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Neutrophils release neutrophil extracellular traps (NETs) which ensnare pathogens and have pathogenic functions in diverse diseases. We examined the NETosis pathways induced by five stimuli; PMA, the calcium ionophore A23187, nigericin, Candida albicans and Group B Streptococcus. We studied NET production in neutrophils from healthy donors with inhibitors of molecules crucial to PMA-induced NETs including protein kinase C, calcium, reactive oxygen species, the enzymes myeloperoxidase (MPO) and neutrophil elastase. Additionally, neutrophils from chronic granulomatous disease patients, carrying mutations in the NADPH oxidase complex or a MPO-deficient patient were examined. We show that PMA, C. albicans and GBS use a related pathway for NET induction, whereas ionophores require an alternative pathway but that NETs produced by all stimuli are proteolytically active, kill bacteria and composed mainly of chromosomal DNA. Thus, we demonstrate that NETosis occurs through several signalling mechanisms, suggesting that extrusion of NETs is important in host defence. The immune system protects the body against microorganisms that can cause infections and diseases. Neutrophils are a type of immune cell that patrol the blood in search of germs. Once they encounter potentially harmful microbes, neutrophils eradicate them in different ways. One way to catch the germs is by using ‘neutrophil extracellular traps’, or NETs for short, to confine and kill the invaders. NETs are web-like structures made up of anti-microbial proteins and the neutrophil’s own DNA. The process of making NETs kills the neutrophil, as it eventually explodes to release the NETs. NETs play a key role in disease prevention, but producing too many NETs or producing them at the wrong time or in the wrong place can actually make certain diseases worse. Therefore, it is important to fully understand the signaling pathways and molecules the neutrophils use to make NETs. Kenny et al. exposed neutrophils from healthy people to five different compounds known to cause the cells to make NETs, including some harmful molecules, a fungus and a bacterium. Microscopy was then used to count how many neutrophils made NETs in response to each of the five stimuli. Further experiments showed that neutrophils from patients with an immune system disorder produced fewer NETs when stimulated with some of the compounds, while the other stimuli caused neutrophils to produce the same levels of NETs as healthy individuals. Kenny et al. also revealed that neutrophils use several different ways to produce and release NETs, depending on the stimulus used. Some of the ways required reactive oxygen species, such as hydrogen peroxide and enzymes, while others produced NETs without the need for these molecules. Lastly, Kenny et al. showed that the way the cells die after the NET is released is unique from other pathways that are known to kill cells. Future work will aim to identify a single molecule that can block neutrophils from releasing NETs at the wrong time and place, without affecting the important role NETs play in fighting germs. Such a molecule could be developed into a drug for people with diseases like lupus or rheumatoid arthritis, where the release of NETs makes the disease worse not better.

Citrullination is a post-translational modification (PTM) of arginine that is crucial for several physiological processes, including gene regulation and neutrophil extracellular trap formation. Despite recent advances, studies of protein citrullination remain challenging due to the difficulty of accessing proteins homogeneously citrullinated at a specific site. Herein, we report a technology that enables the site-specific incorporation of citrulline (Cit) into proteins in mammalian cells. This approach exploits an engineered E. coli -derived leucyl tRNA synthetase-tRNA pair that incorporates a photocaged-citrulline (SM60) into proteins in response to a nonsense codon. Subsequently, SM60 is readily converted to Cit with light in vitro and in living cells. To demonstrate the utility of the method, we biochemically characterize the effect of incorporating Cit at two known autocitrullination sites in Protein Arginine Deiminase 4 (PAD4, R372 and R374) and show that the R372Cit and R374Cit mutants are 181- and 9-fold less active than the wild-type enzyme. This technology possesses the potential to decipher the biology of citrullination. Citrullination of arginine is crucial for several physiological processes. Here the authors report the site-specific incorporation of citrulline into proteins in mammalian cells using an engineered tRNA synthetase/tRNA pair and a photocaged-citrulline.

[...]different phenomena are studied as a separate system, the hydrodynamics of one component ignores the other, spectral properties ignore timing properties or vice versa, and there is not enough instrumental resolution to decipher small scale physics. The study in the spectrotemporal domain also needs proper understanding of spectral state change, quasi periodic oscillation (QPO) frequency evolution, hardness intensity diagram, and line emissions. [...]it is a good time to take a break, recapitulate what has been carried out, what is ongoing, and what can be carried out in the future with upcoming missions, both from a theoretical and observational perspective, bringing together experts across the electromagnetic spectrum to gain new insights into the physical process of accretion. [...]the physical picture of luminous active galactic nuclei (AGN) with powerful jets has been several decades-old problems in high-energy astrophysics. In their high activity state jet is the dominating contributor to the observed features, however, the disk can also contribute when the jet activity is low or moderate ref. [...]distinguishing the central engine responsible for the variabilities in these objects is one of the front line topics.

Citrullination is the conversion of arginine-to-citrulline by protein arginine deiminases (PADs), whose dysregulation is implicated in the pathogenesis of various types of cancers and autoimmune diseases. Consistent with the ability of human cytomegalovirus (HCMV) to induce post-translational modifications of cellular proteins to gain a survival advantage, we show that HCMV infection of primary human fibroblasts triggers PAD-mediated citrullination of several host proteins, and that this activity promotes viral fitness. Citrullinome analysis reveals significant changes in deimination levels of both cellular and viral proteins, with interferon (IFN)-inducible protein IFIT1 being among the most heavily deiminated one. As genetic depletion of IFIT1 strongly enhances HCMV growth, and in vitro IFIT1 citrullination impairs its ability to bind to 5’-ppp-RNA, we propose that viral-induced IFIT1 citrullination is a mechanism of HCMV evasion from host antiviral resistance. Overall, our findings point to a crucial role of citrullination in subverting cellular responses to viral infection. Citrullination is a posttranslational modification of arginines. Here, the authors show that HCMV infection increases citrullination of host and virus proteins to promote infection and that citrullinated interferon-inducible protein IFIT1 is impaired in RNA binding, as a potential mechanism of evasion.

The echo signal amplitude received by radar is very low, necessitating signal amplification for effective detection and processing. A Low Noise Amplifier (LNA) has been developed to boost signal amplitude without adding extra noise. This paper presents innovative techniques such as gm-boosting, RC coupling, an L-type matching network, and common source inductive degeneration. These techniques achieve a wide bandwidth of 1.3 GHz (4.7 to 6 GHz) and a low undesired signal level of 0.8 dB. The bias network enhances transconductance, resulting in a gain of 36-40 dB, while the RC coupling network ensures stability from DC to 36 GHz. The LNA achieves an output 1 dB compression point of 9 dBm, an OIP3 of 23.91 dBm, a sensitivity of -137.8 dBm, a linear dynamic range of 108.8 dB, and a spurious free dynamic range of 87.87 dB. This LNA operates well in RF interference environments and demonstrates strong detection capability and linearity. Built on GaAs technology, the MMIC circuit has a compact footprint of 3.5052 mm². This efficient LNA boosts the tracking range from 128 km to 243 km when integrated into larger systems.

A bstract A detailed analysis of the systematic uncertainties in the calculation of the isovector momentum fraction, 〈 x 〉 u  −  d , helicity moment, 〈 x 〉 Δ u  − Δ d , and the transversity moment, 〈 x 〉 δu  −  δd , of the nucleon is presented using high-statistics data on seven ensembles of gauge configurations generated by the JLab/W&M/LANL/MIT collaborations using 2 + 1-flavors of dynamical Wilson-clover quarks. The much higher statistics have facilitated better control over all systematics compared to previous lattice calculations. The least understood systematic — excited-state contamination — is quantified by studying the variation of the results as a function of different estimates of the mass gap of the first excited state, obtained from two- and three-point correlation functions, and as a function of the pion mass M π . The final results are obtained using a simultaneous fit in the lattice spacing a , pion mass M π and the finite volume parameter M π L keeping leading order corrections. The data show no significant dependence on the lattice spacing and some evidence for finite-volume corrections. Our final results, in the MS ¯ scheme at 2 GeV, are 〈 x 〉 u  −  d = 0 . 155(17)(20), 〈 x 〉 Δ u  − Δ d = 0 . 183(14)(20) and 〈 x 〉 δu  −  δd = 0 . 220(18)(20), where the first error is the overall analysis uncertainty assuming excited-state contributions have been removed, and the second is an additional systematic uncertainty due to possible residual excited-state contributions. These results are consistent with phenomenological global fit values.

A bstract We compute the renormalized running coupling of SU(3) gauge theory coupled to N f = 8 flavors of massless fundamental Dirac fermions. The recently proposed finite volume gradient flow scheme is used. The calculations are performed at several lattice spacings allowing for a controlled continuum extrapolation. The results for the discrete β -function show that it is monotonic without any sign of a fixed point in the range of couplings we cover. As a cross check the continuum results are compared with the well-known perturbative continuum β-function for small values of the renormalized coupling and perfect agreement is found.

AbstractEndotoxemia induced by lipopolysaccharide (LPS) is an extremely severe syndrome identified by global activation of inflammatory responses. Neutrophil extracellular traps (NETs) play an important role in the development of endotoxemia. Histone hypercitrullination catalyzed by peptidylarginine deiminases (PADs) is a key step of NET formation. We have previously demonstrated that simultaneous inhibition of PAD2 and PAD4 with pan-PAD inhibitors can decrease NETosis and improve survival in a mouse model of LPS-induced endotoxic shock. However, the effects of PAD2 specific inhibition during NETosis and endotoxic shock are poorly understood. Therefore, in the present study, we aimed to investigate the effect of the specific PAD2 or PAD4 inhibitor on LPS-induced endotoxic shock in mice. We found that PAD2 inhibition but not PAD4 inhibition improves survival. Also, the levels of proinflammatory cytokines and NETosis were significantly reduced by PAD2 inhibitor. To our knowledge, this study demonstrates for the first time that PAD2 inhibition can reduce NETosis, decrease inflammatory cytokine production, and protect against endotoxin-induced lethality. Our findings provided a novel therapeutic strategy for the treatment of endotoxic shock.

Citrullination, the conversion of peptidyl-arginine into peptidyl-citrulline, is involved in the breakage of self-tolerance in anti-CCP-positive rheumatoid arthritis. This reaction is catalyzed by peptidyl arginine deiminases (PADs), of which PAD2 and PAD4 are thought to play key pathogenic roles. Small-molecule PAD inhibitors such as the pan-PAD inhibitor BB-Cl-amidine, the PAD2-specific inhibitor AFM-30a, and the PAD4-specific inhibitor GSK199 hold therapeutic potential and are useful tools in studies of citrullination. Using an ELISA based on the citrullination of fibrinogen, we found that AFM-30a inhibited the catalytic activity of PADs derived from live PMNs or lysed PBMCs and PMNs and of PADs in cell-free synovial fluid samples from RA patients, while GSK199 had minor effects. In combination, AFM-30a and GSK199 inhibited total intracellular citrullination and citrullination of histone H3 in PBMCs, as determined by Western blotting. They were essentially nontoxic to CD4 + T cells, CD8 + T cells, B cells, NK cells, and monocytes at concentrations ranging from 1 to 20 μM, while BB-Cl-amidine was cytotoxic at concentrations above 1 μM, as assessed by flow cytometric viability staining and by measurement of lactate dehydrogenase released from dying cells. In conclusion, AFM-30a is an efficient inhibitor of PAD2 derived from PBMCs, PMNs, or synovial fluid. AFM-30a and GSK199 can be used in combination for inhibition of PAD activity associated with PBMCs but without the cytotoxic effect of BB-Cl-amidine. This suggests that AFM-30a and GSK199 may have fewer off-target effects than BB-Cl-amidine and therefore hold greater therapeutic potential.

We present a detailed spectral study of the narrow-line Seyfert 1 galaxy, Markarian 335, using eight epoch observations made between 2013 and 2020 with the Nuclear Spectroscopic Telescope Array. The source was variable during this period both in spectral flux and flow geometry. We estimated the height of the Compton cloud from the model fitted parameters for the whole observation period. This allowed us to investigate the underlying physical processes that drive the variability in X-rays. Our model fitted mass varies in a narrow range, between (2.44±0.45−3.04±0.56)×107M⊙, however, given the large error bars, it is consistent with being constant and is in agreement with that known from optical reverberation mapping observations. The disk mass accretion rate reached a maximum of 10% of the Eddington rate during June 2013. Our study sheds light on mass outflows from the system and also compares different aspects of accretion with X-ray binaries.