Transport of 1-kestose across the tonoplast of Jerusalem artichoke tubers

The capacity for 1-kestose uptake into the vacuole of fructan storing Jerusalem artichoke tubers was investigated. 1-kestose serves both as building block for fructan initiation and as a fructose donor for chain elongation. Tonoplast vesicles were isolated from actively storing tubers, and their vesicles were capable of transporting sucrose in a manner indicative of a sucrose/H(+) antiport. Under similar conditions, 1-kestose was not taken up by vesicles energized by either a pH jump or in the presence of ATP. When added together at 2 mM, sucrose uptake was not affected by the presence of 1-kestose. The data argues against the possible synthesis of 1-kestose in the cytosol and subsequent transport to the vacuole. The data also presents definite evidence for the existence a mechanism for sucrose accumulation in fructan storing vacuoles.     

Electrolocation in the platypus--some speculations

In the platypus, electroreceptors are located in rostro-caudal rows in skin of the bill, while mechanoreceptors are uniformly distributed across the bill. The electrosensory area of the cerebral cortex is contained within the tactile somatosensory area, and some cortical cells receive input from both electroreceptors and mechanoreceptors, suggesting a close association between the tactile and electric senses. Platypus can determine the direction of an electric source, perhaps by comparing differences in signal strength across the sheet of electroreceptors as the animal characteristically moves its head from side to side while hunting. The cortical convergence of electrosensory and tactile inputs suggests a mechanism for determining the distance of prey items which, when they move, emit both electrical signals and mechanical pressure pulses. Distance could be computed from the difference in time of arrival of the two signals. Much of the platypus' feeding is done by digging in the bottom of streams with the bill. Perhaps the electroreceptors could also be used to distinguish animate and inanimate objects in this situation where the mechanoreceptors would be continuously stimulated. Much of this is speculation, and there is still much to be learned about electroreception in the platypus and its fellow monotreme, the echidna.     

Die Flora von Kremsier in Mähren

Ohne Zusammenfassung     

Cardiomyopathy Mutations in the Tail of β-Cardiac Myosin Modify the Coiled-coil Structure and Affect Integration into Thick Filaments in Muscle Sarcomeres in Adult Cardiomyocytes

It is unclear why mutations in the filament-forming tail of myosin heavy chain (MHC) cause hypertrophic or dilated cardiomyopathy as these mutations should not directly affect contraction. To investigate this, we first investigated the impact of five hypertrophic cardiomyopathy-causing (N1327K, E1356K, R1382W, E1555K, and R1768K) and one dilated cardiomyopathy-causing (R1500W) tail mutations on their ability to incorporate into muscle sarcomeres in vivo. We used adenoviral delivery to express full-length wild type or mutant enhanced GFP-MHC in isolated adult cardiomyocytes. Three mutations (N1327K, E1356K, and E1555K) reduced enhanced GFP-MHC incorporation into muscle sarcomeres, whereas the remainder had no effect. No mutations significantly affected contraction. Fluorescence recovery after photobleaching showed that fluorescence recovery for the mutation that incorporated least well (N1327K) was significantly faster than that of WT with half-times of 25.1 ± 1.8 and 32.2 ± 2.5 min (mean ± S.E.), respectively. Next, we determined the effects of each mutation on the helical properties of wild type and seven mutant peptides (7, 11, or 15 heptads long) from the myosin tail by circular dichroism. R1382W and E1768K slightly increased the α-helical nature of peptides. The remaining mutations reduced α-helical content, with N1327K showing the greatest reduction. Only peptides containing residues 1301–1329 were highly α-helical suggesting that this region helps in initiation of coiled coil. These results suggest that small effects of mutations on helicity translate into a reduced ability to incorporate into sarcomeres, which may elicit compensatory hypertrophy.     

Estimating Urban Traffic Patterns through Probabilistic Interconnectivity of Road Network Junctions

The emergence of large, fine-grained mobility datasets offers significant opportunities for the development and application of new methodologies for transportation analysis. In this paper, the link between routing behaviour and traffic patterns in urban areas is examined, introducing a method to derive estimates of traffic patterns from a large collection of fine-grained routing data. Using this dataset, the interconnectivity between road network junctions is extracted in the form of a Markov chain. This representation encodes the probability of the successive usage of adjacent road junctions, encoding routes as flows between decision points rather than flows along road segments. This network of functional interactions is then integrated within a modified Markov chain Monte Carlo (MCMC) framework, adapted for the estimation of urban traffic patterns. As part of this approach, the data-derived links between major junctions influence the movement of directed random walks executed across the network to model origin-destination journeys. The simulation process yields estimates of traffic distribution across the road network. The paper presents an implementation of the modified MCMC approach for London, United Kingdom, building an MCMC model based on a dataset of nearly 700000 minicab routes. Validation of the approach clarifies how each element of the MCMC framework contributes to junction prediction performance, and finds promising results in relation to the estimation of junction choice and minicab traffic distribution. The paper concludes by summarising the potential for the development and extension of this approach to the wider urban modelling domain.     

Isolation and characterization of nuclear genes coding for subunits of the yeast ubiquinol-cytochrome c reductase complex

Nuclear genes coding for the M_r 17000, 14000 and 11000 subunits of the ubiquinol-cytochrome c reductase complex (complex III) in yeast have been isolated from a clone bank of yeast nuclear DNA by use of a mRNA hybridization-competition assay. This is based on our observations that levels of mRNAs for these subunits are much reduced during glucose repression and in cytoplasmic petite mutants and the procedure should be of general application for the isolation of other inducible or repressible genes coding for mRNAs present at low levels in the cell. A first characterization of the clones is presented. The genes are not closely linked in the genome and those coding for M_r 14000 and 11000 subunits are present in unique genomic environments, which suggests that there are only single copies of each in the nuclear genome.     

Molecular basis for the functional interaction of dynein light chain with the nuclear-pore complex

Nucleocytoplasmic transport occurs through nuclear pore complexes (NPCs) embedded in the nuclear envelope. Here, we discovered an unexpected role for yeast dynein light chain (Dyn2) in the NPC. Dyn2 is a previously undescribed nucleoporin that functions as molecular glue to dimerize and stabilize the Nup82-Nsp1-Nup159 complex, a module of the cytoplasmic pore filaments. Biochemical analyses showed that Dyn2 binds to a linear motif (termed DID(Nup159)) inserted between the Phe-Gly repeat and coiled-coil domain of Nup159. Electron microscopy revealed that the reconstituted Dyn2-DID(Nup159) complex forms a rigid rod-like structure, in which five Dyn2 homodimers align like 'pearls on a string' between two extented DID(Nup159) strands. These findings imply that the rigid 20 nm long Dyn2-DID(Nup159) filament projects the Nup159 Phe-Gly repeats from the Nup82 module. Thus, it is possible that dynein light chain plays a role in organizing natively unfolded Phe-Gly repeats within the NPC scaffold to facilitate nucleocytoplasmic transport.     

pH-induced conformational change of the rotavirus VP4 spike: implications for cell entry and antibody neutralization

The rotavirus spike protein, VP4, is a major determinant of infectivity and neutralization. Previously, we have shown that trypsin-enhanced infectivity of rotavirus involves a transformation of the VP4 spike from a flexible to a rigid bilobed structure. Here we show that at elevated pH the spike undergoes a drastic, irreversible conformational change and becomes stunted, with a pronounced trilobed appearance. These particles with altered spikes, at a normal pH of 7.5, despite the loss of infectivity and the ability to hemagglutinate, surprisingly exhibit sialic acid (SA)-independent cell binding in contrast to the SA-dependent cell binding exhibited by native virions. Remarkably, a neutralizing monoclonal antibody that remains bound to spikes throughout the pH changes (pH 7 to 11 and back to pH 7) completely prevents this conformational change, preserving the SA-dependent cell binding and hemagglutinating functions of the virion. A hypothesis that emerges from the present study is that high-pH treatment triggers a conformational change that mimics a post-SA-attachment step to expose an epitope recognized by a downstream receptor in the rotavirus cell entry process. This process involves sequential interactions with multiple receptors, and the mechanism by which the antibody neutralizes is by preventing this conformational change.     

Epitope mapping of a single repetitive unit of the B13 Trypanosoma cruzi antigen as fusions to Escherichia coli LamB protein

B13, one of the immunodominant antigens of Trypanosoma cruzi, is composed of repeats of a 12-amino-acid motif. Using synthetic peptides, the sequence FGQAAAGDK was previously shown to contain the B13 immunodominant epitope recognized by chagasic patients sera. To investigate the effects of neighboring sequences in the immunodominance, we tested serum recognition of two B13 sequences fused to LamB. GDKPSPFGQAAA-LamB and FGQAAAGDKPSP-LamB were recognized, respectively, by 15% and 80% of 80 sera reactive to B13 antigen. Recognition of FGQAAAGDKPSP-LamB was inhibited by AAAGDK-containing synthetic peptides. FGQAAAGDKPSP-LamB competed with a B13 recombinant protein containing 16.6 repeats for binding to chagasic antibodies. These results strengthen previous conclusions on the immunodominant epitope of B13 and provide a comparison of two methods for epitope mapping.