Evidence for strong host clone-parasite species interactions in the Daphnia microparasite system

Abstract Organisms are often confronted with multiple enemy species. Defenses against different parasite species may be traded off against each other. However, if resistance is based on (potentially costly) general defense mechanisms, it may be positively correlated among parasites. In an experimental study, we confronted 19 clones from one Daphnia magna population with two bacterial and three microsporidian parasite species. All parasites were isolated from the same pond as the hosts. Host clones were specific in their susceptibility towards different parasite species, and parasite species were host-clone specific in their infectivity, spore production, and virulence, resulting in highly significant host-parasite interactions. Since the Daphnia 's resistance to different parasite species showed no obvious correlation, neither general defense mechanisms nor trade-offs in resistance explain our findings. None of the Daphnia clones were resistant to all parasite species, and the average level of resistance was quite similar among clones. This may reflect a cost of defense, so that the cumulative cost of being resistant to all parasite species might be too high.     

Endogenous RGS proteins facilitate dopamine D(2S) receptor coupling to G(alphao) proteins and Ca2+ responses in CHO-K1 cells

The role of RGS proteins on dopaminergic D2S receptor (D2SR) signalling was investigated in Chinese hamster ovary (CHO)-K1 cells, using recombinant RGS protein- and PTX-insensitive G alphao proteins. Dopamine-mediated [35S]GTPgammaS binding was attenuated by more than 60% in CHO-K1 D2SR cells coexpressing a RGS protein- and PTX-insensitive G(alphao)Gly184Ser:Cys351Ile protein versus cells coexpressing a similar amount of PTX-insensitive G alphaoCys351Ile protein. Dopamine-agonist-mediated Ca2+ responses were dependent on the coexpression with a G alphao Cys351Ile protein and were fully abolished upon coexpression with a G alphaoGly184Ser:Cys351Ile protein. These results suggest that interactions between the G alphao protein and RGS proteins are involved in efficient D2SR signalling.     

PAK interacts with NCK and MLK2 to regulate the activation of jun N-terminal kinase

The p21-GTPase activated kinase, PAK1, and the mixed lineage kinase, MLK2, have been implicated in the activation of jun N-terminal kinase (JNK). However, the role of PAK1 in JNK activation is still not understood. Here we show that over-expression of the SH3-SH2 adapter Nck 'squelches' JNK activation but this squelching is relieved by over-expression of PAK1. In turn, PAK1 squelches activation of JNK by MLK2 and these kinases interact via their catalytic domains. The data suggest that PAK1 recruits MLK2 to an activated receptor via the adapter Nck, but cannot itself induce activation of the JNK cascade.     

Nitric oxide binding properties of neuroglobin. A characterization by EPR and flash photolysis

Neuroglobin is a recently discovered member of the globin superfamily. Combined electron paramagnetic resonance and optical measurements show that, in Escherichia coli cell cultures with low O(2) concentration overexpressing wild-type mouse recombinant neuroglobin, the heme protein is mainly in a hexacoordinated deoxy ferrous form (F8His-Fe(2+)-E7His), whereby for a small fraction of the protein the endogenous protein ligand is replaced by NO. Analogous studies for mutated neuroglobin (mutation of E7-His to Leu, Val, or Gln) reveal the predominant presence of the nitrosyl ferrous form. After sonication of the cells wild-type neuroglobin oxidizes rapidly to the hexacoordinated ferric form, whereas NO ligation initially protects the mutants from oxidation. Flash photolysis studies of wild-type neuroglobin and its E7 mutants show high recombination rates (k(on)) and low dissociation rates (k(off)) for NO, indicating a high intrinsic affinity for this ligand similar to that of other hemoglobins. Since the rate-limiting step in ligand combination with the deoxy-hexacoordinated wild-type form involves the dissociation of the protein ligand, NO binding is slower than for the related mutants. Structural and kinetic characteristics of neuroglobin and its mutants are analyzed. NO production in rapidly growing E. coli cell cultures is discussed.     

Biochemical and molecular characterization of a hydroxyjasmonate sulfotransferase from Arabidopsis thaliana

12-Hydroxyjasmonate, also known as tuberonic acid, was first isolated from Solanum tuberosum and was shown to have tuber-inducing properties. It is derived from the ubiquitously occurring jasmonic acid, an important signaling molecule mediating diverse developmental processes and plant defense responses. We report here that the gene AtST2a from Arabidopsis thaliana encodes a hydroxyjasmonate sulfotransferase. The recombinant AtST2a protein was found to exhibit strict specificity for 11- and 12-hydroxyjasmonate with K(m) values of 50 and 10 microm, respectively. Furthermore, 12-hydroxyjasmonate and its sulfonated derivative are shown to be naturally occurring in A. thaliana. The exogenous application of methyljasmonate to A. thaliana plants led to increased levels of both metabolites, whereas treatment with 12-hydroxyjasmonate led to increased level of 12-hydroxyjasmonate sulfate without affecting the endogenous level of jasmonic acid. AtST2a expression was found to be induced following treatment with methyljasmonate and 12-hydroxyjasmonate. In contrast, the expression of the methyljasmonate-responsive gene Thi2.1, a marker gene in plant defense responses, is not induced upon treatment with 12-hydroxyjasmonate indicating the existence of independent signaling pathways responding to jasmonic acid and 12-hydroxyjasmonic acid. Taken together, the results suggest that the hydroxylation and sulfonation reactions might be components of a pathway that inactivates excess jasmonic acid in plants. Alternatively, the function of AtST2a might be to control the biological activity of 12-hydroxyjasmonic acid.     

Effect of different densities of live and dead Chlorella vulgaris on the population growth of rotifers Brachionus calyciflorus and Brachionus patulus (Rotifera)

In order to maintain rotifer populations during periods of low algal production, it is necessary to offer alternate diets, some of which include forms of preserved algae. The present work is based on the effect of live and dead Chlorella vulgaris on the population growth of Brachionus calyciflorus and Brachionus patulus. The experimental design consisted of three algal levels (0.5 x 10(6), 1.5 x 10(6) and 4.5 x 10(6) cells ml-1) offered in three forms (living, frozen and heat-killed). The maximal population density values for B. calyciflorus ranged from 55 +/- 1 ind. ml-1 (at 0.5 x 10(6) cells ml-1) to 471 +/- 72 ind. ml-1 (at 4.5 x 10(6) cells ml-1) with live Chlorella, but was much lower (6 +/- 1 to 26 +/- 6 ind. ml-1) with frozen or heat-killed alga under comparable food levels. However, the maximum population density of B. patulus under live or or heat-killed Chlorella was similar at comparable algal levels but when offered frozen algae it was four times less. The highest mean peak population density was 1,277 +/- 83 ind. ml-1 under 4.5 x 10(6) cells ml-1. The rate of population increase for B. calyciflorus varied from 0.50 to 0.79 using live Chlorella, but under comparable conditions, this range was lower (0.21 to 0.31) for B. patulus. Results have been discussed in light of possible application for aquaculture.     

Characterization of Ca2+ release from heterogeneous Ca2+ stores in sarcoplasmic reticulum isolated from arterial and gastric smooth muscle

Ca2+ transport was investigated in vesicles of sarcoplasmic reticulum subfractionated from bovine main pulmonary artery and porcine gastric antrum using digitonin binding and zonal density gradient centrifugation. Gradient fractions recovered at 15-33% sucrose were studied as the sarcoplasmic reticulum component using Fluo-3 fluorescence or 45Ca2+ Millipore filtration. Thapsigargin blocked active Ca2+ uptake and induced a slow Ca2+ release from actively loaded vesicles. Unidirectional 45Ca2+ efflux from passively loaded vesicles showed multicompartmental kinetics. The time course of an initial fast component could not be quantitatively measured with the sampling method. The slow release had a half-time of several minutes. Both components were inhibited by 20 microM ruthenium red and 10 mM Mg2+. Caffeine, inositol 1,4,5-trisphosphate, ATP, and diltiazem accelerated the slow component. A Ca2+ release component activated by ryanodine or cyclic adenosine diphosphate ribose was resolved with Fluo-3. Comparison of tissue responses showed that the fast Ca2+ release was significantly smaller and more sensitive to inhibition by Mg2+ and ruthenium red in arterial vesicles. They released more Ca2+ in response to inositol 1,4,5-trisphosphate and were more sensitive to activation by cyclic adenosine diphosphate ribose. Ryanodine and caffeine, in contrast, were more effective in gastric antrum. In each tissue, the fraction of the Ca2+ store released by sequential application of caffeine and inositol 1,4,5-trisphosphate depended on the order applied and was additive. The results indicate that sarcoplasmic reticulum purified from arterial and gastric smooth muscle represents vesicle subpopulations that retain functional Ca2+ channels that reflect tissue-specific pharmacological modulation. The relationship of these differences to physiological responses has not been determined.