications of the miR-149-GIT1 pathway in clinical diagnosis and therapeutics.

Non-resonant and resonant x-ray scattering studies on multiferroic TbMn2O5.

Comprehensive x-ray scattering studies, including resonant scattering at Mn L, Tb L, and M edges, were performed on single crystals of TbMn2O5 for crystallographic data to elucidate the nature of its commensurate and incommensurate phases. The scattering results provide direct evidence of symmetry lowering to the ferroelectric phase driven by magnetically induced lattice modulations and show the presence of multiple magnetic orders. The competing orders under spin-frustrated geometry are believed to cause discommensuration and result in the commensurate-to-incommensurate phase transition around 24 K. It is proposed that the low temperature incommensurate phase consists of commensurate domains separated by antiphase domain walls which change both signs of spontaneous polarizations and x-ray scattering amplitudes for forbidden reflections.

Alterations in Ca2+ signaling, and c-fos and nur77 expression are associated with sodium butyrate-induced differentiation of C6 glioma cell.

Sodium butyrate is well known in stimulating growth and differentiation of cancer cells. In the present study, butyrate treatment caused decreases in thymidine incorporation in the early passages (45-60) of C6 glioma cells. In addition, butyrate also caused decreases in inositol incorporation and transient ATP-stimulated Ca2+ mobilization suggesting that butyrate altered general mechanisms of Ca2+ signaling in these cells. To gain direct insight into the crosstalk between sodium butyrate and Ca2+ signaling in transcriptional regulation, we investigated the induction of the Ca2+-sensitive immediate early genes (IEGs), c-fos, nur77 and c-myc. Sodium butyrate per se enhanced the expression of c-fos mRNA, and the enhanced levels were maintained for 24 h, but over the same time period, the initially increased levels of nur77 expression tailed off, while c-myc expression was slightly reduced. Increasing intracellular Ca2+ concentration ([Ca2+]i) by thapsgargin and A23187 induced the expression of both c-fos and nur77 mRNA expression, and synergistic effects were observed when cells were incubated with sodium butyrate plus thapsgargin and A23187. However, removal of both extracellular Ca2+ by EGTA, or intracellular free Ca2+ with BAPTA did not affect the sodium butyrate-induced c-fos and nur77 mRNA. These results suggest that although sodium butyrate altered Ca2+ signaling which is an important regulatory mechanism for c-fos and nur77 expression, nevertheless the sodium butyrate-induced c-fos and nur77 expression may be not in fact mediated through Ca2+ signaling.

Altered phospholipid metabolism in sodium butyrate-induced differentiation of C6 glioma cells.

We examined the changes in phospholipid metabolisms in sodium butyrate-treated C6 glioma cells. Treatment of 2.5 mM sodium butyrate for 24 h induced an increase in the activity of glutamine synthetase, suggesting that these cells were under differentiation. Similar treatment was associated with (i) increased arachidonic acid incorporation into phosphatidylcholine, and (ii) decreased arachidonic acid incorporation into phosphatidylinositol and (iii) phosphatidylethanolamine. These effects were subsequently investigated by examining the acylation process, de novo biosynthesis, and the agonist-stimulated phosphoinositides hydrolysis in these cells. Our results indicated that sodium butyrate stimulated the acylation of arachidonic acid into lysophosphatidylcholine, lysophosphatidylethanolamine, and lysophosphatidylinositol. The glycerol incorporation into these lipids was not affected, but the inositol incorporation into total chloroform extracts and Pl and phosphatidylinositol 4-phosphate was decreased in the sodium butyrate-treated cells. Moreover, the accumulation of the rapid histamine-stimulated phosphoinositide metabolites, i.e., inositol monophosphate, inositol diphosphate, and inositol triphosphate (IP3) was decreased in these cells. To elucidate whether the decreased inositol phosphates were due to a decrease in the phosphoinositides hydrolysis, we measured the transient IP3 production directly by a receptor-binding assay. Our results indicated that histamine-stimulated transient IP3 formations were decreased. Taken together, these results indicated that multiple changes by multiple mechanisms of phospholipid metabolisms were found in sodium butyrate-treated C6 glioma cells. The decreased IP3 formation and its subsequent action, i.e., Ca2+ mobilization, may play an early but pivotal role by which sodium butyrate induces C6 glioma cell differentiation.

Effects of mercury on serotonin concentration in the brain of tilapia, Oreochromis mossambicus.

In order to know the effect of mercury pollution on the serotonergic system of fish, serotonin concentrations in a discrete brain region of tilapia, Oreochromis mossambicus, were examined. Serotonin concentration was measured using a high performance liquid chromatography system with electrochemical detector. In male fish, the concentrations of serotonin were 1.468 +/- 0.350, 0.811 +/- 0.190 and 0.330 +/- 0.061 micrograms/g wet tissue in hypothalamus, telencephalon and optic lobe, respectively. The serotonin content was significantly different between each region; the hypothalamus had a higher content than that of the telencephalon and optic lobe. The serotonin concentration in female hypothalamus was 1.102 +/- 0.112 micrograms/g wet tissue which was significantly lower than that in males. However, serotonin concentration in the telencephalon and optic lobe showed no difference between male and female. After exposure to 0.015 and 0.03 ppm HgCl2 for 6 months beginning 7 days posthatching, male sample fish showed a significantly dose-dependent decrease in serotonin concentration in the hypothalamus. But a similar phenomenon was not found in other regions of the brain. These results suggest that exposure to HgCl2 results in an attenuated development of the serotonergic system in the hypothalamus of fish.