Sialic acid-containing lipopolysaccharides in purple nonsulfur bacteria

Lipopolysaccharides (LPS) from a number of purple nonsulfur bacteria and of phylogenetically related species were analyzed for the presence of sialic acid by gas chromatography/mass spectrometry. Species and strains of the genera Rhodobacter, Rhodopseudomonas, Rhodomicrobium, Rhodospirillum, Rhodocyclus and Rhodopila were investigated, sialic acid, however, was found only in the genus Rhodobacter. It occurs in strains of Rhodobacter capsulatus, R. sphaeroides, R. sulfidophilus and R. veldkampii. All these species belong to the -3 subgroup of purple bacteria as defined by 16S rRNA catalogues. Approximately equimolar ratios of sialic acid and of 2-keto-3-deoxy-octonate (KDO) were found in isolated LPSs. Sodium deoxycholate gel electrophoresis of these LPS-samples also suggested a location of sialic acid in the LPS core region. Sialic acid was present only in those LPSs, which exhibited a complete core region.Abbreviations LPS Lipopolysaccharide - Neu5Ac N-acetylneuraminic acid - KDO 2-Keto-3-deoxy-octonate - Neuß2Me neuraminic acid-2-methyl glycoside - DOC sodium deoxycholate - PAGE polyacrylamide gel electrophoresis - HPLC high performance liquid chromatography - TLC thin layer chromatography     

Sialylation is modulated through maturation in hemocytes from Macrobrachium rosenbergii

In this work we identified in adult and juvenile freshwater prawn, Macrobrachium rosenbergii, three major type of circulating hemocytes: fusiform; rounded; and large ovoid hemocytes. Rounded and large hemocytes represent the first defense line, since this type of cells exerts phagocytic activity as well as lectin synthesis. Considering that glycosylation plays important roles in cell communication and as a target for pathogenic microorganisms, in this report was also described the main glycosidic modifications that occur in the large and rounded hemocytes from the freshwater prawn during maturation as determined with lectins. Neu5Acalpha2,6Gal, was identified homogeneously distributed in the membrane in 90% of hemocytes from juvenile organisms. Maturation of the freshwater prawn induced a decrease or complete loss of Neu5Acalpha2,6Gal residues that were replaced with Neu5Acalpha2,3 molecules in practically all hemocytes from adult organisms. This change was paralleled by a diminution in 9-O-acetyl-neuraminic acid (Neu5,9Ac(2)) expression. T and Tn antigens (Galbetal,3 GalNAcalpha1-0-Ser/Thr or GalNAcalpha1-0-Ser/Thr, respectively), as well as N-glycosidically linked glycans, seem to be highly conserved throughout maturation. Our results show that sialylation of freshwater prawn hemocytes is modulated throughout the maturation process.     

Malondialdehyde generated from peroxidized linolenic acid causes protein modification in heat-stressed plants

When polyunsaturated fatty acids (PUFAs) in biomembrane are peroxidized, a great diversity of aldehydes is formed, and some of which are highly reactive. Thus they are thought to have biological impacts in stressed plants; however, the detailed mechanism of generation and biochemical effects are unknown. In this study, we show that chloroplasts are major organelles in which malondialdehyde (MDA) generated from peroxidized linolenic acid modifies proteins in heat-stressed plants. First, to clarify the biochemical process of MDA generation from PUFAs and its attachment to proteins, we carried out in vitro experiments using model proteins (BSA and Rubisco) and methylesters of C18 PUFAs that are major components of plant biomembrane. Protein modification was detected by Western blotting using monoclonal antibodies that recognize MDA binding to proteins. Results showed that peroxidation of linolenic acid methylester by reactive oxygen species was essential for protein modification by MDA, and the MDA modification was highly dependent on temperature, leading to a loss of Rubisco activity. When isolated spinach thylakoid membrane was peroxidized at 37 degrees C, oxygen-evolving complex 33kDa protein (OEC33) was modified by MDA. These model experiments suggest that protein modification by MDA preferentially occurs under higher temperatures and oxidative conditions, thus we examined protein modification in heat-stressed plants. Spinach plants were heat-stressed at 40 degrees C under illumination, and modification of OEC33 protein by MDA was detected. In heat-stressed Arabidopsis plants, light-harvesting complex protein was modified by MDA under illumination. This modification was not observed in linolenic acid-deficient mutants (fad3fad7fad8 triple mutant), suggesting that linolenic acid is a major source of protein modification by MDA in heat-stressed plants.     

Sialic acid attenuates puromycin aminonucleoside-induced desialylation and oxidative stress in human podocytes

Sialoglycoproteins make a significant contribution to the negative charge of the glomerular anionic glycocalyx—crucial for efficient functioning of the glomerular permselective barrier. Defects in sialylation have serious consequences on podocyte function leading to the development of proteinuria. The aim of the current study was to investigate potential mechanisms underlying puromycin aminonucleosisde (PAN)-induced desialylation and to ascertain whether they could be corrected by administration of free sialic acid.     

Abscisic acid concentrations are correlated with leaf area reductions in two salt-stressed rapid-cycling Brassica species

The greater sensitivity of B. carinata to salinity in comparison to B. napus has been linked to a greater reduction in net assimilation rate. Apparently this is not due to ion toxicity; the cause is unknown. In this report, we test the hypothesis that increases in abscisic acid (ABA) are involved in the reduction of growth by salinity. Salinity (8 dS m^–1) caused an increase of ABA concentrations in the shoot, root and callus of both species. ABA concentrations were lower in the salt-tolerant species, B. napus, than the salt-sensitive species, B. carinata, both in the whole plant and callus. Leaf expansion for both species was equally sensitive to ABA; salt stress did not significantly alter sensitivity to applied ABA. The growth inhibition increased in a hyperbolic manner with an increase in endogenous ABA concentration. These results indicate that ABA in salt-stressed plants may play a role in the inhibition of growth. The photosynthesis of salt-sensitive species, B. carinata, was also decreased by salinity, corresponding to the reduction in growth. The decreased photosynthesis does not appear to be the cause of the growth reduction, because photosynthesis was not inhibited by short-term exposure to salinity and photosynthesis was poorly correlated with endogenous ABA concentrations.     

A decade of progress in understanding vitamin E synthesis in plants

The chloroplasts of higher plants contain and elaborate many unique biochemical pathways that produce an astonishing array of compounds that are vital for plastid function and are also important from agricultural and nutritional perspectives. One such group of compounds is the tocochromanols (more commonly known as Vitamin E), which is a class of four tocopherols and four toctorienols, lipid-soluble antioxidants that are only synthesized by plants and other oxygenic, photosynthetic organisms. Though the essential nature of tocopherols in mammalian diets was recognized over 80 years ago and the biosynthetic pathway in plants and algae elucidated in the late 1970s and early 80s, it has only been in the past decade that the genes and proteins for tocopherol synthesis have finally been isolated and characterized. The use of model plant and cyanobacterial systems has driven this gene discovery to the point that manipulation of tocopherol levels and types in various plant tissues and crops is becoming a reality. This article reviews progress since 1996 in the molecular and genetic understanding of tocopherol synthesis in the model photosynthetic organisms Arabidopsis thaliana and Synechocystis PCC6803 as a primer for current and future efforts to manipulate the levels of this essential nutrient in food crops by breeding and transgenic approaches.     

Effects of tunicamycin on protein glycosylation and development inVolvox carteri

Summary The involvement of protein glycosylation in regulation of the development of the multicellular green alga,Volvox carteri, was studied using the antibiotic, tunicamycin. Three specific developmental processes were found to be affected by the antibiotic: reproductive cell maturation; establishment of polar cellular organization during embryogenesis and release of progeny spheroids from the parental spheroids. Tunicamycin inhibited the transfer of GlcNAc-1-phosphate to dolichyl phosphate which is catalyzed byVolvox membrane preparations. Changes in the glycosylation of several secreted and cellular glycoproteins were observed when proteins were labelled with radioactive amino acids and sugars in the absence and presence of tunicamycin and then electrophoresed on sodium dodecylsulfate-polyacrylamide slab gels. The levels of a few secreted proteins were reduced in tunicamycin treated cultures and one protein band appeared exclusively in the treated cells. Tunicamycin treatment also altered the electrophoretic mobility of radio-iodinated surface macromolecules. Binding of concanavalin A by tunicamycin treatedVolvox spheroids was drastically reduced. It is there-fore likely that the aberrant development results from inhibition of protein glycosylation and the consequent changes in the structure of the cellular, secreted and surface glycoproteins.     

Synthesis of the starfish ganglioside LLG-3 tetrasaccharide

The first synthesis of the ganglioside LLG-3 tetrasaccharide, which has attractive biological activities as well as a unique structure, is described. A C8-methoxy decorated sialic acid building block was initially prepared and a glycolic acid moiety was then introduced by sialylation. Amide condensation between the sialyl glycolic acid and an amino group at C5 on the sialyllactoside unit afforded the fully protected LLG-3 tetrasaccharide. Finally, the desired tetrasaccharide part of LLG-3 was obtained after careful global deprotection.     

Sialic acid attenuates the cytotoxicity of the lipid hydroperoxides HpODE and HpETE

Reduction of peroxide molecular species is an essential function in living organisms. In previous studies, we proposed a new function for the sialic acid N-acetylneuraminic acid (Neu5Ac)—that of antioxidant/hydrogen peroxide scavenging agent. On the basis of the reaction scheme, Neu5Ac is thought to act as a general antioxidant of all hydroperoxide-type species (R-OOHs). The concentration of tert-butyl hydroperoxide (t-BuOOH) decreased after co-incubation with N-acetylneuraminic acid. Neu5Ac also decreased the R-OOH concentration in solutions of peroxylinolenic acid (13(S)-hydroperoxy-(9Z,11E)-octadecadienoic acid, HpODE) and peroxyarachidonic acid (15(S)-hydroperoxy-(5Z,8Z,11Z,13E)-eicosatetraenoic acid, HpETE)—two lipid hydroperoxides that participate in many physiological events. Moreover, the cytotoxicity of both these lipid hydroperoxides was attenuated by reaction with Neu5Ac acid. Our results suggest that N-acetylneuraminic acid is a potential antioxidant of most hydroperoxides that accumulate in organisms.     

Sialic acids acquired by Pseudomonas aeruginosa are involved in reduced complement deposition and siglec mediated host-cell recognition

The opportunism of Pseudomonas aeruginosa (PA) in immunocompromised hosts prompted us to explore the potential role of sialic acids (Sia) in this phenomenon. Culture of PA in the presence of exogenous Sia resulted in linkage-specific incorporation of Sia which was associated with decreased complement deposition on the bacteria. Sia acquired by PA mediated enhanced binding of bacteria to recombinant-CHO cells expressing human siglec-7 or siglec-9, as well as to human NK-cells and monocytes naturally expressing these siglecs. Therefore, Sia may be acquired by PA in the host and contribute to bacterial pathogenicity and host-cell interactions via reduction of complement deposition and siglec-dependent recognition.     

Spatial distribution of leaf form and the self-thinning exponent are affected by the sensitivity of the response to abscisic acid in anArabidopsis thaliana population

The spatial distribution of leaves is related to the exponent of the self-thinning relationship in plant populations. In this study, we evaluated the fractal dimension of rosette leaves of wild-type (WT)Arabidopsis thaliana and of an abscisic acid (ABA) -insensitive mutant (abi2-1) to test a model of the spatial distribution of leaf form in anArabidopsis population based on subdivision of a cube surrounding the leaf into uniform boxes and to investigate ABA’s affect on this model of the leaf. The values of the self-thinning exponent were -1.31 and -1.45 for WT andabi2-1. The mean dimensions of the box used to model the spatial distribution of leaf form, estimated using our model, were 2.08 and 2.03, respectively. By assuming that the box dimension equals the fractal dimension within the populations, the predicted self-thinning exponent equaled -1.40 for WT and -1.49 forabi2-1. When exogenous ABA was applied to both genotypes, the self-thinning exponent became -1.26 and -1.43 for WT andabi2-1, and the exponents predicted using the dimensions of the box were -1.37 and -1.46, respectively. The empirically predicted exponent equaled that predicted using the dimensions of the box (95% confidence interval). Empirical prediction of the spatial pattern using the two genotypes with and without ABA showed that ABA influenced the spatial form of the rosette leaves. Therefore, sensitivity to ABA can affect self-thinning through genetically determined changes in leaf form and its spatial distribution.     

Interactions of abscisic acid and sugar signalling in the regulation of leaf senescence

Leaf senescence can be triggered by a high availability of carbon relative to nitrogen or by external application of abscisic acid (ABA). Most Arabidopsis mutants with decreased sugar sensitivity during early plant development are either ABA insensitive (abi mutants) or ABA deficient (aba mutants). To analyse the interactions of carbon, nitrogen and ABA in the regulation of senescence, wild-type Arabidopsis thaliana (L.) Heynh. and aba and abi mutants were grown on medium with varied glucose and nitrogen supply. On medium containing glucose in combination with low, but not in combination with high nitrogen supply, senescence was accelerated and sucrose, glucose and fructose accumulated strongly. In abi mutants that are not affected in sugar responses during early development (abi1-1 and abi2-1), we observed no difference in the sugar-dependent regulation of senescence compared to wild-type plants. Similarly, senescence was not affected in the sugar-insensitive abi4-1 mutant. In contrast, the abi5-1 mutant did exhibit a delay in senescence compared to its wild type. As ABA has been reported to induce senescence and ABA deficiency results in sugar insensitivity during early development, we expected senescence to be delayed in aba mutants. However, the aba1-1 and aba2-1 mutants showed accelerated senescence compared to their wild types on glucose-containing medium. Our results show that, in contrast to sugar signalling in seedlings, ABA is not required for the sugar-dependent induction of leaf senescence. Instead, increased sensitivity to osmotic stress could have triggered early senescence in the aba mutants.Abbreviations ABA Abscisic acid - aba Abscisic acid deficient - abi Abscisic acid insensitive - F_v/F_m Maximum efficiency of photosystem II photochemistry     

In vivo demonstration of acid phosphatase activity in the rhizosphere of soil-grown plants

For in vivo demonstration of acid phosphatase activity in the rhizosphere of soil-grown plants filter papers were treated with a mixture of 1-naphthyl phosphate as substrate and the diazonium salt Fast Red TR as an indicator. After enzymatic hydrolysis, 1-naphthol forms a red complex with Fast Red TR. This method was applied to 8-day old maize plants and 3-year old Norway spruce plants growing in rhizoboxes in soil under non-sterile conditions. The treated filter paper is placed at the surface of roots and soil and acid phosphatase activity is visualized as a red-coloured root print on the filter paper. The method can be used as a qualitative analysis of acid phosphatase in the rhizosphere. It also allows a rough estimate of phosphatase activity in different root zones.     

Acetylsalicylic acid induces programmed cell death in Arabidopsis cell cultures

Acetylsalicylic acid (ASA), a derivative from the plant hormone salicylic acid (SA), is a commonly used drug that has a dual role in animal organisms as an anti-inflammatory and anticancer agent. It acts as an inhibitor of cyclooxygenases (COXs), which catalyze prostaglandins production. It is known that ASA serves as an apoptotic agent on cancer cells through the inhibition of the COX-2 enzyme. Here, we provide evidences that ASA also behaves as an agent inducing programmed cell death (PCD) in cell cultures of the model plant Arabidopsis thaliana, in a similar way than the well-established PCD-inducing agent H(2)O(2), although the induction of PCD by ASA requires much lower inducer concentrations. Moreover, ASA is herein shown to be a more efficient PCD-inducing agent than salicylic acid. ASA treatment of Arabidopsis cells induces typical PCD-linked morphological and biochemical changes, namely cell shrinkage, nuclear DNA degradation, loss of mitochondrial membrane potential, cytochrome c release from mitochondria and induction of caspase-like activity. However, the ASA effect can be partially reverted by jasmonic acid. Taking together, these results reveal the existence of common features in ASA-induced animal apoptosis and plant PCD, and also suggest that there are similarities between the pathways of synthesis and function of prostanoid-like lipid mediators in animal and plant organisms.     

The proteome of the chloroplast lumen of higher plants

Recent research in proteomics of the higher plant chloroplast has achieved considerable progress and added to our knowledge of lumenal chloroplast proteins. This work shows that chloroplast lumen has its own specific proteome and may comprise as many as 80 proteins. Although the new map of the lumenal proteome provides a great deal of information, it also raises numerous questions because the physiological functions of most of the novel lumenal proteins are unknown. In this Minireview, we summarize the latest discoveries regarding lumenal proteins and present the currently known facts about the lumenal chloroplast proteome of higher plants.     

Participation of proteolytic enzymes in the interaction of plants with phytopathogenic microorganisms

Different forms of participation of proteolytic enzymes in pathogenesis and plant defense are reviewed. Together with extracellular proteinases, phytopathogenic microorganisms produce specific effectors with proteolytic activity and are able to act on proteins inside the plant cell. In turn, plants use both extracellular and intracellular proteinases for defense against phytopathogenic microorganisms. Among the latter, a special role belongs to vacuolar processing enzymes (legumains), which perform the function of caspases in the plant cell.