CircRNA hsa_circRNA_101996 increases cervical cancer proliferation and invasion through activating TPX2 expression by restraining miR-8075

In recent years, circular RNAs have been shown to serve as essential regulators in several human cancers. Nevertheless, the function and mechanism of CircRNA in cervical cancer remain elusive. In the present study, we showed that hsa_circRNA_101996 was highly expressed in cervical cancer tissues compared with matched normal tissues by bioinformatics analysis. We showed that the expression level of hsa_circRNA_101996 in cervical cancer tissues was positively correlated with TNM stage, tumor size, and lymph node metastasis. Moreover, higher levels of hsa_circRNA_101996 were related to poor outcomes of cervical cancer patients. We found that knockdown of hsa_circRNA_101996 significantly inhibited the proliferation, cell cycle, migration, and invasion of cervical cancer cells. Mechanistically, we demonstrated that hsa_circRNA_101996 served as a sponge of miR-8075, which targeted TPX2 in cervical cancer cells. We showed that miR-8075 that was downregulated in cervical cancer tissues repressed cervical cancer cell proliferation, migration, and invasion. Furthermore, we validated that upregulation of TPX2 by hsa_circRNA_101996-mediated inhibition of miR-8075 contributed to cervical cancer proliferation, migration, and invasion. Taken together, our findings revealed a novel mechanism that hsa_circRNA_101996-miR-8075-TPX2 network promoted cervical cancer progression.     

金铁锁转录因子ptMYC2的克隆和生物信息学分析

金铁锁是 “云南白药”等多种中成药的重要组成,其有效成分为三萜皂苷,MYC类转录因子在调节植物三萜类次生代谢积累中有重要作用。为研究ptMYC2 基因在金铁锁三萜皂苷合成代谢途径的调控机制,该研究基于金铁锁转录组测序数据,克隆得到ptMYC2转录因子的两个全长基因; 通过生物信息学软件对两条转录因子的同源性、理化性质、疏水性、跨膜结构、亚细胞定位、结构域、靶基因结合位点等进行初步预测分析。结果表明:两条转录因子所编码的蛋白属于亲水性蛋白,不存在跨膜区域,均是非分泌蛋白质,且不存在信号肽; 两条转录因子的亚细胞定位于细胞核; 结构域分析显示,两个基因都含有bHLH家族结构域; 预测得到金铁锁三萜皂苷合成途径中HMGR、FPS、SE、β-AS等基因的启动子可能存在与MYC2结合的E-box特异性结合位点。该研究结果将为进一步研究ptMYC2基因在金铁锁三萜皂苷合成代谢途径的调节机制奠定基础。     

Overexpression of GLT1 in fps1DeltagpdDelta mutant for optimum ethanol formation by Saccharomyces cerevisiae

Glycerol is the main byproduct produced under anaerobic ethanol fermentations by Saccharomyces cerevisiae and consumes a considerable amount of substrate. To verify the metabolic phenotype predications for increasing ethanol formation, two engineered S. cerevisiae KAM-14, KAM-15 strains were constructed for possible redirection of glycerol carbon flux into ethanol by overexpression of GLT1 in the fps1DeltagpdDelta mutant. The engineered strains KAM-14 and KAM-15 compared to the control strain KAM-2, produced 12.24% and 10.42% higher ethanol, 39.72% and 31.03% lower glycerol yield during anaerobic batch fermentations, respectively. The maximum specific growth rates of KAM-14 and KAM-15 were found to be relatively lower than that of KAM-2 during the exponential growth phase. In the meantime, the biomass concentrations of both KAM-14 and KAM-15 were similar to KAM-2. Acetate and pyruvate concentrations of KAM-14 and KAM-15 were greatly decreased comparing to those of KAM-2, respectively. These experimental results approved the metabolic pathway strategies to improve ethanol formation.     

Bi-functional peptides with both trypsin-inhibitory and antimicrobial activities are frequent defensive molecules in Ranidae amphibian skins

Amphibian skins act as the first line against noxious aggression by microorganisms, parasites, and predators. Anti-microorganism activity is an important task of amphibian skins. A large amount of gene-encoded antimicrobial peptides (AMPs) has been identified from amphibian skins. Only a few of small protease inhibitors have been found in amphibian skins. From skin secretions of 5 species (Odorrana livida, Hylarana nigrovittata, Limnonectes kuhlii, Odorrana grahami, and Amolops loloensis) of Ranidae frogs, 16 small serine protease inhibitor peptides have been purified and characterized. They have lengths of 17-20 amino acid residues (aa). All of them are encoded by precursors with length of 65-70 aa. These small peptides show strong trypsin-inhibitory abilities. Some of them can exert antimicrobial activities. They share the conserved GCWTKSXXPKPC fragment in their primary structures, suggesting they belong to the same families of peptide. Signal peptides of precursors encoding these serine protease inhibitors share obvious sequence similarity with those of precursors encoding AMPs from Ranidae frogs. The current results suggest that these small serine protease inhibitors are the common defensive compounds in frog skin of Ranidae as amphibian skin AMPs.     

Inhibition of Klebsiella pneumoniae ATCC 13883 cells by hexane extract of Halimeda discoidea (Decaisne) and the identification of its potential bioactive compounds

The inhibitory effect of the Klebsiella pneumoniae ATCC 13883 strain caused by the hexane extract of Halimeda discoidea (Nor Afifah et al., 2010) was further evaluated by means of the microscopy view and its growth curves. The morphological changes of the K. pneumoniae ATCC 13883 cells were observed under the scanning electron microscope (SEM) and transmission electron microscope (TEM) after they were treated at minimum inhibitory concentration (MIC; 0.50 mg/ml) (Nor Afifah et al., 2010) for 12, 24, and 36 h. The results showed the severity of the morphological deteriorations experienced by the treated cells. The killing curve assay was performed for 48 h at three different extract concentrations (1/2 MIC, MIC, and 2 MIC). An increase in the extract concentration of up to 2 MIC value did significantly reduce the number of cells by approximately 1.9 log10, as compared with the control. Identification of the potential compounds of the extract responsible for the antibacterial activity was carried out through the gas chromatography-mass spectrum (GCMS) analysis of the active subfraction, and the compound E-15-heptadecenal was identified and suggested as the most potential antibacterial compound of this extract. The subsequent cellular degenerations showed by the data might well explain the inhibitory mechanisms of the suggested antibacterial compound. All of these inhibitory effects have further proven the presence of an antibacterial compound within H. discoidea that can inhibit the growth of K. pneumoniae ATCC 13883.     

Experimental study of intracellular ice growth in human umbilical vein endothelial cells

Study of the intracellular ice formation (IIF) and growth is essential to the mechanistic understanding of cellular damage through freezing. In the aid of high speed and high resolution cryo-imaging technology, the transient intracellular ice formation and growth processes of the attached human umbilical vein endothelial cells (HUVEC) were successfully captured during freezing. It was found that the intracellular ice nucleation site was on the cell membrane closer to the nucleus. The ice growth was directional and toward the nucleus, which covered the whole nucleus before growing into the cytoplasm. The crystal growth rate in the nucleus was much larger than that in the cytoplasm, and its morphology was influenced by the cooling rate. During the thawing process, small crystals fused into larger ones inside the nucleus. Moreover, the cumulative fraction of the HUVEC with IIF was mainly dependent on the cooling rate not the confluence of the cells attached.     

The inhaled phosphodiesterase 4 inhibitor GSK256066 reduces allergen challenge responses in asthma

GSK256066 is a selective phosphodiesterase 4 inhibitor that can be given by inhalation, minimising the potential for side effects. We evaluated the effects of GSK256066 on airway responses to allergen challenge in mild asthmatics.

Methods

In a randomised, double blind, cross-over study, 24 steroid naive atopic asthmatics with both early (EAR) and late (LAR) responses to inhaled allergen received inhaled GSK256066 87.5 mcg once per day and placebo for 7 days, followed by allergen challenge. Methacholine reactivity was measured 24 h post-allergen. Plasma pharmacokinetics were measured. The primary endpoint was the effect on LAR.

Results

GSK256066 significantly reduced the LAR, attenuating the fall in minimum and weighted mean FEV1 by 26.2% (p = 0.007) and 34.3% (p = 0.005) respectively compared to placebo. GSK256066 significantly reduced the EAR, inhibiting the fall in minimum and weighted mean FEV1 by 40.9% (p = 0.014) and 57.2% (p = 0.014) respectively compared to placebo. There was no effect on pre-allergen FEV1 or methacholine reactivity post allergen. GSK256066 was well tolerated, with low systemic exposure; plasma levels were not measurable after 4 hours in the majority of subjects.

Conclusions

GSK256066 demonstrated a protective effect on the EAR and LAR. This is the first inhaled PDE4 inhibitor to show therapeutic potential in asthma.

Trial Registration

This study is registered on clinicaltrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00380354","term_id":"NCT00380354"}}NCT00380354     

Functional analysis of the YopE GTPase-activating protein (GAP) activity of Yersinia pseudotuberculosis

YopE of Yersinia pseudotuberculosis inactivates three members of the small RhoGTPase family (RhoA, Rac1 and Cdc42) in vitro and mutation of a critical arginine abolishes both in vitro GTPase-activating protein (GAP) activity and cytotoxicity towards HeLa cells, and renders the pathogen avirulent in a mouse model. To understand the functional role of YopE, in vivo studies of the GAP activity in infected eukaryotic cells were conducted. Wild-type YopE inactivated Rac1 as early as 5 min after infection whereas RhoA was down regulated about 30 min after infection. No effect of YopE was found on the activation state of Cdc42 in Yersinia-infected cells. Single-amino-acid substitution mutants of YopE revealed two different phenotypes: (i) mutants with significantly lowered in vivo GAP activity towards RhoA and Rac1 displaying full virulence in mice, and (ii) avirulent mutants with wild-type in vivo GAP activity towards RhoA and Rac1. Our results show that Cdc42 is not an in vivo target for YopE and that YopE interacts preferentially with Rac1, and to a lesser extent with RhoA, during in vivo conditions. Surprisingly, we present results suggesting that these interactions are not a prerequisite to establish infection in mice. Finally, we show that avirulent yopE mutants translocate YopE in about sixfold higher amount compared with wild type. This raises the question whether YopE's primary function is to sense the level of translocation rather than being directly involved in downregulation of the host defence.