Leishmania Promastigotes Lack Phosphatidylserine but Bind Annexin V upon Permeabilization or Miltefosine Treatment

The protozoan parasite Leishmania is an intracellular pathogen infecting and replicating inside vertebrate host macrophages. A recent model suggests that promastigote and amastigote forms of the parasite mimic mammalian apoptotic cells by exposing phosphatidylserine (PS) at the cell surface to trigger their phagocytic uptake into host macrophages. PS presentation at the cell surface is typically analyzed using fluorescence-labeled annexin V. Here we show that Leishmania promastigotes can be stained by fluorescence-labeled annexin V upon permeabilization or miltefosine treatment. However, combined lipid analysis by thin-layer chromatography, mass spectrometry and (31)P nuclear magnetic resonance (NMR) spectroscopy revealed that Leishmania promastigotes lack any detectable amount of PS. Instead, we identified several other phospholipid classes such phosphatidic acid, phosphatidylethanolamine; phosphatidylglycerol and phosphatidylinositol as candidate lipids enabling annexin V staining.     

Achieving High Accuracy Prediction of Minimotifs

The low complexity of minimotif patterns results in a high false-positive prediction rate, hampering protein function prediction. A multi-filter algorithm, trained and tested on a linear regression model, support vector machine model, and neural network model, using a large dataset of verified minimotifs, vastly improves minimotif prediction accuracy while generating few false positives. An optimal threshold for the best accuracy reaches an overall accuracy above 90%, while a stringent threshold for the best specificity generates less than 1% false positives or even no false positives and still produces more than 90% true positives for the linear regression and neural network models. The minimotif multi-filter with its excellent accuracy represents the state-of-the-art in minimotif prediction and is expected to be very useful to biologists investigating protein function and how missense mutations cause disease.     

Location-dependent effects of inhibition on local spiking in pyramidal neuron dendrites

Cortical computations are critically dependent on interactions between pyramidal neurons (PNs) and a menagerie of inhibitory interneuron types. A key feature distinguishing interneuron types is the spatial distribution of their synaptic contacts onto PNs, but the location-dependent effects of inhibition are mostly unknown, especially under conditions involving active dendritic responses. We studied the effect of somatic vs. dendritic inhibition on local spike generation in basal dendrites of layer 5 PNs both in neocortical slices and in simple and detailed compartmental models, with equivalent results: somatic inhibition divisively suppressed the amplitude of dendritic spikes recorded at the soma while minimally affecting dendritic spike thresholds. In contrast, distal dendritic inhibition raised dendritic spike thresholds while minimally affecting their amplitudes. On-the-path dendritic inhibition modulated both the gain and threshold of dendritic spikes depending on its distance from the spike initiation zone. Our findings suggest that cortical circuits could assign different mixtures of gain vs. threshold inhibition to different neural pathways, and thus tailor their local computations, by managing their relative activation of soma- vs. dendrite-targeting interneurons.     

Bioluminescence resonance energy transfer assays reveal ligand-specific conformational changes within preformed signaling complexes containing delta-opioid receptors and heterotrimeric G proteins

Heptahelical receptors communicate extracellular information to the cytosolic compartment by binding an extensive variety of ligands. They do so through conformational changes that propagate to intracellular signaling partners as the receptor switches from a resting to an active conformation. This active state has been classically considered unique and responsible for regulation of all signaling pathways controlled by a receptor. However, recent functional studies have challenged this notion and called for a paradigm where receptors would exist in more than one signaling conformation. This study used bioluminescence resonance energy transfer assays in combination with ligands of different functional profiles to provide in vivo physical evidence of conformational diversity of delta-opioid receptors (DORs). DORs and alpha(i1)beta(1)gamma(2) G protein subunits were tagged with Luc or green fluorescent protein to produce bioluminescence resonance energy transfer pairs that allowed monitoring DOR-G protein interactions from different vantage points. Results showed that DORs and heterotrimeric G proteins formed a constitutive complex that underwent structural reorganization upon ligand binding. Conformational rearrangements could not be explained by a two-state model, supporting the idea that DORs adopt ligand-specific conformations. In addition, conformational diversity encoded by the receptor was conveyed to the interaction among heterotrimeric subunits. The existence of multiple active receptor states has implications for the way we conceive specificity of signal transduction.     

Cyclic enkephalin analogs containing various para-substituted phenylalanine derivatives in place of Tyr1 are potent opioid agonists.

The cyclic enkephalin analog H-Tyr-c[D-Cys-Gly-Phe(pNO(2))-D-Cys]NH(2) is a highly potent opioid agonist with IC(50)s of 35 pm and 19 pm in the guinea-pig ileum (GPI) and mouse vas deferens (MVD) assays, respectively. The Phe(1)-analog of this peptide showed 370-fold and 6790-fold lower agonist potency in the GPI and MVD assays, respectively, indicating the importance of the Tyr(1) hydroxyl-group in the interaction with mu and delta opioid receptors. In the present study, the effect of various substituents (-NH(2), -NO(2), -CN, -CH(3), -COOH, -COCH(3), -CONH(2)) introduced in the para-position of the Phe(1)-residue of H-Phe-c[D-Cys-Gly-Phe(pNO(2))-D-Cys]NH(2) on the in vitro opioid activity profile was examined. Most analogs showed enhanced mu and delta agonist potencies in the two bioassays, except for the Phe(pCOOH)(1)-analog, which was weakly active, probably as a consequence of the negative charge. The most potent compounds were the Phe(pCOH(3))(1)- and the Phe(pCONH(2))(1)-analogs. The latter compound showed subnanomolar mu and delta agonist potencies and represents the most potent enkephalin analog lacking the Tyr(1) hydroxyl-group reported to date. Taken together, these results indicate that various substituents introduced in the para-position of Phe(1) enhance opioid activity via hydrogen bonding or hydrophobic interactions with the receptor. Comparison with existing structure-activity relationship on phenolic hydroxyl replacements in morphinans indicates that these nonpeptide opiates and some of the cyclic enkephalin analogs described here may have different modes of binding to the receptor.     

Causes of death in hemophilia patients in East Germany

By means of the Central Card Index of Haemophilia, a working material of the Section of Haemophilia in the Society of Haematology and Blood Transfusion of the GDR, 52 patients with heriditary haemorrhagic diatheses could be registered. These persons had died between 1962 and 1984. The average age of death amounted to 33.8 years. With 23 cases intracranial bleedings were the dominating cause of death in all haemophiliacs. From this fact the absolute necessity arises to initiate a substitutive therapy with a sufficiently high dosage, if there is an assumption of a craniocerebral trauma and, in addition, to treat hypertension in such a way that normotonic values are achieved. Even retrospectively no signs of an acquired immunodeficiency syndrome could be found in any of the patients. As haemophilic bleedings apparently represented the life limiting factors in the majority of the deceased, the importance of an early adequate substitutive therapy is stressed, the principles of which should be known to all general practioners.     

Zur Frage der Gezeitenwirkung auf die emergierenden Algen

Zusammenfassung Fucus virsoides lebt im Adriatischen Meere vom Triester Golfe an bis Ragusa, wird aber gegen Süden immer kleiner und zeigt auch anatomische Veränderungen.Diese morphologisch-anatomischen Unterschiede der Nord- und Südpflanzen sind durch die von Norden (durchschnittlich 60 cm) gegen die mittlere Adria (etwa 10–20 cm) abnehmenden Gezeitenunterschiede bedingt, wobei die Pflanze immer kürzere Zeit, oft tagelang überhaupt nicht, aus dem Wasser auftaucht.Da die Fluthöhe von der mittleren gegen die südliche Adria wieder zunimmt, so kann das Aufhören derFucus-Vegetation beiRagusa nicht durch die Gezeiten bedingt sein. Die Ursache ist die hohe mittlere Jahrestemperatur des Wassers. Denn bis dorthin reicht der erwärmende Einfluß des einströmenden subtropischen Mittelmeerwassers, was die nur bis in die mittlere Adria vom Süden vordringenden tropischen und subtropischen Planktonorganismen ersichtlich machen.Die GattungFucus ist an kaltes bzw. temperiertes Wasser gebunden.     

Histopathology of hepatic and pulmonary granulomata experimentally induced with eggs of Schistosoma mansoni

Hepatic granulomata were experimentally induced in previously unexposed white mice, albino rats and golden hamsters by injecting viable exogenous eggs of Schistosoma mansoni via the mesenterico-portal system. Histopathologic studies of livers of these animals showed that the lesions were similar to those in infections resulting form exposure to cercariae as occurs naturally in Mansonian schistosomiasis. Comparable observations made of the lungs of animals that had received egg injections via their tail veins, showed striking differences with respect to timing of the occurrence of various histopathologic stages, mean size of granulomata, cellular composition and pathologic manifestations.     

Immunotyping of freshly isolated strains of Pseudomonas aeruginosa

During 1972-1982 the bacteriological study of 1391 patients with thermal burns was carried out. As the result of clinico-bacteriological studies, the occurrence of P. aeruginosa was found to increase from 39.3% to 70.5% during this period. The immunotyping of P. aeruginosa cultures isolated in 3 burn-treatment centers showed that strains belonging to immunotypes 2, 3, 7 and 3/7 were most frequently isolated from burn wounds. These strains were found to be the cause of hospital infections in burn-treatment hospitals. In connection with the data thus obtained immunological preparations intended for the prophylaxis and treatment of P. aeruginosa infection should include P. aeruginosa strains, immunotypes 2, 3, 7 and 3/7.