The PSF.p54nrb complex is a novel Mnk substrate that binds the mRNA for tumor necrosis factor alpha

To identify new potential substrates for the MAP kinase signal-integrating kinases (Mnks), we employed a proteomic approach. The Mnks are targeted to the translational machinery through their interaction with the cap-binding initiation factor complex. We tested whether proteins retained on cap resin were substrates for the Mnks in vitro, and identified one such protein as PSF (the PTB (polypyrimidine tract-binding protein)-associated splicing factor). Mnks phosphorylate PSF at two sites in vitro, and our data show that PSF is an Mnk substrate in vivo. We also demonstrate that PSF, together with its partner, p54nrb, binds RNAs that contain AU-rich elements (AREs), such as those for proinflammatory cytokines (e.g. tumor necrosis factor ? (TNF?)). Indeed, PSF associates specifically with the TNF? mRNA in living cells. PSF is phosphorylated at two sites by the Mnks. Our data show that Mnk-mediated phosphorylation increases the binding of PSF to the TNF? mRNA in living cells. These findings identify a novel Mnk substrate. They also suggest that the Mnk-catalyzed phosphorylation of PSF may regulate the fate of specific mRNAs by modulating their binding to PSF·p54nrb

Evaluation of mTOR-regulated mRNA translation.

mTOR, the mammalian target of rapamycin, regulates protein synthesis (mRNA translation) by affecting the phosphorylation or activity of several translation factors. Here, we describe methods for studying the impact of mTOR signalling on protein synthesis, using inhibitors of mTOR such as rapamycin (which impairs some of its functions) or mTOR kinase inhibitors (which probably block all functions).To assess effects of mTOR inhibition on general protein synthesis in cells, the incorporation of radiolabelled amino acids into protein is measured. This does not yield information on the effects of mTOR on the synthesis of specific proteins. To do this, two methods are described. In one, stable-isotope labelled amino acids are used, and their incorporation into new proteins is determined using mass spectrometric methods. The proportions of labelled vs. unlabeled versions of each peptide from a given protein provide quantitative information about the rate of that protein's synthesis under different conditions. Actively translated mRNAs are associated with ribosomes in polyribosomes (polysomes); thus, examining which mRNAs are found in polysomes under different conditions provides information on the translation of specific mRNAs under different conditions. A method for the separation of polysomes from non-polysomal mRNAs is describe

Shockwaves in converging geometries

Plate impact experiments are a powerful tool in equation of state (EOS) development, but are inherently limited by the range of impact velocities accessible to the gun. In an effort to dramatically increase the range of pressures which can be studied with available impact velocities, a new experimental technique is being developed. The possibility of using a confined converging target to focus Shockwaves and produce a large amplitude pressure pulse is examined. When the planar shock resulting from impact enters the converging target the impedance mismatch at the boundary of the confinement produces reflected Mach waves and the subsequent wave interactions produce a diffraction cycle resulting in increases in the shock strength with each cycle. Since this configuration is limited to relatively low impedance targets, a second technique is proposed in which the target is two concentric cylinders designed such that the inner cylinder will have a lower shock velocity than the much larger shock velocity in the outer cylinder. The resulting dispersion in the wave front creates converging shocks, which will interact and eventually result in a steady Mach configuration with an increase in pressure in the Mach disk. Numerical simulations indicate a significant increase in pressure for both methods and show promise for the proposed concepts

Advances in Shock Compression of Mantle Materials and Implications

Hugoniots of lower mantle mineral compositions are sensitive to the conditions where they cross phase boundaries including both polymorphic phase transitions and partial to complete melting. For SiO_2, the Hugoniot of fused silica passes from stishovite to partial melt (73 GPa, 4600 K) whereas the Hugoniot of crystal quartz passes from CaCi_2 structure to partial melt (116 GPa, 4900 K). For Mg_2SiO_4, the forsterite Hugoniot passes from the periclase +MgSiO_3 (perovskite) assemblage to melt before 152 GPa and 4300 K, whereas the wadsleyite Hugoniot transforms first to periclase +MgSiO_3 (post-perovskite) and then melts at 151 GPa and 4160 K. Shock states achieved from crystal enstatite are molten above 160 GPa. High-pressure Grüneisen parameters for molten states of MgSiO_3 and Mg_2SiO_4 increase markedly with compression, going from 0.5 to 1.6 over the 0 to 135 GPa range. This gives rise to a very large (>2000 K) isentropic rise in temperature with depth in thermal models of a primordial deep magma ocean within the Earth. These magma ocean isentropes lead to models that have crystallization initiating at mid-lower mantle depths. Such models are consistent with the suggestion that the present ultra-low velocity zones, at the base of the lowermost mantle, represent a dynamically stable, partially molten remnant of the primordial magma ocean. The new shock melting data for silicates support a model of the primordial magma ocean that is concordant with the Berkeley-Caltech iron core model [1] for the temperature at the center of the Earth

A review of CO2 coolants for sustainable machining

In many machining operations, metalworking fluids (MWFs) play an invaluable role. Often, proper application of an intelligent MWF strategy allows manufacturing processes to benefit from a multitude of operational incentives, not least of which are increased tool life, improved surface integrity and optimised chip handling. Despite these clearly positive implications, current MWF strategies are often unable to accommodate the environmental, economic and social conscience of industrial environments. In response to these challenges, CO2 coolants are postulated as an operationally viable, environmentally benign MWF solution. Given the strong mechanistic rationale and historical evidence in support of cryogenic coolants, this review considers the technological chronology of cryogenic MWF’s in addition to the current state-of-the-art approaches. The review also focuses on the use of CO2 coolants in the context of the machining of a multitude of material types in various machining conditions. In doing so, cryogenic assisted machining is shown to offer a litany of performance benefits for both conventional emulsion (flood) cooling and near dry strategies, i.e., minimum quantity lubrication (MQL), as well as aerosol dry lubrication (ADL)

Mycobacterium tuberculosis subverts negative regulatory pathways in human macrophages to drive immunopathology.

Tuberculosis remains a global pandemic and drives lung matrix destruction to transmit. Whilst pathways driving inflammatory responses in macrophages have been relatively well described, negative regulatory pathways are less well defined. We hypothesised that Mycobacterium tuberculosis (Mtb) specifically targets negative regulatory pathways to augment immunopathology. Inhibition of signalling through the PI3K/AKT/mTORC1 pathway increased matrix metalloproteinase-1 (MMP-1) gene expression and secretion, a collagenase central to TB pathogenesis, and multiple pro-inflammatory cytokines. In patients with confirmed pulmonary TB, PI3Kδ expression was absent within granulomas. Furthermore, Mtb infection suppressed PI3Kδ gene expression in macrophages. Interestingly, inhibition of the MNK pathway, downstream of pro-inflammatory p38 and ERK MAPKs, also increased MMP-1 secretion, whilst suppressing secretion of TH1 cytokines. Cross-talk between the PI3K and MNK pathways was demonstrated at the level of eIF4E phosphorylation. Mtb globally suppressed the MMP-inhibitory pathways in macrophages, reducing levels of mRNAs encoding PI3Kδ, mTORC-1 and MNK-1 via upregulation of miRNAs. Therefore, Mtb disrupts negative regulatory pathways at multiple levels in macrophages to drive a tissue-destructive phenotype that facilitates transmission

Ribosomal stress activates eEF2K-eEF2 pathway causing translation elongation inhibition and recruitment of Terminal Oligopyrimidine (TOP) mRNAs on polysomes

The synthesis of adequate amounts of ribosomes is an essential task for the cell. It is therefore not surprising that regulatory circuits exist to organize the synthesis of ribosomal components. It has been shown that defect in ribosome biogenesis (ribosomal stress) induces apoptosis or cell cycle arrest through activation of the tumor suppressor p53. This mechanism is thought to be implicated in the pathophysiology of a group of genetic diseases such as Diamond Blackfan Anemia which are called ribosomopathies. We have identified an additional response to ribosomal stress that includes the activation of eukaryotic translation elongation factor 2 kinase with a consequent inhibition of translation elongation. This leads to a translational reprogramming in the cell that involves the structurally defined group of messengers called terminal oligopyrimidine (TOP) mRNAs which encode ribosomal proteins and translation factors. In fact, while general protein synthesis is decreased by the impairment of elongation, TOP mRNAs are recruited on polysomes causing a relative increase in the synthesis of TOP mRNA-encoded proteins compared to other proteins. Therefore, in response to ribosomal stress, there is a change in the translation pattern of the cell which may help restore a sufficient level of ribosomes

Composite nature of fresh skin revealed during compression

Biological systems are subjected to moderate-to-high strain rates in blast-type traumatic injuries. An improved understanding of the responses of cells and tissues to extreme mechanical stresses could improve mitigation and post-injury treatment strategies. A key aim of this research is to create biologically meaningful injury models of soft tissues. Here the authors examine the material and cellular properties of freshly harvested porcine skin in compression. The data presented suggest that fresh skin differentially responds low to moderate strain rates as a composite rather than that of a homogeneous polymer. The implications of this work are discussed in terms of creating improved analytical models to describe the material properties of fresh skin. The Centre for Blast Injury Studies acknowledges The Royal British Legion for its support, as well as Imperial College London. The Institute of Shock Physics acknowledges the support of the Atomic Weapon Establishment, Aldermaston, UK and Imperial College London.This is the accepted manuscript. The final version is available at http://www.icevirtuallibrary.com/content/article/10.1680/bbn.14.00028