Stepwise phosphorylation of p65 promotes NF-kappa B activation and NK cell responses during target cell recognition

NF-κB is a key transcription factor that dictates the outcome of diverse immune responses. How NF-κB is regulated by multiple activating receptors that are engaged during natural killer (NK)-target cell contact remains undefined. Here we show that sole engagement of NKG2D, 2B4 or DNAM-1 is insufficient for NF-κB activation. Rather, cooperation between these receptors is required at the level of Vav1 for synergistic NF-κB activation. Vav1-dependent synergistic signalling requires a separate PI3K-Akt signal, primarily mediated by NKG2D or DNAM-1, for optimal p65 phosphorylation and NF-κB activation. Vav1 controls downstream p65 phosphorylation and NF-κB activation. Synergistic signalling is defective in X-linked lymphoproliferative disease (XLP1) NK cells entailing 2B4 dysfunction and required for p65 phosphorylation by PI3K-Akt signal, suggesting stepwise signalling checkpoint for NF-κB activation. Thus, our study provides a framework explaining how signals from different activating receptors are coordinated to determine specificity and magnitude of NF-κB activation and NK cell responses

Stimulation of Na⁺/H⁺ Exchanger Isoform 1 Promotes Microglial Migration

Regulation of microglial migration is not well understood. In this study, we proposed that Na+/H+ exchanger isoform 1 (NHE-1) is important in microglial migration. NHE-1 protein was co-localized with cytoskeletal protein ezrin in lamellipodia of microglia and maintained its more alkaline intracellular pH (pHi). Chemoattractant bradykinin (BK) stimulated microglial migration by increasing lamellipodial area and protrusion rate, but reducing lamellipodial persistence time. Interestingly, blocking NHE-1 activity with its potent inhibitor HOE 642 not only acidified microglia, abolished the BK-triggered dynamic changes of lamellipodia, but also reduced microglial motility and microchemotaxis in response to BK. In addition, NHE-1 activation resulted in intracellular Na+ loading as well as intracellular Ca2+ elevation mediated by stimulating reverse mode operation of Na+/Ca2+ exchange (NCXrev). Taken together, our study shows that NHE-1 protein is abundantly expressed in microglial lamellipodia and maintains alkaline pHi in response to BK stimulation. In addition, NHE-1 and NCXrev play a concerted role in BK-induced microglial migration via Na+ and Ca2+ signaling. © 2013 Shi et al

Facile Synthesis of Amine-Functionalized Eu3+-Doped La(OH)3 Nanophosphors for Bioimaging

Here, we report a straightforward synthesis process to produce colloidal Eu3+-activated nanophosphors (NPs) for use as bioimaging probes. In this procedure, poly(ethylene glycol) serves as a high-boiling point solvent allowing for nanoscale particle formation as well as a convenient medium for solvent exchange and subsequent surface modification. The La(OH)3:Eu3+ NPs produced by this process were ~3.5 nm in diameter as determined by transmission electron microscopy. The NP surface was coated with aminopropyltriethoxysilane to provide chemical functionality for attachment of biological ligands, improve chemical stability and prevent surface quenching of luminescent centers. Photoluminescence spectroscopy of the NPs displayed emission peaks at 597 and 615 nm (λex = 280 nm). The red emission, due to 5D0 → 7F1 and 5D0 → 7F2 transitions, was linear with concentration as observed by imaging with a conventional bioimaging system. To demonstrate the feasibility of these NPs to serve as optical probes in biological applications, an in vitro experiment was performed with HeLa cells. NP emission was observed in the cells by fluorescence microscopy. In addition, the NPs displayed no cytotoxicity over the course of a 48-h MTT cell viability assay. These results suggest that La(OH)3:Eu3+ NPs possess the potential to serve as a luminescent bioimaging probe

Top-up operation at Pohang Light Source-II

After three years of upgrading work, PLS-II (S. Shin, Commissioning of the PLS-II, JINST, January 2013) is now successfully operating. The top-up operation of the 3 GeV linear accelerator had to be delayed because of some challenges encountered, and PLS-II was run in decay mode at the beginning in March 2012. The main difficulties encountered in the top-up operation of PLS-II are different levels between the linear accelerator and the storage ring, the 14 narrow gap in-vacuum undulators in operation, and the full energy injection by 3 GeV linear accelerator. Large vertical emittance and energy jitter of the linac were the major obstacles that called for careful control of injected beam to reduce beam loss in the storage ring during injection. The following measures were taken to resolve these problems: (1) The high resolution Libera BPM (see http://www.i-tech.si) was implemented to measure the beam trajectory and energy. (2) Three slit systems were installed to filter the beam edge. (3) De-Qing circuit was applied to the modulator system to improve the energy stability of injected beam. As a result, the radiation by beam loss during injection is reduced drastically, and the top-up mode has been successfully operating since 19th March 2013. In this paper, we describe the experimental results of the PLS-II top-up operation and the improvement plan. (C) 2014 AIP Publishing LLC.open111011sciescopu

TRAIL promotes caspase-dependent pro-inflammatory responses via PKCδ activation by vascular smooth muscle cells

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is best known for its selective cytotoxicity against transformed tumor cells. Most non-transformed primary cells and several cancer cell lines are not only resistant to death receptor-induced apoptosis, but also subject to inflammatory responses in a nuclear factor-κB (NF-κB)-dependent manner. Although the involvement of TRAIL in a variety of vascular disorders has been proposed, the exact molecular mechanisms are unclear. Here, we aimed to delineate the role of TRAIL in inflammatory vascular response. We also sought possible molecular mechanisms to identify potential targets for the prevention and treatment of post-angioplastic restenosis and atherosclerosis. Treatment with TRAIL increased the expression of intercellular adhesion molecule-1 by primary human vascular smooth muscle cells via protein kinase C (PKC)δ and NF-κB activation. Following detailed analysis using various PKCδ mutants, we determined that PKCδ activation was mediated by caspase-dependent proteolysis. The protective role of PKCδ was further confirmed in post-traumatic vascular remodeling in vivo. We propose that the TRAIL/TRAIL receptor system has a critical role in the pathogenesis of inflammatory vascular disorders by transducing pro-inflammatory signals via caspase-mediated PKCδ cleavage and subsequent NF-κB activation

Quality control and beam test of GEM detectors for future upgrades of the CMS muon high rate region at the LHC

Gas Electron Multipliers (GEM) are a proven position sensitive gas detector technology which nowadays is becoming more widely used in High Energy Physics. GEMs offer an excellent spatial resolution and a high particle rate capability, with a close to 100% detection efficiency. In view of the high luminosity phase of the CERN Large Hadron Collider, these aforementioned features make GEMs suitable candidates for the future upgrades of the Compact Muon Solenoid (CMS) detector. In particular, the CMS GEM Collaboration proposes to cover the high-eta region of the muon system with large-area triple-GEM detectors, which have the ability to provide robust and redundant tracking and triggering functions. In this contribution, after a general introduction and overview of the project, the construction of full-size trapezoidal triple-GEM prototypes will be described in more detail. The procedures for the quality control of the GEM foils, including gain uniformity measurements with an x-ray source will be presented. In the past few years, several CMS triple-GEM prototype detectors were operated with test beams at the CERN SPS. The results of these test beam campaigns will be summarised

The Cell Shape-determining Csd6 Protein from Helicobacter pylori Constitutes a New Family of L,D-Carboxypeptidase

Helicobacter pylori causes gastrointestinal diseases, including gastric cancer. Its high motility in the viscous gastric mucosa facilitates colonization of the human stomach and depends on the helical cell shape and the flagella. In H. pylori, Csd6 is one of the cell shape-determining proteins that play key roles in alteration of cross-linking or by trimming of peptidoglycan muropeptides. Csd6 is also involved in deglycosylation of the flagellar protein FlaA. To better understand its function, biochemical, biophysical, and structural characterizations were carried out. We show that Csd6 has a three-domain architecture and exists as a dimer in solution. The N-terminal domain plays a key role in dimerization. The middle catalytic domain resembles those of L,D-transpeptidases, but its pocket-shaped active site is uniquely defined by the four loops I to IV, among which loops I and III show the most distinct variations from the known L,D-transpeptidases. Mass analyses confirm that Csd6 functions only as an L,D-carboxypeptidase and not as an L,D-transpeptidase. The D-Ala-complexed structure suggests possible binding modes of both the substrate and product to the catalytic domain. The C-terminal nuclear transport factor 2-like domain possesses a deep pocket for possible binding of pseudaminic acid, and in silico docking supports its role in deglycosylation of flagellin. On the basis of these findings, it is proposed that H. pylori Csd6 and its homologs constitute a new family of L,D-carboxypeptidase. This work provides insights into the function of Csd6 in regulating the helical cell shape and motility of H. pylori.1165Ysciescopu

Web-based monitoring tools for Resistive Plate Chambers in the CMS experiment at CERN

The Resistive Plate Chambers (RPC) are used in the CMS experiment at the trigger level and also in the standard offline muon reconstruction. In order to guarantee the quality of the data collected and to monitor online the detector performance, a set of tools has been developed in CMS which is heavily used in the RPC system. The Web-based monitoring (WBM) is a set of java servlets that allows users to check the performance of the hardware during data taking, providing distributions and history plots of all the parameters. The functionalities of the RPC WBM monitoring tools are presented along with studies of the detector performance as a function of growing luminosity and environmental conditions that are tracked over time

Radiation background with the CMS RPCs at the LHC

The Resistive Plate Chambers (RPCs) are employed in the CMS Experiment at the LHC as dedicated trigger system both in the barrel and in the endcap. This article presents results of the radiation background measurements performed with the 2011 and 2012 proton-proton collision data collected by CMS. Emphasis is given to the measurements of the background distribution inside the RPCs. The expected background rates during the future running of the LHC are estimated both from extrapolated measurements and from simulation