Heat stress: an overview of molecular responses in photosynthesis

The primary targets of thermal damage in plants are the oxygen evolving complex along with the associated cofactors in photosystem II (PSII), carbon fixation by Rubisco and the ATP generating system. Recent investigations on the combined action of moderate light intensity and heat stress suggest that moderately high temperatures do not cause serious PSII damage but inhibit the repair of PSII. The latter largely involves de novo synthesis of proteins, particularly the D1 protein of the photosynthetic machinery that is damaged due to generation of reactive oxygen species (ROS), resulting in the reduction of carbon fixation and oxygen evolution, as well as disruption of the linear electron flow. The attack of ROS during moderate heat stress principally affects the repair system of PSII, but not directly the PSII reaction center (RC). Heat stress additionally induces cleavage and aggregation of RC proteins; the mechanisms of such processes are as yet unclear. On the other hand, membrane linked sensors seem to trigger the accumulation of compatible solutes like glycinebetaine in the neighborhood of PSII membranes. They also induce the expression of stress proteins that alleviate the ROS-mediated inhibition of repair of the stress damaged photosynthetic machinery and are required for the acclimation process. In this review we summarize the recent progress in the studies of molecular mechanisms involved during moderate heat stress on the photosynthetic machinery, especially in PSII.     

Hydrogen peroxide activates the Gas6-Axl pathway in vascular smooth muscle cells

Axl, a receptor tyrosine kinase, is involved in cell survival, proliferation, and migration. We have shown that Axl expression increases in the neointima of balloon-injured rat carotids. Because oxidative stress is known to play a major role in remodeling of injured vessels, we hypothesized that H(2)O(2) might activate Axl by promoting autophosphorylation. H(2)O(2) rapidly stimulated Axl tyrosine phosphorylation in rat vascular smooth muscle cells within 1 min that was maximal at 5 min (6-fold). The response to H(2)O(2) was concentration-dependent with EC(50) of approximately 500 microm. Axl phosphorylation was partly dependent on production of its endogenous ligand, growth arrest gene 6 (Gas6), because Axl-Fc, a fragment of Axl extracellular domain that neutralizes Gas6, inhibited H(2)O(2)-induced Axl phosphorylation by 50%. Axl phosphorylation by H(2)O(2) was also attenuated by warfarin, which inhibits Gas6 activity by preventing post-translational modification. In intact vessels Axl was phosphorylated by H(2)O(2), and Axl phosphorylation was inhibited by warfarin treatment in balloon-injured carotids. Akt, a downstream target of Axl, was phosphorylated by H(2)O(2)in Axl(+/+) mouse aorta but significantly inhibited in Axl(-/-) aorta. Intimal proliferation was decreased significantly in a cuff injury model in Axl(-/-) mice compared with Axl(+/+) mice. In summary, Axl is an important signaling mediator for oxidative stress in cultured vascular smooth muscle cells and intact vessels and may represent an important therapeutic target for vascular remodeling and response to injury.     

Regulation of CD147 cell surface expression: involvement of the proline residue in the CD147 transmembrane domain

CD147, also known as extracellular matrix metalloproteinase inducer, is a regulator of matrix metalloproteinase production and serves as a signaling receptor for extracellular cyclophilins. Here we demonstrate that the cell surface expression of CD147 is regulated by cyclophilins via the transmembrane domain of CD147. Solution binding experiments demonstrated that the transmembrane domain was both necessary and sufficient for CD147 binding to cyclophilin A (CypA). Treatment with cyclosporin A significantly reduced surface expression of CD147 and of CD8-CD147 fusion protein carrying the extracellular domain of CD8 fused to the transmembrane and cytoplasmic domains of CD147, but did not affect expression of CD8. Peptide binding studies demonstrated specific interaction between CypA and the proline-containing peptide from the CD147 transmembrane domain. Mutation of this proline residue reduced binding of CD147-derived peptides to CypA and also diminished transport of CD147 to the plasma membrane without reducing the total level of CD147 expression. These results suggest involvement of a cyclophilin-related protein in CD147 cell surface expression and provide molecular details for regulation of CD147 trafficking by cyclophilins.     

Synthesis and characterization of pyridoxine,nicotine and nicotinamide salts of dithiophosphoric acids as antibacterial agents against resistant wound infection

The pyridine-derived biomolecules are of considerable interest in developing medicinal compounds with various specific activities. Novel ammonium salts of pyridoxine, (S)-(–)-nicotine and nicotinamide with O,O-diorganyl dithiophosphoric acids (DTPA) were synthesized and characterized. The complexation of chiral monoterpenyl DTPA, including (S)-(–)-menthyl, (R)-(+)-menthyl, (1R)-endo-(+)-fenchyl, (1S,2S,3S,5R)-(+)-isopinocampheolyl derivatives, with pyridoxine and nicotine provided effective antibacterial compounds 3a,b,e,f, and 5a,b,d,f with MIC values against Gram-positive bacteria as low as 10?µM (6?µg/mL). Two selected pyridoxine and nicotine salts based on menthyl DTPA 3a and 5a were similarly active against antibiotic-resistant bacteria from burn wounds including MRSA. The compounds had enhanced amphiphilic and hemolytic properties and effectively altered surface characteristics and matrix-secreting ability of P. aeroginosa and S. aureus. MBC/MIC ratios of 3a and 5a suggested the bactericidal mode of their action. Furthermore, the compounds exhibited moderate cytotoxicity towards human skin fibroblasts (IC₅₀?=?48.6 and 57.6?µM, respectively, 72?h), encouraging their further investigation as potential antimicrobials against skin and wound infections.     

5'-carbamoylphosphonyl-[6-3H]-AZT as a tool for studying metabolic transformations of the nonradioactive counterpart, an inhibitor of HIV replication

An effective synthesis of 5'-carbamoylphosphonyl-[6-3H]-AZT was developed from [6-3H]-AZT.For the synthesized compound, chemical and enzymatic stability were determined and its penetration across HL-60 cell membranes was studied.     

Quantitative assessment of intrinsic carbonic anhydrase activity and the capacity for bicarbonate oxidation in photosystem II

On the basis of equilibrium isotopic distribution experiments using (18)O-labeled water, it is generally accepted that water is the sole substrate for O(2) production by photosystem II (PSII). Nevertheless, recent studies indicating a direct interaction between bicarbonate and the donor side of PSII have been used to hypothesize that bicarbonate may have been a physiologically important substrate for O(2) production during the evolution of PSII [Dismukes, G. C., Klimov, V. V., Baranov, S. V., Kozlov, Y. N., DasGupta, J., and Tyryshikin, A. (2001) Proc. Natl. Acad. Sci. U.S.A. 98, 2170-2175]. To test out this hypothesis and to determine whether contemporary oxygenic organisms have the capacity to oxidize bicarbonate, we employed special rapid-mixing isotopic experiments using (18)O/(13)C-labeled bicarbonate to quantify the inherent carbonic anhydrase activity in PSII samples and the potential flux of oxygen from bicarbonate into the photosynthetically produced O(2). The measurements were made on PSII samples prepared from spinach, Thermosynechococcus elongatus, and Arthrospira maxima. For the latter organism, a strain was used that grows naturally in an alkaline, high (bi)carbonate soda lake in Africa. The results reveal that bicarbonate is not the substrate for O(2) production in these contemporary oxygenic photoautotrophs when assayed under single turnover conditions.     

A new ethanolamine phosphate-containing variant of the O-antigen of Shigella flexneri type 4a

The O-specific polysaccharide (O-antigen) structure of a Shigella flexneri type 4a strain from the Dysentery Reference Laboratory (London, UK) was elucidated in 1978 and its characteristic feature was found to be α-d-glucosylation of GlcNAc at position 6, which defines O-factor IV. Our NMR spectroscopic studies of the O-specific polysaccharides of two other strains belonging to S. flexneri type 4a (G1668 from Adelaide, Australia, and 1359 from Moscow, Russia) confirmed the carbohydrate backbone structure but revealed in both strains an additional component, ethanolamine phosphate (EtnP), attached at position 3 of one of the rhamnose residues:

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Phosphorylation has not been hitherto reported in any S. flexneri O-antigen. Reinvestigation of the O-specific polysaccharide of S. flexneri type 4b showed that it is not phosphorylated and confirmed its structure established earlier.     

Structural Insights into Inhibition of Sterol 14α-Demethylase in the Human Pathogen Trypanosoma cruzi

Trypanosoma cruzi causes Chagas disease (American trypanosomiasis), which threatens the lives of millions of people and remains incurable in its chronic stage. The antifungal drug posaconazole that blocks sterol biosynthesis in the parasite is the only compound entering clinical trials for the chronic form of this infection. Crystal structures of the drug target enzyme, Trypanosoma cruzi sterol 14α-demethylase (CYP51), complexed with posaconazole, another antifungal agent fluconazole and an experimental inhibitor, (R)-4′-chloro-N-(1-(2,4-dichlorophenyl)-2-(1H-imid-azol-1-yl)ethyl)biphenyl-4-carboxamide (VNF), allow prediction of important chemical features that enhance the drug potencies. Combined with comparative analysis of inhibitor binding parameters, influence on the catalytic activity of the trypanosomal enzyme and its human counterpart, and their cellular effects at different stages of the Trypanosoma cruzi life cycle, the structural data provide a molecular background to CYP51 inhibition and azole resistance and enlighten the path for directed design of new, more potent and selective drugs to develop an efficient treatment for Chagas disease.