Tyrosine sulfation of glycoprotein I(b)alpha. Role of electrostatic interactions in von Willebrand factor binding

Glycoprotein I(b)alpha (GP I(b)alpha), the ligand binding subunit of the platelet glycoprotein Ib-IX-V complex, is sulfated on three tyrosine residues (Tyr-276, Tyr-278, and Tyr-279). This posttranslational modification is known to be critical for von Willebrand factor (vWF) binding; yet it remains unclear whether it provides a specific structure or merely contributes negative charges. To investigate this issue, we constructed cell lines expressing GP I(b)alpha polypeptides with the three tyrosine residues converted to either Glu or Phe and studied the ability of these mutants to bind vWF in the presence of modulators or shear stress. The mutants were expressed normally on the cell surface as GP Ib-IX complexes, with the conformation of the ligand-binding domain preserved, as judged by the binding of conformation-sensitive monoclonal antibodies. In contrast to their normal expression, both mutants were functionally abnormal. Cells expressing the Phe mutant failed to bind vWF in the presence of either ristocetin or botrocetin. These cells adhered to and rolled on immobilized vWF only when their surface receptor density was increased to twice the level that supported adhesion of cells expressing the wild-type receptor and even then only 20% as many rolled and rolled significantly faster than wild-type cells. Cells expressing the Glu mutant, on the other hand, were normal with respect to ristocetin-induced vWF binding and adhesion to immobilized vWF but were markedly defective in botrocetin-induced vWF binding. These results indicate that GP I(b)alpha tyrosine sulfation influences the interaction of this polypeptide with vWF primarily by contributing negative charges under physiological conditions and when the interaction is induced by ristocetin but contributes a specific structure to the botrocetin-induced interaction.     

Human brain synembryn interacts with Gsalpha and Gqalpha and is translocated to the plasma membrane in response to isoproterenol and carbachol

Heterotrimeric G-proteins transduce signals from heptahelical transmembrane receptors to different effector systems, regulating diverse complex intracellular pathways and functions. In brain, facilitation of depolarization-induced neurotransmitter release for synaptic transmission is mediated by Gsalpha and Gqalpha. To identify effectors for Galpha-proteins, we performed a yeast two-hybrid screening of a human brain cDNA library, using the human Galphas protein as a bait. We identified a protein member of the synembryn family as one of the interacting proteins. Extending the study to other Galpha subunits, we found that Gqalpha also interacts with synembryn, and these interactions were confirmed by in vitro pull down studies and by in vivo confocal laser microscopy analysis. Furthermore, synembryn was shown to translocate to the plasma membrane in response to carbachol and isoproterenol. This study supports recent findings in C. elegans where, through genetic studies, synembryn was shown to act together with Gqalpha regulating neuronal transmitter release. Based on these observations, we propose that synembryn is playing a similar role in human neuronal cells.     

Antioxidant and insect growth regulatory activities of stilbenes and extracts from Yucca periculosa

The methanol extract from the bark of Yucca periculosa F. Baker afforded 4,4'-dihydroxstilbene, resveratrol and 3,3',5,5'-tetrahydroxy-4-methoxystilbene and had growth regulatory activity against the Fall Army worm (Spodoptera frugiperda J.E. Smith, Lepidoptera:Noctuidae) an insect pest of corn. The most active compound was 3,3',5,5'-tetrahydroxy-4-methoxystilbene which had significant effects at 3 microg/g in diets. In addition to the inhibitory activity on bleaching of crocin induced by alkoxyl radicals, these compounds also demonstrated scavenging properties toward 2,2-diphenyl-1-picrylhydrazyl in TLC autographic and spectrophotometric assays. Our results indicate that these compounds could be involved in interference of sclerotization and moulting. These compounds appear to have selective effects on the pre-emergence metabolism of the insect. The results were fully comparable to known natural insect growth inhibitors such as gedunin and Cedrela extracts and have had a possible role as natural insecticidal agents.     

A hidden sensory function for motor cortex

Sensory perception has traditionally been attributed to the activation of sensory cortices. However, by inducing an illusory perception of movement, Naito and colleagues show in this issue of Neuron that the illusory perception of movement is related to activation of primary motor cortex.     

Continental-scale patterns of nutrient and fish effects on shallow lakes: synthesis of a pan-European mesocosm experiment

1. Results are analysed from 11 experiments in which effects of fish addition and nutrient loading on shallow lakes were studied in mesocosms. The experiments, five in 1998, six in 1999, were carried out in six lakes, distributed from Finland to southern Spain, according to a standard protocol. 2. Effects of the treatments on 29 standard chemical, phytoplankton and zooplankton variables are examined to assess the relative importance of bottom‐up (nutrient enrichment) and top‐down (fish predation) effects. For each year, the experiments in different locations are treated as replicates in a meta‐analysis. Results of individual experiments are then compared in terms of the patterns of significant influences of nutrient addition and fish predation with these overall results (the baseline), and between years in the same location. 3. The overall meta‐analysis gave consistent results across the 2 years, with nutrient loading influencing all of the chemical variables, and on average 31% of primary producer and 39% of zooplankton variables. In contrast, fish influenced none of the chemical variables, 11% of the primary producer and 44% of the zooplankton variables. Nutrient effects on the system were thus about three times greater than fish effects, although fish effects were not inconsiderable. 4. The relative importance of nutrients and fish in individual experiments often differed between years at the same location and effects deviated to varying degrees from the baseline. These deviations were treated as measures of consistency (predictability) of conclusions in repeat experiments. Consistency increased southwards and this is interpreted as a consequence of more variable annual weather northwards. 5. The influence of nutrient loading was greater southwards and this was probably manifested through naturally greater annual macrophyte abundance in warmer locations in consequence of the longer plant growing‐season. There was no trend in the relative importance of fish effects with latitude but this may partly be an artefact of the simple fish community used. These findings suggest that nutrient control should be a greater priority than biomanipulation in the restoration of eutrophicated shallow lakes in warm temperate regions. 6. Starting conditions affected the outcome of experiments. High initial concentrations of total phosphorus and planktonic chlorophyll a concentration (created by local conditions prior to the experiment) led to de‐emphasis of the importance of nutrient loading in the experiment.     

The plant traits that drive ecosystems: Evidence from three continents

Question: A set of easily‐measured (‘soft’) plant traits has been identified as potentially useful predictors of ecosystem functioning in previous studies. Here we aimed to discover whether the screening techniques remain operational in widely contrasted circumstances, to test for the existence of axes of variation in the particular sets of traits, and to test for their links with ‘harder’ traits of proven importance to ecosystem functioning. Location: central‐western Argentina, central England, northern upland Iran, and north‐eastern Spain. Recurrent patterns of ecological specialization: Through ordination of a matrix of 640 vascular plant taxa by 12 standardized traits, we detected similar patterns of specialization in the four floras. The first PCA axis was identified as an axis of resource capture, usage and release. PCA axis 2 appeared to be a size‐related axis. Individual PCA for each country showed that the same traits remained valuable as predictors of resource capture and utilization in all of them, despite their major differences in climate, biogeography and land‐use. The results were not significantly driven by particular taxa: the main traits determining PCA axis 1 were very similar in eudicotyledons and monocotyledons and Asteraceae, Fabaceae and Poaceae. Links between recurrent suites of ‘soft’ traits and ‘hard’ traits: The validity of PCA axis 1 as a key predictor of resource capture and utilization was tested by comparisons between this axis and values of more rigorously established predictors (‘hard’ traits) for the floras of Argentina and England. PCA axis 1 was correlated with variation in relative growth rate, leaf nitrogen content, and litter decomposition rate. It also coincided with palatability to model generalist herbivores. Therefore, location on PCA axis 1 can be linked to major ecosystem processes in those habitats where the plants are dominant. Conclusion: We confirm the existence at the global scale of a major axis of evolutionary specialization, previously recognised in several local floras. This axis reflects a fundamental trade‐off between rapid acquisition of resources and conservation of resources within well‐protected tissues. These major trends of specialization were maintained across different environmental situations (including differences in the proximate causes of low productivity, i.e. drought or mineral nutrient deficiency). The trends were also consistent across floras and major phylogenetic groups, and were linked with traits directly relevant to ecosystem processes.     

15-Hydroxy-acetylerioflorin and other constituents from Viguiera linearis

Aerial parts of Viguiera linearis afforded 16α-hydroxy-ent-kauranoic acid, viguiestenin, leptocarpin, acetylleptocarpin, budlein B, clovandiol and the new heliangolide 15-hydroxy-acetylerioflorin.     

Terpenoids and a flavan-3-ol from viguiera quinqueradiata

The isolation is reported of the new natural products from Viguiera quinqueradiata, acetylleptocarpin and (2R,3S-4′-hydroxy-3′,5,7-tri-O-methyl-flavan-3-ol. The diterpenes 15α-angeloyloxy-ent-kaur-16-en-19-oic acid, 15α-tigloyloxy-ent-kaur-16-en-19-oic acid and the sesquiterpene lactones leptocarpin and budlein A were also found.     

Physiological presynaptic facilitation of dopamine release in the cat caudate nucleus

Using halothane-anaesthetized cats implanted with push-pull cannulae, we investigated the effects of GABA application into the VM/VL on the release of [3H] DA continuously synthetized from [3H] tyrosine in the ipsilateral CN and SN and on single unit activity of nigral DA cells. GABA was applied (30 min) at a concentration of 10(-3) or 10(-5) M since the higher concentration reduces the local multi-unit activity in the VM/VL while the opposite response is observed with the lower one. The application of GABA into the VM/VL at a concentration of 10(-3) M resulted in an increase in nigral [3H] DA release, an inhibition of DA cell firing and a decrease in [3H] DA release in the CN. The latter effect is likely due to the inhibition of DA neuron activity which is triggered through DA autoreceptors by DA released from dendrites. In contrast, when applied at a concentration of 10(-5) M into the VM/VL, GABA stimulated [3H] DA release in the CN despite its inhibitory effect on single unit activity of DA cells. Furthermore, the nigral release of [3H] DA was no longer affected. These results indicated that DA release from nerve terminals was not dependent on nerve activity and they favour the existence of a potent facilitatory presynaptic regulation of DA release. The intervention of a presynaptic mechanism was further established by examining the effect of GABA (10(-5) M) application into the VM/VL on [3H] DA release in the CN shortly after a complete ipsilateral hemisection of the brain made at the meso-diencephalic level.(ABSTRACT TRUNCATED AT 250 WORDS)