Potent and selective chemical probes are valuable tools for discovery of novel treatments for human diseases. NF-κB-inducing kinase (NIK) is a key trigger in the development of liver injury and fibrosis. Whether inhibition of NIK activity by chemical probes ameliorates liver inflammation and injury is largely unknown. In this study, a small-molecule inhibitor of NIK, B022, was found to be a potent and selective chemical probe for liver inflammation and injury. B022 inhibited the NIK signaling pathway, including NIK-induced p100-to-p52 processing and inflammatory gene expression, both in vitro and in vivo Furthermore, in vivo administration of B022 protected against not only NIK but also CCl-induced liver inflammation and injury. Our data suggest that inhibition of NIK is a novel strategy for treatment of liver inflammation, oxidative stress, and injury.-Ren, X., Li, X., Jia, L., Chen, D., Hou, H., Rui, L., Zhao, Y., Chen, Z. A small-molecule inhibitor of NF-κB-inducing kinase (NIK) protects liver from toxin-induced inflammation, oxidative stress, and injury.
Rice cultivation has been challenged by increasing food demand and water scarcity. We examined the responses of water use, grain yield, and water productivity to various modes of field water managements in Chinese double rice systems. Four treatments were studied in a long-term field experiment (1998–2015): continuous flooding (CF), flooding—midseason drying—flooding (F-D-F), flooding—midseason drying—intermittent irrigation without obvious standing water (F-D-S), and flooding—rain-fed (F-RF). The average precipitation was 483 mm in early-rice season and 397 mm in late-rice season. The irrigated water for CF, F-D-F, F-D-S, and F-RF, respectively, was 263, 340, 279, and 170 mm in early-rice season, and 484, 528, 422, and 206 mm in late-rice season. Grain yield for CF, F-D-F, F-D-S, and F-RF, respectively, was 4,722, 4,597, 4,479, and 4,232 kgha-1 in early-rice season, and 5,420, 5,402, 5,366, and 4,498 kgha-1 in late-rice season. Compared with CF, F-D-F consumed more irrigated water, which still decreased grain yield, leading to a decrease in water productivity by 25% in early-rice season and by 8% in late-rice season. Compared with F-D-F, F-D-S saved much irrigated water with a small yield reduction, leading to an increase in water productivity by 22% in early-rice season and by 26% in late-rice season. The results indicate that CF is best for early-rice and FDS is best for late-rice in terms of grain yield and water productivity.
The cold shock domain (CSD) is an evolutionarily conserved nucleic acid binding domain that exhibits binding activity to RNA, ssDNA, and dsDNA. Mammalian CRHSP-24 contains CSD, but its structure-functional relationship has remained elusive. Here we report the crystal structure of human CRHSP-24 and characterization of the molecular trafficking of CRHSP-24 between stress granules and processing bodies in response to oxidative stress. The structure of CRHSP-24 determined by single-wavelength anomalous dispersion exhibits an ␣-helix and a compact -barrel formed by five curved anti-parallel  strands. Ligand binding activity of the CSD is orchestrated by residues Ser 41 to Leu 43 . Interestingly, a phosphomimetic S41D mutant abolishes the ssDNA binding in vitro and causes CRHSP-24 liberated from stress granules in vivo without apparent alternation of its localization to the processing bodies. This new class of phosphorylation-regulated interaction between the CSD and nucleic acids is unique in stress granule plasticity. Importantly, the association of CRHSP-24 with stress granules is blocked by PP4/ PP2A inhibitor calyculin A as PP2A catalyzes the dephosphorylation of Ser 41 of CRHSP-24. Therefore, we speculate that CRHSP-24 participates in oxidative stress response via a dynamic and temporal association between stress granules and processing bodies.
Deciphering the activity-conformation relationship of PTPase is of great interest to understand howPTPase activity is determined by its conformation. Here we studied the activity and conformational transitions of PTPase from thermus thermophilus HB27 in the presence of sodium dodecyl sulfate (SDS). Activity assays showed the inactivation of PTPase induced by SDS was in a concentration-dependent manner. Fluorescence and circular dichroism spectra suggested SDS induced significant conformational transitions of PTPase, which resulted in the inactivation of PTPase, and the changes of α-helical structure and tertiary structure of PTPase. Structural analysis revealed a number of hydrophobic and charged residues around the active sites of PTPase may be involved in the hydrophobic and ionic bonds interactions of PTPase and SDS, which are suggested to be the major driving force to result in PTPase inactivation and conformational transitions induced by SDS. Our results suggested the hydrophobic and charged residues around the active sites were essential for the activity and conformation of PTPase. Our study promotes a better understanding of the activity and conformation of PTPase.Although we have discussed the conformational transitions and the inactivation of PTPase induced by SDS in detail, more experimental evidences such as the complex structure of PTPase-SDS and the dynamics of PTPase-SDS interactions are still required to clarify the details of molecular interaction. Our study is valuable toward the long-term goal to better understand the activity and conformation of PTPase. Materials and MethodsReagents and materials. All chemicals used in this research such as para-nitrophenyl phosphate (pNPP), Isopropyl-β-D-1-thiogalactopyranoside (IPTG), Dithiothreitol (DTT), SDS and 1-anilinonaphtalene-8-sulfonate (ANS) were of the highest purity commercially available. PTPase from thermus thermophilus HB27 was cloned into pET-28a (+) vector (Novagen, Germany) and overexpressed in E. coli BL21 (DE3). PTPase was purified as described 12 . The concentration of recombinant PTPase was determined by BCA protein assay kit (Pierce, USA).PTPase activity assay. PTPase activity was assayed as described previously 25,44 . Briefly, pNPP (10 mM) and PTPase (2.4 μM) were added into 200 µL, 50 mM acetic acid-sodium acetate buffer (pH 3.8) plus 5 mM DTT. After incubation at 30 °C for 10 min, NaOH (1 M, 1 mL) was added into the mixture to terminate the reaction. The absorption change at 405 nm was recorded on a Helios-γ UV-VIS spectrophotometer (Thermo Scientific, USA).
Protein crystallization is of great importance because protein crystals have a number of different important applications, including large-scale purification of proteins, determination of protein structure, nanoparticle preparation, and theoretical studies of crystallization. An approach often used to efficiently crystallize proteins is the use of nucleants or seeds (small fragments of protein crystals) that can help increase the probability of protein crystallization. Due to the very positive effect that seeding has on protein crystallization, seeds are now widely accepted and utilized in practical protein crystallization. Here, we show that cross-linked protein crystals (CLPCs), which retain the crystal structure but are much more stable than noncross-linked crystals, can also be used as a new type of seed for promoting protein crystallization. Seeding with CLPCs has effects on both the reproducibility and screening of protein crystals and could improve the optical perfection (well-defined facets) of protein crystals and the probability of obtaining protein crystals. In addition, the cross-linked protein crystals may reduce the concentration of protein molecules needed to obtain protein crystals. Furthermore, CLPCs are very stable in air, and no protective medium is necessary for the long-term storage of CLPCs. This feature makes the CLPC seeding method a potentially powerful technique in practical protein crystallization on either a laboratorial or an industrial scale.
During the last two decades, C-type lectin receptors (CLRs) have been demonstrated to play key roles in initiating the host immune response against fungal infection. It is well established that CLRs, such as Dectin-1, Dectin-2, Dectin-3 and Mincle recognize the cell wall component from the infected microorganisms by using their carbohydrate recognition domain (CRD). Upon stimulation, CLRs induce multiple signal transduction cascades through their own immunereceptor tyrosine-based activation motifs (ITAMs) or interacting with ITAM-containing adaptor proteins such as FcRγ, which then lead to the activation of nuclear factor kappa B (NF-κB) through Syk- and CARD9-dependent pathway. Dissecting CLR signal cascades and their effects on host immune cells is essential to understand the molecular mechanisms in regulating host antifungal immunity. Recently, the activated CLRs including Dectin-1 and Dectin-2 are reported to undergo lysome-mediated degradation by an E3 ubiquitin ligase CBL-b. Moreover, structural analysis will help understand the molecular mechanism of these CLRs and provide clues to rational design for effective anti-fungal drugs. Overall, we summarize the current knowledge on activating and inhibitory CLRs and discuss how to boost host immune system to fight against invasive fungal infection.
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