14-3-3ε Boosts Bleomycin-induced DNA Damage Response by Inhibiting the Drug-resistant Activity of MVP

Major vault protein (MVP) is the predominant constituent of the vault particle, the largest known ribonuclear protein complex. Although emerging evidence have been establishing the links between MVP (vault) and multidrug resistance (MDR), little is known regarding exactly how the MDR activity of MVP is modulated during cellular response to drug-induced DNA damage (DDR). Bleomycin (BLM), an anticancer drug, induces DNA double-stranded breaks (DSBs) and consequently triggers the cellular DDR. Due to its physiological implications in hepatocellular carcinoma (HCC) and cell fate decision, 14-3-3ε was chosen as the pathway-specific bait protein to identify the critical target(s) responsible for HCC MDR. By using an LC–MS/MS-based proteomic approach, MVP was first identified in the BLM-induced 14-3-3ε interactome formed in HCC cells. Biological characterization revealed that MVP possesses specific activity to promote the resistance to the BLM-induced DDR. On the other hand, 14-3-3ε enhances BLM-induced DDR by interacting with MVP. Mechanistic investigation further revealed that 14-3-3ε, in a phosphorylation-dependent manner, binds to the phosphorylated sites at both Thr52 and Ser864 of the monomer of MVP. Consequently, the phosphorylation-dependent binding between 14-3-3ε and MVP inhibits the drug-resistant activity of MVP for an enhanced DDR to BLM treatment. Our findings provide an insight into the mechanism underlying how the BLM-induced interaction between 14-3-3ε and MVP modulates MDR, implicating novel strategy to overcome the chemotherapeutic resistance through interfering specific protein–protein interactions

Possibilities of increasing interest in foster care in the Czech Republic

For most people, family represents a place where they feel safe and loved. But not everyone is lucky enough to have a place like this, to have a functional family. If the biological family does not work as it should and a baby is threatened in some way, the state has several options to help the child. If there are solvable problems in the family, the state can try to help it by various interventions to improve the situation. But if these actions do not help and the child is forced to leave this family, it is necessary to ensure alternative environment for baby. The child can either be placed in institutional care, or to substitutional family care. Considering the shortcomings of institutional care, various forms of substitutional care are mainly supported. Probably the best known form of substitutional care is adoption. In the case of adoption, baby becomes the member of new family and relationships between child and his biological family vanishes. This form is only possible for a small percentage of abandoned children, because most of them still has some kind of relationship with biological family. For these children, there is an alternative, namely foster care. In the case of foster care, child grows up in a foster family, foster parents care for him every day, but the child still has a..

ACC Deaminase from <i>Lysobacter gummosus</i> OH17 Can Promote Root Growth in <i>Oryza sativa</i> Nipponbare Plants

Although <i>Lysobacter</i> species are a remarkable source of natural compounds with antibacterial and antifungal activities, the ability of these bacteria to produce plant growth promoters remains practically unknown. In this work, the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) has been isolated from the secretions of <i>Lysobacter gummosus</i> OH17, indicating the presence of an ACC deaminase, which was shown to be encoded in the gene <i>peg_1256</i>. The recombinant enzyme could not only deaminate ACC to provide 2-oxobutanoic acid but also catalyzed the amination of the 2-oxobutanoic acid, demonstrating, for the first time, that ACC deaminases can produce ACC. After the treatment of rice <i>Oryza sativa</i> Nipponbare plants with OH17 ACC deaminase, the ethylene production levels were 44% higher in comparison with the control experiments, allowing significant improvements in root, 10%, and stem, 14%, growth

PPE38 of Mycobacterium marinum Triggers the Cross-Talk of Multiple Pathways Involved in the Host Response, As Revealed by Subcellular Quantitative Proteomics

The PE/PPE family of proteins which are in high abundance in pathogenic species such as Mycobacterium tuberculosis and M. marinum, play the critical role in generating antigenic variation and evasion of host immune responses. However, little is known about their functional roles in mycobacterial pathogenesis. Previously, we found that PPE38 is associated with the virulence of mycobacteria, presumably by modulating the host immune response. To clarify the link between PPE38 and host response, we employed a subcellular, amino acid-coded mass tagging (AACT)/SILAC-based quantitative proteomic approach to determine the proteome changes during host response to M. marinum PPE38. As a result, 291 or 290 proteins were found respectively to be up- or down-regulated in the nucleus. Meanwhile, 576 upregulated and 272 downregulated proteins were respectively detected in the cytosol. The data of quantitative proteomic changes and concurrent biological validations revealed that M. marinum PPE38 could trigger extensive inflammatory responses in macrophages, probably through interacting with toll-like receptor 2 (TLR2). We also found that PPE38 may arrest MHC-1 processing and presentation in infected macrophages. Using bioinformatics tools to analyze global changes in the host proteome, we obtained a PPE38-respondor network involved in various transcriptional factors (TFs) and TF-associated proteins. The results of our systems investigation now indicate that there is cross-talk involving a broad range of diverse biological pathways/processes that coordinate the host response to M. marinum PPE38

PPE38 of Mycobacterium marinum Triggers the Cross-Talk of Multiple Pathways Involved in the Host Response, As Revealed by Subcellular Quantitative Proteomics

The PE/PPE family of proteins which are in high abundance in pathogenic species such as Mycobacterium tuberculosis and M. marinum, play the critical role in generating antigenic variation and evasion of host immune responses. However, little is known about their functional roles in mycobacterial pathogenesis. Previously, we found that PPE38 is associated with the virulence of mycobacteria, presumably by modulating the host immune response. To clarify the link between PPE38 and host response, we employed a subcellular, amino acid-coded mass tagging (AACT)/SILAC-based quantitative proteomic approach to determine the proteome changes during host response to M. marinum PPE38. As a result, 291 or 290 proteins were found respectively to be up- or down-regulated in the nucleus. Meanwhile, 576 upregulated and 272 downregulated proteins were respectively detected in the cytosol. The data of quantitative proteomic changes and concurrent biological validations revealed that M. marinum PPE38 could trigger extensive inflammatory responses in macrophages, probably through interacting with toll-like receptor 2 (TLR2). We also found that PPE38 may arrest MHC-1 processing and presentation in infected macrophages. Using bioinformatics tools to analyze global changes in the host proteome, we obtained a PPE38-respondor network involved in various transcriptional factors (TFs) and TF-associated proteins. The results of our systems investigation now indicate that there is cross-talk involving a broad range of diverse biological pathways/processes that coordinate the host response to M. marinum PPE38

Enhancing Multiphoton Upconversion from NaYF₄:Yb/Tm@NaYF₄ Core–Shell Nanoparticles <i>via</i> the Use of Laser Cavity

We discover that emission efficiency of Tm³⁺-doped upconversion nanoparticles can be enhanced through the use of a laser cavity. With suitable control of the lasing conditions, the population of the intermediate excited states of the Tm³⁺ can be clamped at a required value above the excitation threshold. As a result, upconversion efficiency for the 300–620 nm emission band of the Tm³⁺-doped nanoparticles under 976 nm excitation can be enhanced by an order of magnitude over the case without a laser cavity. This is because the intrinsic recombination process of the intermediate excited states is suppressed and the surplus of excitation power directly contributes to the enhancement of multiphoton upconversion. Furthermore, our theoretical investigation has shown that the improvement of upconversion emission efficiency is mainly dependent on the cavity loss, so that this strategy can also be extended to other lanthanide-doped systems

14-3-3ε Mediates the Cell Fate Decision-Making Pathways in Response of Hepatocellular Carcinoma to Bleomycin-Induced DNA Damage

<div><p>Background</p><p>Lack of understanding of the response of hepatocellular carcinoma (HCC) to anticancer drugs causes the high mortality of HCC patients. Bleomycin (BLM) that induces DNA damage is clinically used for cancer therapy, while the mechanism underlying BLM-induced DNA damage response (DDR) in HCC cells remains ambiguous. Given that 14-3-3 proteins are broadly involved in regulation of diverse biological processes (BPs)/pathways, we investigate how a 14-3-3 isoform coordinates particular BPs/pathways in BLM-induced DDR in HCC.</p> <p>Methodology/Principal Findings</p><p>Using dual-tagging quantitative proteomic approach, we dissected the 14-3-3ε interactome formed during BLM-induced DDR, which revealed that 14-3-3ε <i>via</i> its associations with multiple pathway-specific proteins coordinates multiple pathways including chromosome remodeling, DNA/RNA binding/processing, DNA repair, protein ubiquitination/degradation, cell cycle arrest, signal transduction and apoptosis. Further, “zoom-in” investigation of the 14-3-3ε interacting network indicated that the BLM-induced interaction between 14-3-3ε and a MAP kinase TAK1 plays a critical role in determining cell propensity of apoptosis. Functional characterization of this interaction further revealed that BLM triggers site-specific phosphorylations in the kinase domain of TAK1. These BLM-induced changes of phosphorylations directly correlate to the strength of the TAK1 binding to 14-3-3ε, which govern the phosphorylation-dependent TAK1 activation. The enhanced 14-3-3ε-TAK1 association then inhibits the anti-apoptotic activity of TAK1, which ultimately promotes BLM-induced apoptosis in HCC cells. In a data-dependent manner, we then derived a mechanistic model where 14-3-3ε plays the pivotal role in integrating diverse biological pathways for cellular DDR to BLM in HCC.</p> <p>Conclusions</p><p>Our data demonstrated on a systems view that 14-3-3ε coordinates multiple biological pathways involved in BLM-induced DDR in HCC cells. Specifically, 14-3-3ε associates with TAK1 in a phosphorylation-dependent manner to determine the cell fate of BLM-treated HCC cells. Not only individual proteins but also those critical links in the network could be the potential targets for BLM-mediated therapeutic intervention of HCC.</p> </div

Flight of a Cytidine Deaminase Complex with an Imperfect Transition State Analogue Inhibitor: Mass Spectrometric Evidence for the Presence of a Trapped Water Molecule

Cytidine deaminase (CDA) binds the inhibitor zebularine as its 3,4-hydrate (<i>K</i>_d ∼ 10^–12 M), capturing all but ∼5.6 kcal/mol of the free energy of binding expected of an ideal transition state analogue (<i>K</i>_tx ∼ 10^–16 M). On the basis of its entropic origin, that shortfall was tentatively ascribed to the trapping of a water molecule in the enzyme–inhibitor complex, as had been observed earlier for product uridine [Snider, M. J., and Wolfenden, R. (2001) <i>Biochemistry 40</i>, 11364–11371]. Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) of CDA nebularized in the presence of saturating 5-fluorozebularine reveals peaks corresponding to the masses of E₂Zn₂W₂ (dimeric Zn-CDA with two water molecules), E₂Zn₂W₂Fz, and E₂Zn₂W₂Fz₂, where Fz represents the 3,4-hydrate of 5-fluorozebularine. In the absence of an inhibitor, E₂Zn₂ is the only dimeric species detected, with no additional water molecules. Experiments conducted in H₂¹⁸O indicate that the added mass W represents a trapped water molecule rather than an isobaric ammonium ion. This appears to represent the first identification of an enzyme-bound water molecule at a subunit interface (active site) using FTICR-MS. The presence of a 5-fluoro group appears to retard the decomposition of the inhibitory complex kinetically in the vapor phase, as no additional dimeric complexes (other than E₂Zn₂) are observed when zebularine is used in place of 5-fluorozebularine. Substrate competition assays show that in solution zebularine is released from CDA (<i>k</i>_off > 0.14 s^–1) much more rapidly than is 5-fluorozebularine (<i>k</i>_off = 0.014 s^–1), despite the greater thermodynamic stability of the zebularine complex

Exceptional Resilience of Small-Scale Au₃₀Cu₂₅Zn₄₅ under Cyclic Stress-Induced Phase Transformation

Shape memory alloys that produce and recover from large deformation driven by martensitic transformation are widely exploited in biomedical devices and microactuators. Generally their actuation work degrades significantly within first a few cycles and is reduced at smaller dimensions. Further, alloys exhibiting unprecedented reversibility have relatively small superelastic strain, 0.7%. These raise the questions of whether high reversibility is necessarily accompanied by small work and strain and whether high work and strain is necessarily diminished at small scale. Here we conclusively demonstrate that these are not true by showing that Au₃₀Cu₂₅Zn₄₅ pillars exhibit 12 MJ m^–3 work and 3.5% superelastic strain even after 100 000 phase transformation cycles. Our findings confirm that the lattice compatibility dominates the mechanical behavior of phase-changing materials at nano to micron scales and points a way for smart microactuators design having the mutual benefits of high actuation work and long lifetime

Analysis of the Proteins Secreted from the <i>Oryza meyeriana</i> Suspension-Cultured Cells Induced by <i>Xanthomonas oryzae</i> pv. <i>oryzae</i>

<div><p><i>Oryza meyeriana</i>, a wild species of rice from China, shows high resistance to <i>Xanthomonas oryzae</i> pv. <i>oryzae</i> (Xoo), the cause of rice bacterial blight, one of the most serious rice pathogens. To better understand the resistance mechanism, a proteomic study was conducted to identify changes in the proteins secreted in embryo cell suspension cultures in response to Xoo. After two-dimensional difference gel electrophoresis (2D-DIGE), 72 differentially expressed protein spots corresponding to 34 proteins were identified by Matrix-Assisted Laser Desorption/ Ionization Time of Flight Mass Spectrometry. Of the 34 proteins, 10 were up regulated and 24 down regulated. The secreted proteins identified were predicted to be involved in various biological processes, including signal transduction, defense, ROS and cell wall modification. 77% of the 34 proteins were predicted to have a signal peptide by Signal P. Quantitative Real-Time PCR showed that transcript levels of 14 secreted proteins were not well correlated with secreted protein levels. Peroxidase activity was up regulated in both <i>O</i>. <i>meyriana</i> and susceptible rice but was about three times higher in <i>O</i>. <i>meyeriana</i>. This suggests that peroxidases may play an important role in the early response to Xoo in <i>O</i>. <i>meyeriana</i>. These results not only provide a better understanding of the resistance mechanism of <i>O</i>. <i>meyeriana</i>, but have implications for studies of the interactions between other plants and their pathogens.</p></div