Chromosomes move poleward in anaphase along stationary microtubules that coordinately disassemble from their kinetochore ends

The binding sites on the nicotinic acetylcholine receptor of labels specific for the alpha-, beta-, and delta-subunits were determined by electron image analysis, using tubular crystals of receptors grown from the postsynaptic membranes of Torpedo marmorata electric organ. The labels were alpha-bungarotoxin (which attaches to the acetylcholine binding sites on the pair of alpha-subunits), Fab35 (a monoclonal antibody Fab fragment directed against the main immunogenic region of the alpha-subunit), Fab111 (a monoclonal antibody Fab fragment directed against a cytoplasmic site on the beta-subunit), and wheat germ agglutinin (which binds to N-acetylglucosamine residues on the delta-subunit). These labels, bound to receptors in the crystals, were located by comparing labeled with native structures, averaged in each case over more than 5,000 molecules. From the assignments made, we find that the clockwise arrangement of subunits around the receptor, viewed from the synaptic face, is: alpha, beta, alpha, gamma, and delta; that the main immunogenic region is at (or close to) the side of the alpha-subunit; and that the two acetylcholine binding sites are at the synaptic end of the alpha-subunits, 27-28 A from the central axis and approximately 53 A apart. In the crystal lattice, neighboring molecules are paired so that their delta- and alpha-subunits are juxtaposed, an organization that appears to relate closely to the grouping of receptors in vivo.     

Photosynthesis in an Australian rainforest tree,Argyrodendron peralatum,during the rapid development and relief of water deficits in the dry season

Summary Rates of apparent photosynthesis were measured in situ at five positions between the upper crown and a lower branch of a 34 m tall Argyrodendron peralatum (F.M. Bailey) H.L. Edlin ex I.H. Boas tree, and on an understorey sapling of the same species growing in a northern Australian rainforest. At the end of the dry season, rapid reductions in photosynthetic rates occurred in the upper crown within three days after a rain event, but changes in the lower crown and the sapling were less marked. Complete recovery of photosynthesis followed a second rain event. At high photon flux densities, stomatal conductance to water vapour decreased in a curvilinear fashion as the vapour pressure difference between leaf and air increased. Apparent photosynthesis was linearly related to stomatal conductance on the first clear day after each rain event, but there was no relationship between these parameters at the end of a brief natural drying cycle. Under conditions of adequate water supply, stomatal conductances of both upper crown and understorey leaves increased linearly with increasing photon flux density up to about 300 mol m^-2 s^-1. During water deficits, stomatal conductances in leaves from the understorey increased much more rapidly at very low photon flux densities than did conductances in leaves from the upper canopy.     

Properties of microtubule sliding disintegration in isolated tetrahymena cilia

Properties of the sliding disintegration response of demembranated tetrahymena cilia have been studied by measuring the spectrophotomeric response or turbidity of cilia suspensions at a wavelength of 350 nm relative to changes in the dynein substrate (MgATP(2-)) concentration. The maximum decrease in turbidity occurs in 20 muM ATP, and 90 percent of the decrease occurs in approximately 5.9 s. At lower ATP concentrations (1-20 muM), both the velocity and magnitude of the turbidity decreases are proportional to ATP concentration. The velocity data for 20 muM ATP permit construction of a reaction velocity curve suggesting that changes in turbidity are directly proportional to the extent and velocity of disintegration. At ATP concentrations more than 20 muM (50muM to 5mM), both velocity and magnitude of the turbidimetric response are reduced by approximately 50 percent. This apparent inhibition results in a biphasic response curve that may be related to activation of residual shear resistance or regulatory components at the higher ATP concentrations. The inhibitory effects of elevated ATP can be eliminated by mild trypsin proteolysis, whereupon the reaction goes to completion at any ATP concentration. The turbidimetric responses of the axoneme-substrate suspensions are consistent with the extent and type of axoneme disintegration revealed by electron microscope examination of the various suspensions, suggesting that the turbidimetric assay may prove to be a reliable means for assessing the state of axoneme integrity.     

Field analyses of insect heat budgets: Reflectance,size and heating rates

Summary This paper outlines simple techniques for determining rates of heat gain and loss in relation to the weight and reflectance of insects caught in their natural habitats. In particular the construction of a new reflectometer is described. The results thus obtained permit estimates of the relative importance of size and colour in determining rates of heat exchange and temperature excesses, so allowing better predictions of heat budgets for a given species.     

Electron microscopy of the stacked disk aggregate of tobacco mosaic virus protein. I. Three-dimensional image reconstruction

The three-dimensional structure of the stacked disk aggregate of tobacco mosaic virus protein has been determined from “phase plate” electron micrographs to an effective resolution of about 12 Å. It is a long rod comprised of paired rings of protein (disks), the subunits of which have different conformations according to which ring they belong. The two subunit conformations are such that the rings come close together within a disk near the outer surface of the particle, but between disks on the inside. This property, interpreted on the basis of a polar packing of the subunits, was established from an earlier, lower resolution, study by Finch &; Klug (1971). The present study shows, in addition, that the pairing is contributed mainly by axial distortions of the subunits in one of the rings, the axial distortions of the subunits in the other being largely replaced at lower radii by a tilt or twist and, at higher radii, by a slew. The subunits in the latter ring appear to have a conformation similar to that of the protein molecules in the virus.     

High lift function of the pteroid bone and forewing of pterosaurs

The pteroid bone is a rod-like element found only in pterosaurs, the flying reptiles of the Mesozoic. It articulated at the wrist, and supported a membranous forewing in front of the inner part of the wing spar. The function of this bone, particularly its orientation, has been much debated. It is widely believed that it pointed towards the body, and that the forewing was relatively narrow. An alternative hypothesis states that it was directed forwards during flight, resulting in a much broader forewing that acted as a leading edge flap. We tested scale models in a wind tunnel to determine the aerodynamic consequences of these conflicting hypotheses, and found that performance is greatly improved if the pteroid is directed forwards: the lift: drag ratios are superior and the maximum lift is exceptionally high in comparison with conventional aerofoils. This high lift capability may have enabled even the largest pterosaurs to take off and land without difficulty.     

Heavy metal poisoning: the effects of cadmium on the kidney

The heavy metal cadmium (Cd) is known to be a widespread environmental contaminant and a potential toxin that may adversely affect human health. Exposure is largely via the respiratory or gastrointestinal tracts; important non-industrial sources of exposure are cigarette smoke and food (from contaminated soil and water). The kidney is the main organ affected by chronic Cd exposure and toxicity. Cd accumulates in the kidney as a result of its preferential uptake by receptor-mediated endocytosis of freely filtered and metallothionein bound Cd (Cd-MT) in the renal proximal tubule. Internalised Cd-MT is degraded in endosomes and lysosomes, releasing free Cd²⁺ into the cytosol, where it can generate reactive oxygen species (ROS) and activate cell death pathways. An early and sensitive manifestation of chronic Cd renal toxicity, which can be useful in individual and population screening, is impaired reabsorption of low molecular weight proteins (LMWP) (also a receptor-mediated process in the proximal tubule) such as retinol binding protein (RBP). This so-called ‘tubular proteinuria’ is a good index of proximal tubular damage, but it is not usually detected by routine clinical dipstick testing for proteinuria. Continued and heavy Cd exposure can progress to the clinical renal Fanconi syndrome, and ultimately to renal failure. Environmental Cd exposure may be a significant contributory factor to the development of chronic kidney disease, especially in the presence of other co-morbidities such as diabetes or hypertension; therefore, the sources and environmental impact of Cd, and efforts to limit Cd exposure, justify more attention.     

Simultaneous analysis of relative protein expression levels across multiple samples using iTRAQ isobaric tags with 2D nano LC-MS/MS

In this paper, we describe the use of iTRAQ (isobaric Tags for Relative and Absolute Quantitation) tags for comparison of protein expression levels between multiple samples. These tags label all peptides in a protein digest before labeled samples are pooled, fractionated and analyzed using mass spectrometry (MS). As the tags are isobaric, the intensity of each peak is the sum of the intensity of this peptide from all samples, providing a moderate enhancement in sensitivity. On peptide fragmentation, amino-acid sequence ions also show this summed intensity, providing a sensitivity enhancement. However, the distinct distribution of isotopes in the tags is such that, on further fragmentation, a tag-specific reporter ion is released. The relative intensities of these ions represent the relative amount of peptide in the analytes. Integration of the relative quantification data for the peptides allows relative quantification of the protein. This protocol discusses the rationale behind design, optimization and performance of experiments, comparing protein samples using iTRAQ chemistries combined with strong cation exchange chromatographic fractionation and MS.