Leukotriene B4 level in neutrophils from allergic and healthy subjects stimulated by low concentration of calcium ionophore A23187. Effect of exogenous arachidonic acid and possible endogenous source.

Peripheral blood neutrophils from patients with allergic rhinitis and from normal subjects were incubated for 5 min at 37 degrees C with 0.15 microM calcium ionophore A23187 in the absence or presence of exogenous arachidonic acid (2.5 to 10 microM). In neutrophils from allergic patients, the leukotriene B4 (LTB4) level was significantly increased by exogenous arachidonic acid in a concentration-dependent manner (16.2 +/- 4.2 and 38.1 +/- 6.8 pmol/5 min per 2 X 10(6) cells in the absence and presence of 10 microM arachidonic acid, respectively; P less than 0.005; n = 8). The LTB4 level in neutrophils from healthy subjects was only 0.97 +/- 0.17 pmol/5 min per 2 x 10(6) cells (n = 5) and was not enhanced by exogenous arachidonate. When cells from allergic patients were challenged in the presence of exogenous [1-14C]arachidonic acid, released LTB4 was radiolabeled and the incorporated radioactivity increased with the labeled arachidonate concentration. Labeled LTB4 was never detectable after incubating neutrophils from normal donors with exogenous labeled arachidonate. When neutrophils were incubated with [1-14C]arachidonate for 1 h, the different lipid pools of the two cell populations were labeled but both types of neutrophils produced unlabeled LTB4 in response to ionophore stimulation. The hydrolysis of choline and ethanolamine phospholipids into diacyl-, alkenylacyl- and alkylacyl-species revealed that solely the alkylacyl-subclass of phosphatidylcholine was unlabeled. We conclude (i) that neutrophils from allergic patients stimulated by low ionophore concentration produce more LTB4 than neutrophils from healthy subjects and incorporate exogenous arachidonate, (ii) that endogenous arachidonate converted to LTB4 by the 5-lipoxygenase pathway may provide only from 1-O-alkyl-2-arachidonoyl-glycero-3-phosphocholine.     

Localization of general and regulatory proteolysis in Bacillus subtilis cells

Protein degradation mediated by ATP-dependent proteases, such as Hsp100/Clp and related AAA+ proteins, plays an important role in cellular protein homeostasis, protein quality control and the regulation of, e.g. heat shock adaptation and other cellular differentiation processes. ClpCP with its adaptor proteins and other related proteases, such as ClpXP or ClpEP of Bacillus subtilis, are involved in general and regulatory proteolysis. To determine if proteolysis occurs at specific locations in B. subtilis cells, we analysed the subcellular distribution of the Clp system together with adaptor and general and regulatory substrate proteins, under different environmental conditions. We can demonstrate that the ATPase and the proteolytic subunit of the Clp proteases, as well as the adaptor or substrate proteins, form visible foci, representing active protease clusters localized to the polar and to the mid-cell region. These clusters could represent a compartmentalized place for protein degradation positioned at the pole close to where most of the cellular protein biosynthesis and also protein quality control are taking place, thereby spatially separating protein synthesis and degradation.     

The Arabidopsis phosphatase PP2C49 negatively regulates salt tolerance through inhibition of AtHKT1;1

Type 2C protein phosphatases (PP2Cs) are the largest protein phosphatase family. PP2Cs dephosphorylate substrates for signaling in Arabidopsis, but the functions of most PP2Cs remain unknown. Here, we characterized PP2C49 (AT3G62260, a Group G PP2C), which regulates Na⁺ distribution under salt stress and is localized to the cytoplasm and nucleus. PP2C49 was highly expressed in root vascular tissues and its disruption enhanced plant tolerance to salt stress. Compared with wild type, the pp2c49 mutant contained more Na⁺ in roots but less Na⁺ in shoots and xylem sap, suggesting that PP2C49 regulates shoot Na⁺ extrusion. Reciprocal grafting revealed a root‐based mechanism underlying the salt tolerance of pp2c49. Systemic Na⁺ distribution largely depends on AtHKT1;1 and loss of function of AtHKT1;1 in the pp2c49 background overrode the salt tolerance of pp2c49, resulting in salt sensitivity. Furthermore, compared with plants overexpressing PP2C49 in the wild‐type background, plants overexpressing PP2C49 in the athtk1;1 mutant background were sensitive to salt, like the athtk1;1 mutants. Moreover, protein–protein interaction and two‐voltage clamping assays demonstrated that PP2C49 physically interacts with AtHKT1;1 and inhibits the Na⁺ permeability of AtHKT1;1. This study reveals that PP2C49 negatively regulates AtHKT1;1 activity and thus determines systemic Na⁺ allocation during salt stress.     

Raiding and Foraging Behavior of the Blood-Red Ant,Formica sanguinea Latr. (Hymenoptera,Formicidae)

In this paper we report the results of a detailed study on the behavioral ecology of slave raiding and foraging activity in the European blood-red ant, Formica sanguinea Latr. The field study was conducted over an unbroken period of 78 days, during which the activity of two dulotic colonies of this facultative slave-maker was observed for 10 h each day. It was possible to observe 26 raids distributed over 23 days, among which 18 were followed by the sacking of nests belonging to the species F. cunicularia, F. fusca, and Lasius emarginatus, whereas 8 failed. Simple, continuous, and simultaneous raids occurred. We recorded the timing, frequency, distance, and direction of slave raids, including the number of participants and the type of booty. Particular attention was devoted to the scouting behavior and raiding organization. Moreover, every day, we observed foraging and predatory behavior, during which adult insects (mainly ants), seeds, and berries were retrieved to the dulotic colonies. On the basis of our observations F. sanguinea seems to be a very efficient slave-maker and predatory species of the Raptiformica subgenus. Moreover, its dulotic behavior may be regarded as a continuation and an expansion of its foraging and predatory behavior, as predicted by Darwin's hypothesis for the origin and evolution of slavery in ants.     

Selective acceptance of the brood of two formicine slave-making ants by host and non-host related species

Summary In a laboratory choice-test, free-living ant workers ofFormica cunicularia andF. rufibarbis (subgenusServiformica), both potentially slave species of the obligatory slave-makerPolyergus rufescens, cared for cocoons of this parasite and for homocolonial cocoons at comparable rates. Both potential hosts did not differ in their capacity to rear the parasite brood. This fact is discussed in relation to host selection and specificity inP. rufescens. No such attraction and/or tolerance was found towards cocoons of the facultative slave-makerFormica sanguinea, which also enslaves both host species. Workers ofF. lugubris, a species which is never enslaved, destroyed cocoons from both slave-making species. The attractiveness of the brood ofP. rufescens for both potentially slave species could be due to an interspecific brood pheromone in addition to brood mimicry. An alternative hypothesis is a close phylogenetic distance between this slave-maker andServiformica species. The capacity to gain acceptance by adult slave workers might be one of the crucial evolutionary steps separating obligatory from facultative slave-making ants.     

Substructural specificity and polyvalent carbohydrate recognition by the Entamoeba histolytica and rat hepatic N- acetylgalactosamine/galactose lectins

Both the Entamoeba histolytica lectin, a virulence factor for the causative agent of amebiasis, and the mammalian hepatic lectin bind to N-acetylgalactosamine (GalNAc) and galactose (Gal) nonreducing termini on oligosaccharides, with preference for GalNAc. Polyvalent GalNAc- derivatized neoglycoproteins have >1000-fold enhanced binding affinity for both lectins (Adler,P., Wood,S.J., Lee,Y.C., Lee,R.T., Petri,W.A.,Jr. and Schnaar,R.L.,1995, J. Biol. Chem ., 270, 5164-5171). Substructural specificity studies revealed that the 3-OH and 4-OH groups of GalNAc were required for binding to both lectins, whereas only the E.histolytica lectin required the 6-OH group. Whereas GalNAc binds with 4-fold lower affinity to the E.histolytica lectin than to the mammalian hepatic lectin, galactosamine and N-benzoyl galactosamine bind with higher affinity to the E. histolytica lectin. Therefore, a synthetic scheme for converting polyamine carriers to poly-N-acyl galactosamine derivatives (linked through the galactosamine primary amino group) was developed to test whether such ligands would bind the E.histolytica lectin with high specificity and high affinity. Contrary to expectations, polyvalent derivatives including GalN6lys5, GalN4desmosine, GalN4StarburstTMdendrimer, and GalN8StarburstTMdendrimer demonstrated highly enhanced binding to the mammalian hepatic lectin but little or no enhancement of binding to the E.histolytica lectin. We propose that the mammalian hepatic lectin binds with greatest affinity to GalNAc "miniclusters," which mimic branched termini of N-linked oligosaccharides, whereas the E.histolytica lectin binds most effectively to "maxiclusters," which may mimic more widely spaced GalNAc residues on intestinal mucins.      

Cytochemical demonstration of hydrogen peroxide in polymorphonuclear leukocyte phagosomes

Phagocytosis by polymorphonuclear leukocytes (PMN) is accompanied by specific morphological and metabolic events which may result in the killing of internalized micro-organism. Hydrogen peroxide is produced in increased amounts during phagocytosis (17) and in combination with myeloperoxidase and halide ions constitute a potent, microbicidal mechanism (8,9,11). There can be direct iodination of micro-organisms (10), or alternatively, other intermediate reaction products, i.e. chloramines and aldehydes (21), can exert a microbicidal effect. The H2O2-peroxidase-halide system is presumed to operate within the phagocytic vacuole (12,18). Myeloperoxidase, present in the primary granules of PMN, enters the phagocytic vacuole during degranulation (1,4,7), and halide ions are probably derived from the extracellular medium or are present in the PMN (see 11, 18). For the operation of this system in intact cells, the presence of H2O2 in the phagocytic vacuole is necessary, and indeed this has been suggested by the work of several investigators (12, 18, 21). In the present investigation, the diaminobenzidine reaction of Graham and Karnovsky (5), modified to utilize endogenous myeloperoxidase and hydrogen peroxide, has been applied to actively phagocytizing PMN to demonstrate cytochemically the presence of H2O2 in the phagocytic vacuole.     

Tobamovirus evolution: gene overlaps, recombination, and taxonomic implications

Tobamoviruses, mostly isolated from solanaceous plants, may represent ancient virus lineages that have codiverged with their hosts. Recently completed nucleotide sequences of six nonsolanaceous tobamoviruses allowed assessment of the codivergence hypothesis and support a third subgroup within tobamoviruses. The genomic sequences of 12 tobamoviruses and the partial sequences of 11 others have been analyzed. Comparisons of the predicted protein sequences revealed three clusters of tobamoviruses, corresponding to those infecting solanaceous species (subgroup 1), those infecting cucurbits and legumes (subgroup 2), and those infecting crucifers. The orchid-infecting odontoglossum ringspot tobamovirus was associated with subgroup 1 genomes by its coat and movement protein sequences, but with the crucifer-pathogenic tobamoviruses by the remainder of its genome, suggesting that it is the progeny of a recombinant. For four of five genomic regions, subgroup 1 and 3 genomes were equidistant from a subgroup 2 genome chosen for comparison, suggesting uniform rates of evolution. A phylogenetic tree of plant families based on the tobamoviruses they harbor was congruent with that based on rubisco sequences but had a different root, suggesting that codivergence was tempered by rare events of viruses of one family colonizing another family. The proposed subgroup 3 viruses probably have an origin of virion assembly in the movement protein gene, a large (25-codon) overlap of movement and coat protein open reading frames, and a comparably shorter genome. Codon-position- dependent base compositions and codon prevalences suggested that the coat protein frame of the overlap region was ancestral. Bootstrapped parsimony analysis of the nucleotides in the overlap region and of the sequences translated from the -1 frame (the subgroup 3 movement protein frame) of this region produced trees inconsistent with those deduced from other regions. The results are consistent with a model in which a no or short overlap organization was ancestral. Despite encoding of subgroup 2 and 3 movement protein C-termini by nonhomologous nucleotides, weak similarities between their amino acid sequences suggested convergent sequence evolution.      

Secondary hip dysplasia increases risk for early coxarthritis after Legg-Calve-Perthes disease. A study of 255 hips

Abstract

The biomechanical parameters of the hip joint articular surface were analysed in 141 adult hips after Legg-Calve Perthes Disease, and 114 contralateral unaffected hips (controls), by using HIPSTRESS mathematical models. Geometrical parameters, assessed from anteroposterior and axial radiograms, were used as input to models for resultant hip force and contact hip stress. Results confirm previous indications that head enlargement after the Legg-Calve-Perthes Disease compensates the values of hip stress. Furthermore, it was found that an increased risk for coxarthritis development after the disease is secondary to concomitant hip dysplasia, with considerable and statistically significantly lower centre-edge angle and unfavourable distribution of stress.