Melatonin, a novel selective ATF-6 inhibitor, induces human hepatoma cell apoptosis through COX-2 downregulation

AIM To clarify the mechanisms involved in the critical endoplasmic reticulum(ER) stress initiating unfolded protein response pathway modified by melatonin.METHODS Hepatoma cells, Hep G2, were cultured in vitro. Flow cytometry and TUNEL assay were used to measure Hep G2 cell apoptosis. Western blotting and quantitative reverse transcription-polymerase chain reaction methods were used to determine the protein and messenger RNA levels of ER stress and apoptosis related genes' expression, respectively. Tissue microarray construction from patients was verified by immunohistochemical analysis.RESULTS In the present study, we first identified that melatoninselectively blocked activating transcription factor 6(ATF-6) and then inhibited cyclooxygenase-2 (COX-2) expression, leading to enhanced liver cancer cell apoptosis under ER stress condition. Dramatically increased CCAAT-enhancer-binding protein homologous protein level, suppressed COX-2 and decreased Bcl-2/Bax ratio by melatonin or ATF-6 si RNA contributed the enhanced Hep G2 cell apoptosis under tunicamycin (an ER stress inducer) stimulation. In clinical hepatocellular carcinoma patients, the close relationship between ATF-6 and COX-2 was further confirmed.CONCLUSION These findings indicate that melatonin as a novel selective ATF-6 inhibitor can sensitize human hepatoma cells to ER stress inducing apoptosis.

Murine hepatocellular carcinoma derived stem cells reveal epithelial-to-mesenchymal plasticity

AIM To establish a model to enrich and characterize stemlike cells from murine normal liver and hepatocellular carcinoma(HCC) cell lines and to further investigate stem-like cell association with epithelial-to-mesenchymal transition(EMT).METHODS In this study,we utilized a stem cell conditioned serumfree medium to enrich stem-like cells from mouse HCC and normal liver cell lines,Hepa 1-6 and AML12,respectively.We isolated the 3-dimensional spheres and assessed their stemness characteristics by evaluating theRNA levels of stemness genes and a cell surface stem cell marker by quantitative reverse transcriptase-PCR(q RTPCR).Next,we examined the relationship between stem cells and EMT using q RT-PCR.RESULTS Three-dimensional spheres were enriched by culturing murine HCC and normal hepatocyte cell lines in stem cell conditioned serum-free medium supplemented with epidermal growth factor,basic fibroblast growth factor and heparin sulfate.The 3-dimensional spheres had enhanced stemness markers such as Klf4 and Bmi1 and hepatic cancer stem cell(CSC) marker Cd44 compared to parental cells grown as adherent cultures.We report that epithelial markers E-cadherin and ZO-1 were downregulated,while mesenchymal markers Vimentin and Fibronectin were upregulated in 3-dimensional spheres.The 3-dimensional spheres also exhibited changes in expression of Snai,Zeb and Twist family of EMT transcription factors.CONCLUSION Our novel method successfully enriched stem-like cells which possessed an EMT phenotype.The isolation and characterization of murine hepatic CSCs could establish a precise target for the development of more effective therapies for HCC.

Expression of transcription factors Slug in the lens epithelial cells undergoing epithelial-mesenchymal transition induced by connective tissue growth factor

AIM: To investigate the expression of transcription factors Slug in human lens epithelial cells(HLECs)undergoing epithelial-mesenchymal transition(EMT)induced by connective tissue growth factor(CTGF).·METHODS: HLECs were treated with CTGF of different concentrations(20, 50 and 100 ng/m L) or without CTGF(control) for 24 h. The morphological changes of HLECs were analysed by microscopy. The expression and cellular localization of Slug was evaluated by immumo-fluorescence. Expressions of Slug, E-cadherin and alpha smooth muscle actin(α-SMA) were further determined by Western blot analysis.·RESULTS: HLECs showed spidle fibrolasts-like characteristics and loosely connected each other after CTGF treatment. The immuno-fluorescence staining indicated that Slug was localized in the nuclei and its expression was induced by CTGF. The relative expressions of Slug protein were 1.64 ±0.11, 1.96 ±0.03,3.12 ±0.10, and 4.08 ±0.14, respectively, in response to control group and treatment with CTGF of 20, 50 and100 ng/m L(F =443.86, P <0.01). The increased Slug protein levels were correlated well with up-expression of α-SMA(0.78±0.05, 0.85±0.06, 2.17±0.15, 2.86±0.10; F =449.85, P <0.01) and down-expression of E-cadherin(2.50 ±0.11,1.79±0.26, 1.05±0.14, 0.63±0.08; F =101.55, P <0.01).·CONCLUSION: Transcription factor Slug may be involved in EMT of HLECs induced by CTGF in vitro.

Hairy Leaf 6, an AP2/ERF Transcription Factor, Interacts with OsWOX3B and Regulates Trichome Formation in Rice

毛状体形成广泛地在植物为表皮的房间区别和房间形态发生作为一个机械学的模型被学习了。然而，位于毛状体形成下面的基因、分子的机制(即，开始和延伸) 在米饭仍然保持大部分不清楚。这里，我们报导一个 AP2/ERF 抄写因素，毛乎乎的叶 6 (HL6 ) ，它在米饭控制毛状体形成。功能的分析表明 HL6 transcriptionally 在米饭调整毛状体延伸，它依赖于功能的 OsWOX3B，在毛状体充当一个关键管理者的包含 homeodomain 蛋白质开始。生物化学、分子的基因分析证明那 HL6 身体上交往与 OsWOX3B，和他们俩在毛状体形成， OsWOX3B 多半与它的直接目标基因之一在提高 HL6 的有约束力的能力期间调整一些植物生长素相关的基因的表达式， OsYUCCA5。人口基因分析显示 HL6 在米饭驯服期间在否定选择下面。一起拿，我们的调查结果提供新卓见进毛状体形成的分子的规章的网络在 ? 米饭。

ABF2, ABF3, and ABF4 Promote ABA-Mediated Chlorophyll Degradation and Leaf Senescence by Transcriptional Activation of Chlorophyll Catabolic Genes and Senescence-Associated Genes in Arabidopsis

叶绿素(Chl ) 降级是叶老朽的一个不可分的过程，并且 NYE1/SGR1 在多样的工厂种类作为 Chl 分解代谢的一个关键管理者被表明了。在这研究，用屏蔽的酵母一个混血儿，我们识别了三 abscisic 酸(骆驼毛的织物) 应答的元素(ABRE ) 绑定抄写因素， ABF2 (AREB1 ) ， ABF3，和 ABF4 (AREB2 ) ， NYE1 倡导者的通常认为的有约束力的蛋白质。通过 transactivation 分析， electrophoretic 活动性移动试金，和染色质 immunoprecipitation，我们表明了直接跳到并且在 vitro 并且在 vivo 激活 NYE1 倡导者的那 ABF2， ABF3，和 ABF4。骆驼毛的织物是叶老朽的一个积极管理者，并且外长地适用骆驼毛的织物能加速 Chl 降级。ABF， abf2abf3abf4，以及二骆驼毛的织物感觉迟钝的异种， abi1-1 和 snrk2.2/2.3/2.6 的三倍的异种，在骆驼毛的织物处理以后的展出停留绿色显型，与 NYE1 和 NYE2 表示的减少的正式就职一起。相反， ABF4 的 overexpression 在骆驼毛的织物处理之上加速了 Chl 降级。有趣地， ABF2/3/4 能也激活二 Chl 分解代谢的酶基因， PAO 和 NYC1 的表达式，由对他们的倡导者直接有约束力。另外， abf2abf3abf4 展出了功能的停留绿色显型，和联系老朽的基因(下垂) 例如 SAG29 (SWEET15 ) ，可能被 ABF 直接调整。一起拿，我们的结果在调停作为关键管理者建议那 ABF2， ABF3，和 ABF4 可能的行为被触发骆驼毛的织物的 Chl 降级和叶老朽一般来说在 Arabidopsis。

Arabidopsis Transcription Factors SPL1 and SPL12 Confer Plant Thermotolerance at Reproductive Stage

植物繁殖机关是脆弱的加热，但是在开花期的热吃惊回答的规定大部分是 uncharacterized。这里，我们报导二 SQUAMOSA 倡导者绑定像蛋白质(系统程序设计语言) 在 Arabidopsis， SPL1 和 SPL12 的 transcriptional 因素，在繁殖阶段在 thermotolerance 冗余地行动。spl1-1 spl12-1 开花期显示了超敏性加热应力，而 SPL1 或 SPL12 的 overexpression 在 Arabidopsis 和烟草提高了 thermotolerance。定序的 RNA 在热应力，一个季度(1,040 ) 在 spl1-1 spl12-1 在之中被调整之上在野类型的开花期揭示了 1939 upregulated 和 1479 downregulated 基因，显示 SPL1 和 SPL12 极大地作出贡献到在开花期的被触发热的 transcriptional reprogramming。尤其是，热应力导致了很多 abscisic 酸(骆驼毛的织物) 应答的基因， 39% 在在 spl1-1 spl12-1 感应的发情被扰乱开花期。骆驼毛的织物的 Preapplication 和 SPL1 的 overexpression 在 spl1-1 spl12-1 并且在 ABA 生合成异种 aba2-1，然而并非在 pyl 恢复了开花期 thermotolerance 在骆驼毛的织物受体 PYR1/PYL1/PYL2/PYL4/PYL5/PYL8 的六倍的变异的有缺陷者。因此， SPL1 和 SPL2 授与的开花期 thermotolerance 包含调停 PYL 的骆驼毛的织物发信号。由 SPL1 和 SPL12 组成的分子的网络这里说明了在繁殖舞台在植物 thermotolerance 的机制上打开新灯。

OsLEC1/OsHAP3E Participates in the Determination of Meristem Identity in Both Vegetative and Reproductive Developments of Rice

In the vegetative phase of plant development, the shoot apical meristem (SAM) produces leaf primordia in regular phyllotaxy, and transforms to the inflorescence meristem when the plant enters reproductive growth, which will undergo a series of identity differentiations and will finally form a complete and fertile panicle. Our previous studies indicated a tissue-specific expression pattern of the OsLEC1 (leafy cotyledon) gene, which is homologous to the Arabidopsis AtLEC1 gene and belongs to the CCAAT-binding protein HAP3 subfamily, during embryo development. Expression of additional OsLEC1 genomic sequences resulted in abnormalities in the development of leaves, panicles and spikelets. The spikelets in particular presented abnormities, including panicle and spikelet-like structures that occurred reiteratively inside prior spikelets, and the occasional spikelet structures that completely transformed into plantlets (a reproductive habit alteration from sexual to asexual called "pseudovivipary"). Analysis showed that OsLEC1 interacts with several SEPALLATA-like MADS transcription factors, suggesting that increased levels of the OsLEC1 protein might interfere with the normal interaction network of these MADS proteins and lead to defective spikelet development. The expression of OsMADS1 was dramatically reduced, and the DNA methylation level of cytosine in certain regions of the OsMADS1 promoter was increased under OsLEC1 overexpression. These results indicate that OsLEC1 affects the development of leaves, panicles and spikelets, and is a key regulator of meristem identity determination in both rice (Oryza sativa) vegetative and reproductive development.

LJbiquitin--Proteasome System in ABA Signaling： From Perception to Action

由 ubiquitination 的蛋白质 translational 以后修正(PTM ) 在工厂生长，开发，和压力回答的许多方面期间被观察了。ubiquitin-proteasome 系统精确调整植物激素由影响蛋白质活动，本地化，汇编，和相互作用能力发信号。Abscisic 酸(骆驼毛的织物) 是主要植物激素，并且在正常或强调的生长条件下面在植物起重要作用。表明小径的骆驼毛的织物由磷酸酶， kinases，抄写因素，和膜离子隧道组成。多重骆驼毛的织物发信号变换器被 ubiquitination 受到规定，这被报导了。特别地，最近的研究识别了 E3 ligases 的不同类型调停在不同房间分隔空间的骆驼毛的织物受体的 ubiquitination。这评论由发生在血浆膜的 monoubiquitination 或 polyubiquitination 集中于这些部件的调整， endomembranes，并且从 cytosol 到原子核；这暗示存在后退并且被 ubiquitination 在骆驼毛的织物发信号调整的 trafficking 过程。很多单个单位的 E3 ligases，多子单元 E3 ligases 的部件， E2s，和涉及骆驼毛的织物信号规定的 26S proteasome 的特定的子单元被讨论。把在 ABA 小径的 ubiquitination 的精确功能可以帮助我们在发信号 ubiquitination 和 PTM 的另外的类型调整的另外的植物激素理解关键因素。

The role of micro RNAs in hepatocyte metabolism and hepatitis B virus replication

Though efficient vaccines against hepatitis B virus(HBV) and antiviral therapies are available,chronic HBV infection is still a global health problem. The process of HBV infection and HBV life cycle are extensively studied in last decades, however, the mechanisms of HBV-induced alterations of host cell metabolisms and host factors involved in modulating of viral replication are not fully understood. Thus, it is an important issue to examine these specific HBV-host interactions for development of novel strategies for antiviral therapies. Recently, microRNAs(miRNAs), a class of post-transcriptional regulatory small RNA, seem to be the relevant fine tuning factors of various cellular activities and pathways, including cell growth, metabolism, and viral replication. In this review, we summarize the up to date knowledge concerning the virus-host interactions and emphasizing on the role of miRNAs in regulation of HBV replication and host cell metabolism.

Conservation and Diversification of the SHR-SCR- SCL23 Regulatory Network in the Development of the Functional Endodermis in Arabidopsis Shoots

部分地， Arabidopsis thaliana 根的功能的内皮的开发被 GRAS 抄写因素控制，也就是短根(SHR ) ，衣衫褴褛的人(SCR ) ，并且像衣衫褴褛的人 23 (SCL23 ) 。最近， SHR-SCR-SCL23 规章的模块为内皮的说明也是必要的，这被显示出(作为捆鞘知道) 在叶子。与在根对 SHR-SCR-SCL23 规章的网络的角色被知道的相比，不过， SHR， SCR，和 SCL23 的分子的相互作用更不在射击被理解。这里，我们证明 SHR 与 SCL23 形成蛋白质建筑群在射击调整 SCL23 的抄写，类似于 SCR 的规定模式表示。我们的结果显示 SHR 充当主人管理者直接激活 SCR 和 SCL23 的表示。在 SHR-SCR-SCL23 网络，我们发现了一以前 SCL23 调制 SHR 的否定反馈循环铺平的 uncharacterized。通过分子、基因、生理、词法的分析，我们也表明 SHR-SCR-SCL23 模块在内皮的形成起一个关键作用(作为淀粉鞘知道) 在胚轴。一起拿，我们的结果在根和射击在内皮开发提供新卓见进 SHR-SCR-SCL23 网络的规章的角色。

Transcriptional and Epigenetic Regulation in Injury-Mediated Neuronal Dendritic Plasticity

Injury to the nervous system induces localized damage in neural structures and neuronal death through the primary insult,as well as delayed atrophy and impaired plasticity of the delicate dendritic fields necessary for interneuronal communication. Excitotoxicity and other secondary biochemical events contribute to morphological changes in neurons following injury. Evidence suggests that various transcription factors are involved in the dendritic response to injury and potential therapies. Transcription factors play critical roles in the intracellular regulation of neuronal morphological plasticity and dendritic growth and patterning. Mounting evidence supports a crucial role for epigenetic modifications via histone deacetylases,histone acetyltransferases,and DNA methyltransferases that modify gene expression in neuronal injury and repair processes.Gene regulation through epigenetic modification is of great interest in neurotrauma research,and an early picture is beginning to emerge concerning how injury triggers intracellular events that modulate such responses. This review provides an overview of injury-mediated influences on transcriptional regulation through epigenetic modification,the intracellular processes involved in the morphological consequences of such changes,and potential approaches to the therapeutic manipulation of neuronal epigenetics for regulating gene expression to facilitate growth and signaling through dendritic arborization following injury.

Identification of WRKY Transcription Factors Related to Saikosaponin Biosynthesis in Adventitious Roots of Bupleurum chinense

Objective To identify the genes of WRKY transcription factors(TFs) from roots of Bupleurum chinense and genes that potentially regulate saikosaponin(SS) biosynthesis.Methods Firstly,the subfamily cluster analysis was mainly based on Arabidopsis thaliana WRKYs for 27 putative WRKY TFs selected from previous transcriptome sequencing data.Secondly,qPCR was used to screen such genes of WRKY TFs that could be induced by NaCI and PEG6000 in adventitious roots of B.chinense.Meanwhile,saikosaponins(SSs) in treated adventitious roots were determined by HPLC.The roots were collected at 0,2,4,8,12,24,48,and 72 h after treatments,and 120 h only for PEG.Finally,the tissue-specific expression was analyzed on screened genes by qPCR.Results The 27 genes were grouped into three categories:There were nine in Group Ⅰ,15 in Group Ⅱ,and two in Group Ⅲ.Four genes of WRKYTFs,BCWRKY6,BCWRKY16,BCWRKY32,and BCWRKY35 were obviously induced by NaCI in adventitious roots of B.chinense,while only BCWRKY32 was induced by PEG.The content of SSs increased at different levels in NaCI and PEG6000 treatment.Three genes including BCWRKY6,BCWRKY32,and BCWRKY35,expressed most in roots,were similar to the accumulation pattern of SS.Conclusion The three WRKY genes,BCWRKY6,BCWRKY32,and BCWRKY35,may be involved in the biosynthesis of SS.

Expression level of pre-B-cell leukemia transcription factor 2 (PBX2) as a prognostic marker for gingival squamous cell carcinoma

Objective: In this study, we investigated the interrelationship between clinicopathologic findings and pre-B-cell leukemia transcription factor 2 (PBX2) expression in gingival squamous cell carcinoma (GSCC). Methods: Expression level of PBX2 was immunohistochemically examined in 66 GSCC subjects (30 men and 36 women) with ages ranging from 42 to 85 (median 64.5) years, in which staining intensity in tumor cells was categorized as either weaker (level 1) or equal to/stronger (level 2) than that in the endothelial cells. Results: PBX2 expression is correlated with valosin-containing protein (VCP) expression. Univariate and multivariate analyses revealed a high level of PBX2 expression to be a poor prognosticator for disease-free survival (DFS) and overall survival (OS), and PBX2 expression was an independent prognostic factor for both DFS and OS in GSCC. Conclusions: PBX2 expression level in GSCC is prognostic. PBX2 may be a useful marker to identify the potential for progression in GSCC.