Soluble TNF-like cytokine (TL1A) production by immune complexes stimulated monocytes in rheumatoid arthritis

TNF-like cytokine (TL1A) is a newly identified member of the TNF superfamily of ligands that is important for T cell costimulation and Th1 polarization. However, despite increasing information about its functions, very little is known about expression of TL1A in normal or pathological states. In this study, we report that mononuclear phagocytes appear to be a major source of TL1A in rheumatoid arthritis (RA), as revealed by their strong TL1A expression in either synovial fluids or synovial tissue of rheumatoid factor (RF)-seropositive RA patients, but not RF-/RA patients. Accordingly, in vitro experiments revealed that human monocytes express and release significant amounts of soluble TL1A when stimulated with insoluble immune complexes (IC), polyethylene glycol precipitates from the serum of RF+/RA patients, or with insoluble ICs purified from RA synovial fluids. Monocyte-derived soluble TL1A was biologically active as determined by its capacity to induce apoptosis of the human erythroleukemic cell line TF-1, as well as to cooperate with IL-12 and IL-18 in inducing the production of IFN-gamma by CD4(+) T cells. Because RA is a chronic inflammatory disease with autoimmune etiology, in which ICs, autoantibodies (including RF), and various cytokines contribute to its pathology, our data suggest that TL1A could be involved in its pathogenesis and contribute to the severity of RA disease that is typical of RF+/RA patients.     

Mechanism of the formation of DNA-protein cross-links by antitumor cisplatin

DNA–protein cross-links are formed by various DNA-damaging agents including antitumor platinum drugs. The natures of these ternary DNA–Pt–protein complexes (DPCLs) can be inferred, yet much remains to be learned about their structures and mechanisms of formation. We investigated the origin of these DPCLs and their cellular processing on molecular level using gel electrophoresis shift assay. We show that in cell-free media cisplatin [cis-diamminedichloridoplatinum(II)] forms DPCLs more effectively than ineffective transplatin [trans-diamminedichloridoplatinum(II)]. Mechanisms of transformation of individual types of plain DNA adducts of the platinum complexes into the DPCLs in the presence of several DNA-binding proteins have been also investigated. The DPCLs are formed by the transformation of DNA monofunctional and intrastrand cross-links of cisplatin. In contrast, interstrand cross-links of cisplatin and monofunctional adducts of transplatin are stable in presence of the proteins. The DPCLs formed by cisplatin inhibit DNA polymerization or removal of these ternary lesions from DNA by nucleotide excision repair system more effectively than plain DNA intrastrand or monofunctional adducts. Thus, the bulky DNA–protein cross-links formed by cisplatin represent a more distinct and persisting structural motif recognized by the components of downstream cellular systems processing DNA damage considerably differently than the plain DNA adducts of this metallodrug.     

A chemically induced new pea (Pisum sativum) mutant SGECdt with increased tolerance to, and accumulation of, cadmium

BACKGROUND AND AIMS: To date, there are no crop mutants described in the literature that display both Cd accumulation and tolerance. In the present study a unique pea (Pisum sativum) mutant SGECd(t) with increased Cd tolerance and accumulation was isolated and characterized. METHODS: Ethylmethane sulfonate mutagenesis of the pea line SGE was used to obtain the mutant. Screening for Cd-tolerant seedlings in the M2 generation was performed using hydroponics in the presence of 6 microm CdCl2. Hybridological analysis was used to identify the inheritance of the mutant phenotype. Several physiological and biochemical characteristics of SGECd(t) were studied in hydroponic experiments in the presence of 3 microm CdCl2, and elemental analysis was conducted. KEY RESULTS: The mutant SGECd(t) was characterized as having a monogenic inheritance and a recessive phenotype. It showed increased Cd concentrations in roots and shoots but no obvious morphological defects, demonstrating its capability to cope well with increased Cd levels in its tissues. The enhanced Cd accumulation in the mutant was accompanied by maintenance of homeostasis of shoot Ca, Mg, Zn and Mn contents, and root Ca and Mg contents. Through the application of La(+3) and the exclusion of Ca from the nutrient solution, maintenance of nutrient homeostasis in Cd-stressed SGECd(t) was shown to contribute to the increased Cd tolerance. Control plants of the mutant (i.e. no Cd treatment) had elevated concentrations of glutathione (GSH) in the roots. Through measurements of chitinase and guaiacol-dependent peroxidase activities, as well as proline and non-protein thiol (NPT) levels, it was shown that there were lower levels of Cd stress both in roots and shoots of SGECd(t). Accumulation of phytochelatins [(PCcalculated) = (NPT)-(GSH)] could be excluded as a cause of the increased Cd tolerance in the mutant. CONCLUSIONS: The SGECd(t) mutant represents a novel and unique model to study adaptation of plants to toxic heavy metal concentrations.     

Electrophysical characteristics of Azospirillum brasilense Sp245 during interaction with antibodies to various cell surface epitopes

This work was undertaken to examine the electrooptical characteristics of cells of Azospirillum brasilense Sp245 during their interaction with antibodies developed to various cell surface epitopes. We used the dependences of the cell suspension optical density changes induced by electroorientation on the orienting field frequency (740, 1000, 1450, 2000, and 2800kHz). Cell interactions with homologous strain-specific antibodies to the A. brasilense Sp245 O antigen and with homologous antibodies to whole bacterial cells brought about considerable changes in the electrooptical properties of the bacterial suspension. When genus-specific antibodies to the flagellin of the Azospirillum sheathed flagellum and antibodies to the serologically distinct O antigen of A. brasilense Sp7 were included in the A. brasilense Sp245 suspension, the changes caused in the electrooptical signal were slight and had values close to those for the above changes. These findings agree well with the immunochemical characteristics of the Azospirillum O antigens and with the data on the topographical distribution of the Azospirillum major cell surface antigens. The obtained results can serve as a basis for the development of a rapid test for the intraspecies detection of microorganisms.     

Arabidopsis PPP family of serine/threonine phosphatases

Serine/threonine-specific phosphoprotein phosphatases (PPPs) are ubiquitous enzymes in all eukaryotes, but their regulatory functions are largely unknown in higher plants. The Arabidopsis genome encodes 26 PPP catalytic subunits related to type 1, type 2A and so-called novel phosphatases, including four plant-specific enzymes carrying large N-terminal kelch-domains, but no apparent homologue of the PP2B family. The catalytic subunits of PPPs associate with regulatory protein partners that target them to well defined cellular locations and modulate their activity. Recent studies of phosphatase partners and their interactions have directed attention again to functional dissection of plant PPP families, and highlight their intriguing roles in the regulation of metabolism, cell cycle and development, as well as their roles in light, stress and hormonal signalling.     

Engineering photosynthetic light capture: impacts on improved solar energy to biomass conversion

The main function of the photosynthetic process is to capture solar energy and to store it in the form of chemical 'fuels'. Increasingly, the photosynthetic machinery is being used for the production of biofuels such as bio-ethanol, biodiesel and bio-H₂. Fuel production efficiency is directly dependent on the solar photon capture and conversion efficiency of the system. Green algae (e.g. Chlamydomonas reinhardtii ) have evolved genetic strategies to assemble large light-harvesting antenna complexes (LHC) to maximize light capture under low-light conditions, with the downside that under high solar irradiance, most of the absorbed photons are wasted as fluorescence and heat to protect against photodamage. This limits the production process efficiency of mass culture. We applied RNAi technology to down-regulate the entire LHC gene family simultaneously to reduce energy losses by fluorescence and heat. The mutant Stm3LR3 had significantly reduced levels of LHCI and LHCII mRNAs and proteins while chlorophyll and pigment synthesis was functional. The grana were markedly less tightly stacked, consistent with the role of LHCII. Stm3LR3 also exhibited reduced levels of fluorescence, a higher photosynthetic quantum yield and a reduced sensitivity to photoinhibition, resulting in an increased efficiency of cell cultivation under elevated light conditions. Collectively, these properties offer three advantages in terms of algal bioreactor efficiency under natural high-light levels: (i) reduced fluorescence and LHC-dependent heat losses and thus increased photosynthetic efficiencies under high-light conditions; (ii) improved light penetration properties; and (iii) potentially reduced risk of oxidative photodamage of PSII.     

PARP inhibition alleviates diabetes-induced systemic oxidative stress and neural tissue 4-hydroxynonenal adduct accumulation: correlation with peripheral nerve function

This study evaluated the role of poly(ADP-ribose) polymerase (PARP) in systemic oxidative stress and 4-hydoxynonenal adduct accumulation in diabetic peripheral neuropathy. Control and streptozotocin-diabetic rats were maintained with or without treatment with the PARP inhibitor, 1,5-isoquinolinediol, 3 mg kg(-1) day(-1), for 10 weeks after an initial 2 weeks. Treatment efficacy was evaluated by poly(ADP-ribosyl)ated protein content in peripheral nerve and spinal cord (Western blot analysis) and dorsal root ganglion neurons and nonneuronal cells (fluorescence immunohistochemistry), as well as by indices of peripheral nerve function. Diabetic rats displayed increased urinary isoprostane and 8-hydroxy-2'-deoxyguanosine excretion (ELISA) and 4-hydroxynonenal adduct accumulation in endothelial and Schwann cells of the peripheral nerve, neurons, astrocytes, and oligodendrocytes of the spinal cord and neurons and glial cells of the dorsal root ganglia (double-label fluorescence immunohistochemistry), as well as motor and sensory nerve conduction velocity deficits, thermal hypoalgesia, and tactile allodynia. PARP inhibition counteracted diabetes-induced systemic oxidative stress and 4-hydroxynonenal adduct accumulation in peripheral nerve and spinal cord (Western blot analysis) and dorsal root ganglion neurons (perikarya, fluorescence immunohistochemistry), which correlated with improvement of large and small nerve fiber function. The findings reveal the important role of PARP activation in systemic oxidative stress and 4-hydroxynonenal adduct accumulation in diabetic peripheral neuropathy.     

TSAR,a new graph–theoretical approach to computational modeling of protein side‐chain flexibility: Modeling of ionization properties of proteins

A new graph–theoretical approach called thermodynamic sampling of amino acid residues (TSAR) has been elaborated to explicitly account for the protein side chain flexibility in modeling conformation‐dependent protein properties. In TSAR, a protein is viewed as a graph whose nodes correspond to structurally independent groups and whose edges connect the interacting groups. Each node has its set of states describing conformation and ionization of the group, and each edge is assigned an array of pairwise interaction potentials between the adjacent groups. By treating the obtained graph as a belief‐network—a well‐established mathematical abstraction—the partition function of each node is found. In the current work we used TSAR to calculate partition functions of the ionized forms of protein residues. A simplified version of a semi‐empirical molecular mechanical scoring function, borrowed from our Lead Finder docking software, was used for energy calculations. The accuracy of the resulting model was validated on a set of 486 experimentally determined pK_a values of protein residues. The average correlation coefficient (R) between calculated and experimental pK_a values was 0.80, ranging from 0.95 (for Tyr) to 0.61 (for Lys). It appeared that the hydrogen bond interactions and the exhaustiveness of side chain sampling made the most significant contribution to the accuracy of pK_a calculations. Proteins 2011; © 2011 Wiley‐Liss, Inc.