LncRNA LINC01140 Inhibits Glioma Cell Migration and Invasion via Modulation of miR-199a-3p/ZHX1 Axis.

PURPOSE: Glioma is an aggressive tumor from the nervous system, which causes more than 70% of primary malignant brain tumors. Considering its severe malignancy, there is an urgent need to investigate more practical markers to understand the pathogenesis of glioma, and potential treatment methods for glioma patients. In the paper, we are focused on examining the roles of LINC01140, miR-199a-3p, and ZHX1 in the progression of gliomas, as well as their inner associations and modulation mechanisms. METHODS: qRT-PCR was employed to examine the expression levels of LINC01140 and miR-199a-3p. We measured the expressions of ZHX1 via qRT-PCR and Western blotting. CCK8 assays, migration assays, and invasion assays were carried out to determine the cell viabilities and abilities of migration and invasion. We also conducted in vivo tumor growth experiments to investigate the roles of LINC01140 in glioma developments. RESULTS: The expressions of LINC01140 were promoted in glioma. Silencing LINC01140 could inhibit glioma cell viabilities, migration, and invasion. In our experiments, miR-199a-3p was inhibited in glioma. LINC01140 negatively regulated the expressions of miR-199a-3p in glioma. MiR-199a-3p could target ZHX1 to inhibit its expression in glioma cells. CONCLUSION: LINC01140 could promote glioma developments by modulating the miR-199a-3p/ZHX1 axis.

New generation NMR bioreactor coupled with high-resolution NMR spectroscopy leads to novel discoveries in Moorella thermoacetica metabolic profiles.

An in situ nuclear magnetic resonance (NMR) bioreactor was developed and employed to monitor microbial metabolism under batch growth conditions in real time. We selected Moorella thermoacetica ATCC 49707 as a test case. M. thermoacetica (formerly Clostridium thermoaceticum) is a strictly anaerobic, thermophilic, acetogenic, gram-positive bacterium with potential for industrial production of chemicals. The metabolic profiles of M. thermoacetica were characterized during growth in batch mode on xylose (a component of lignocellulosic biomass) using the new generation NMR bioreactor in combination with high-resolution NMR (HR-NMR) spectroscopy. In situ NMR measurements were performed using water-suppressed H-1 NMR spectroscopy at 500 MHz, and aliquots of the bioreactor contents were taken for 600-MHz HR-NMR spectroscopy at specific intervals to confirm metabolite identifications and expand metabolite coverage. M. thermoacetica demonstrated the metabolic potential to produce formate, ethanol, and methanol from xylose, in addition to its known capability of producing acetic acid. Real-time monitoring of bioreactor conditions showed a temporary pH decrease, with a concomitant increase in formic acid during exponential growth. Fermentation experiments performed outside of the magnet showed that the strong magnetic field employed for NMR detection did not significantly affect cell metabolism. Use of the in situ NMR bioreactor facilitated monitoring of the fermentation process, enabling identification of intermediate and endpoint metabolites and their correlation with pH and biomass produced during culture growth. Real-time monitoring of culture metabolism using the NMR bioreactor in combination with HR-NMR spectroscopy will allow optimization of the metabolism of microorganisms producing valuable bioproducts.

Coregulation of Terpenoid Pathway Genes and Prediction of Isoprene Production in Bacillus subtilis Using Transcriptomics.

The isoprenoid pathway converts pyruvate to isoprene and related isoprenoid compounds in plants and some bacteria. Currently, this pathway is of great interest because of the critical role that isoprenoids play in basic cellular processes, as well as the industrial value of metabolites such as isoprene. Although the regulation of several pathway genes has been described, there is a paucity of information regarding system level regulation and control of the pathway. To address these limitations, we examined Bacillus subtilis grown under multiple conditions and determined the relationship between altered isoprene production and gene expression patterns. We found that with respect to the amount of isoprene produced, terpenoid genes fall into two distinct subsets with opposing correlations. The group whose expression levels positively correlated with isoprene production included dxs, which is responsible for the commitment step in the pathway, ispD, and two genes that participate in the mevalonate pathway, yhfS and pksG. The subset of terpenoid genes that inversely correlated with isoprene production included ispH, ispF, hepS, uppS, ispE, and dxr. A genome-wide partial least squares regression model was created to identify other genes or pathways that contribute to isoprene production. These analyses showed that a subset of 213 regulated genes was sufficient to create a predictive model of isoprene production under different conditions and showed correlations at the transcriptional level. We conclude that gene expression levels alone are sufficiently informative about the metabolic state of a cell that produces increased isoprene and can be used to build a model that accurately predicts production of this secondary metabolite across many simulated environmental conditions.

Novel monosaccharide fermentation products in Caldicellulosiruptor saccharolyticus identified using NMR spectroscopy.

BACKGROUND: Caldicellulosiruptor saccharolyticus is a thermophilic, Gram-positive, non-spore forming, strictly anaerobic bacterium of interest in potential industrial applications, including the production of biofuels such as hydrogen or ethanol from lignocellulosic biomass through fermentation. High-resolution, solution-state nuclear magnetic resonance (NMR) spectroscopy is a useful method for the identification and quantification of metabolites that result from growth on different substrates. NMR allows facile resolution of isomeric (identical mass) constituents and does not destroy the sample. RESULTS: Profiles of metabolites produced by the thermophilic cellulose-degrading bacterium Caldicellulosiruptor saccharolyticus DSM 8903 strain following growth on different monosaccharides (D-glucose, D-mannose, L-arabinose, D-arabinose, D-xylose, L-fucose, and D-fucose) as carbon sources revealed several unexpected fermentation products, suggesting novel metabolic capacities and unexplored metabolic pathways in this organism. Both 1H and 13C nuclear magnetic resonance (NMR) spectroscopy were used to determine intracellular and extracellular metabolite profiles. One dimensional 1H NMR spectral analysis was performed by curve fitting against spectral libraries provided in the Chenomx software; 2-D homonuclear and heteronuclear NMR experiments were conducted to further reduce uncertainties due to unassigned, overlapping, or poorly-resolved peaks. In addition to expected metabolites such as acetate, lactate, glycerol, and ethanol, several novel fermentation products were identified: ethylene glycol (from growth on D-arabinose), acetoin and 2,3-butanediol (from growth on D-glucose, L-arabinose, and D-xylose), and hydroxyacetone (from growth on D-mannose, L-arabinose, and D-xylose). Production of ethylene glycol from D-arabinose was particularly notable, with around 10% of the substrate carbon converted into this uncommon fermentation product. CONCLUSIONS: The present research shows that C. saccharolyticus, already of substantial interest due to its capability for biological ethanol and hydrogen production, has further metabolic potential for production of higher molecular weight compounds, such as acetoin and 2,3-butanediol, as well as hydroxyacetone and the uncommon fermentation product ethylene glycol. In addition, application of nuclear magnetic resonance (NMR) spectroscopy facilitates identification of novel metabolites, which is instrumental for production of desirable bioproducts from biomass through microbial fermentation.

Enhancing isoprene production by genetic modification of the 1-deoxy-d-xylulose-5-phosphate pathway in Bacillus subtilis.

To enhance the production of isoprene, a volatile 5-carbon hydrocarbon, in the Gram-positive spore-forming rod-shaped bacterium Bacillus subtilis, 1-deoxy-d-xylulose-5-phosphate synthase (Dxs) and 1-deoxy-d-xylulose-5-phosphate reductoisomerase (Dxr) were overexpressed in B. subtilis DSM 10. For the strain that overexpresses Dxs, the yield of isoprene was increased 40% over that by the wild-type strain. In the Dxr overexpression strain, the level of isoprene production was unchanged. Overexpression of Dxr together with Dxs showed an isoprene production level similar to that of the Dxs overproduction strain. The effects of external factors, such as stress factors including heat (48°C), salt (0.3 M NaCl), ethanol (1%), and oxidative (0.005% H(2)O(2)) stress, on isoprene production were further examined. Heat, salt, and H(2)O(2) induced isoprene production; ethanol inhibited isoprene production. In addition, induction and repression effects are independent of SigB, which is the general stress-responsive alternative sigma factor of Gram-positive bacteria.

Exogenous or L-rhamnose-derived 1,2-propanediol is metabolized via a pduD-dependent pathway in Listeria innocua.

1,2-Propanediol (1,2-PD) added exogenously to cultures or produced endogenously from l-rhamnose is metabolized to n-propanol and propionate in Listeria innocua Lin11. The pduD gene, which encodes a diol dehydratase ss subunit homolog, is required for 1,2-PD catabolism. pduD and 16 other genes within the pduA-to-pduF region of a large gene cluster are induced in medium containing 1,2-PD.

Regulation of the mpt operon in Listeria innocua by the ManR protein.

The phosphotransferase system regulation domain (PRD)-containing activator, ManR, is required for glucose-controlled transcription of the mannose permease two (mpt) operon in Listeria innocua. His-871 in ManR PRD-II is needed for mpt repression in glucose-free media. His-506 in PRD-I is needed for mpt induction by glucose.

Dissolved organic matter (DOM) in recycled leachate of bioreactor landfill.

Landfill leachate needs sufficient treatment before safe disposal. Bioreactor landfill technology could effectively degrade the organic matters in recirculated leachate, hence leaving a leachate stream of low biodegradability. This study characterized the dissolved organic matter (DOM) in the leachate from simulated bioreactor landfill columns with or without presence of trace oxygen. The removal efficiencies of this DOM using coagulation-sedimentation or electrolysis processes were demonstrated. Recirculated leachates were sampled from the simulated landfill columns applying conventional mode, intermittent-aeration mode, and natural aeration mode, whose DOM was fractionated into humic acids (HA), fulvic acids (FA) and hydrophilic fractions (HyI) by the XAD-8 resin combined with the cation exchange resin method. The recirculated leachate had low BOD/COD ratio, high humic substances contents, and high aromatic content. Their HA fraction comprised mainly large molecules (>10 k Da), while the FA and HyI were composed of smaller molecules (<50 k and <4 k Da, respectively). With the presence of oxygen, the TOC contents and the contents of HA, FA and HyI in leachate reduced, with FA and HyI fractions of molecular weight (MW) lower than 4 k Da more readily degraded. The organic matters left in leachates from intermittent-aeration mode and natural aeration mode were of low biodegradability. It was tested in the following sections the effects of coagulation-sedimentation process and of electrolysis process on the removal of residual DOM in recirculated leachate. Coagulation-sedimentation tests revealed that poly ferric sulphate (PFS) could remove more COD (58.1%) from leachate than polyaluminum chloride (PACl) (22.9%), particularly on the HA fraction with MW>10 k Da. Coagulation-sedimentation could not remove most of HyI in leachate. Furthermore, the corresponding BOD/COD ratio was not improved through coagulation. Electrolysis test could also effectively removed HA of MW>10 k Da. However, the biodegradability of treated effluent considerably was improved. The electrolysis could decompose high MW substances and increase biodegradability of recirculated leachate from bioreactor landfill.

Novel activator of mannose-specific phosphotransferase system permease expression in Listeria innocua, identified by screening for pediocin AcH resistance.

To identify genes that are important for class IIa bacteriocin interaction and resistance in Listeria species, transposon Tn917 knockout libraries were constructed for Listeria innocua strain Lin11 and screened for mutants that are resistant to pediocin AcH. A highly resistant mutant (G7) (MIC > 20 microg/ml; 1,000-fold less susceptible than the wild type), in which the transposon integrated into the putative promoter of the lin0142 gene, was isolated. lin0142 is located immediately upstream of the mpt operon (mptA/mptC/mptD) that encodes the mannose-specific phosphoenolpyruvate-dependent phosphotransferase system permease EIItMan, which serves as a docking protein for class IIa bacteriocins. The transcription of the mpt operon is known to be positively controlled by sigma54 factor and ManR (a sigma54-associated activator). Transcripts for lin0142 and mpt were undetectable in the G7 mutant, based on quantitative real-time reverse transcriptase PCR analysis. When the wild-type lin0142 gene was expressed at a 7.9-fold-elevated level in the mutant via a multicopy-number plasmid, the level of mpt mRNA became 70% higher than that in the wild-type strain. In addition, the complementation strain reverted back to the pediocin AcH-susceptible phenotype. The levels of manR and rpoN (sigma54) mRNAs were not directly influenced by the level of lin0142 transcription. lin0142 is the only one of the three mpt regulatory genes whose transcription is induced, albeit slightly (1.2-fold), by glucose. The combined results show that the lin0142 gene encodes a novel activator of the mpt operon. The Lin0142 protein contains a winged-helix DNA-binding motif and is distantly related to the Crp-Fnr family of transcription regulators.