Electron microscopic analysis of the modulation of lymphocyte receptor mobility

The topographic distribution of ferritin-labelled concanavalin A (FT-ConA) bound to the surface membrane of mouse lymphocytes has been analysed by examining ultrathin sections and ghost membranes in the electron microscope. Binding of FT-ConA to lymphocytes at 37 °C did not induce redistribution of ConA receptors at any dose of the labelled lectin. In contrast, in cells treated with colchicine, FT-ConA induced both patch and cap formation. These findings suggest that the modulation of receptor mobility by colchicine occurs at the level of individual receptors. Ultrastructural observations revealed both microtubules and microfilaments in lymphocytes consistent with the suggestion that these cellular structures may be involved in the control of receptor mobility. Analyses using freeze-fracture methods indicated that the distribution of intramembranous particles is not correlated with either the movement of surface receptors or the modulation events. An hypothesis on the modulation of surface receptors by cytoplasmic structures is proposed on the basis of these and previous observations.     

Class similarity and viewpoint invariance in the recognition of 3D objects

 In human vision, the processes and the representations involved in identifying specific individuals are frequently assumed to be different from those used for basic level classification, because classification is largely viewpoint-invariant, but identification is not. This assumption was tested in psychophysical experiments, in which objective similarity between stimuli (and, consequently, the level of their distinction) varied in a controlled fashion. Subjects were trained to discriminate between two classes of computer-generated three-dimensional objects, one resembling monkeys and the other, dogs. Both classes were defined by the same set of 56 parameters, which encoded sizes, shapes, and placement of the limbs, ears, snout, etc. Interpolation between parameter vectors of the class prototypes yielded shapes that changed smoothly between monkey and dog. Within-class variation was induced in each trial by randomly perturbing all the parameters. After the subjects reached 90% correct performance on a fixed canonical view of each object, discrimination performance was tested for novel views that differed by up to 60° from the training view. In experiment 1 (in which the distribution of parameters in each class was unimodal) and in experiment 2 (bimodal classes), the stimuli differed only parametrically and consisted of the same geons (parts), yet were recognized virtually independently of viewpoint in the low-similarity condition. In experiment 3, the prototypes differed in their arrangement of geons, yet the subjects’ performance depended significantly on viewpoint in the high-similarity condition. In all three experiments, higher interstimulus similarity was associated with an increase in the mean error rate and, for misorientation of up to 45°, with an increase in the degree of viewpoint dependence. These results suggest that a geon-level difference between stimuli is neither strictly necessary nor always sufficient for viewpoint-invariant performance. Thus, basic and subordinate-level processes in visual recognition may be more closely related than previously thought. Received: 15 November 1993/Accepted in revised form: 14 July 1994     

Involvement of Thylakoid Overenergization in Tentoxin-Induced Chlorosis in Nicotiana spp

The purpose of this work was to clarify the mechanism of tentoxin-induced chlorosis in Nicotiana spp. seedlings. We found that chlorosis does not correlate with the inhibition of chloroplast ATP synthesis in vivo, since it occurs at tentoxin concentrations far higher than that required for the inhibition of photophosphorylation measured in the same seedlings. However, tentoxin-induced chlorosis does correlate with in vivo overenergization of thylakoids. We show that tentoxin induces overenergization in intact plants and isolated thylakoids, probably via multiple interactions with ATP synthase. Furthermore, gramicidin D, a protonophore that relieves overenergization, also relieves chlorosis. Two lines of evidence suggest that reactive oxygen species may be involved in the process of chlorosis: ascorbate, a quencher of oxygen radicals, significantly protects against chlorosis, whereas transgenic Nicotiana spp. mutants overexpressing chloroplast superoxide dismutase are partially resistant to tentoxin-induced chlorosis. It is proposed that chlorosis in developing seedlings results from overenergization of thylakoids, which leads to the generation of oxygen radicals.     

Thermal denaturation of nucleic acids in polyacrylamide gels

A system is described for measuring thermal denaturation of nucleic acid fractions directly in polyacrylamide gels. Total nucleic acids were fractionated by disc gel electrophoresis. The buffer within the gel was then exchanged for one commonly used in denaturation studies. Thermal denaturation profiles of DNA and ribosomal RNA in the gel were determined using a specially constructed Gel Carriage to position the appropriate fraction during spectrophotometric measurements. These profiles were compared with denaturation patterns obtained by classical methods in free solution; the two methods yielded similar patterns.Thermal denaturation profiles were also obtained for chloroplast light ribosomal RNA resolved by gel electrophoresis of total plant nucleic acids. Thus, denaturation patterns of individual, minor components present in complex nucleic acid mixtures can be directly measured in gels.     

CHEMICAL DISSECTION OF MAMMALIAN SPERMATOZOA

Spermatozoa from several mammalian species have been dissected by chemical methods to yield free heads, tails with attached midpieces, and tails from which the mitochondrial components of the midpiece were removed. Mouse and rat spermatozoa were cleaved by brief treatment with trypsin to yield free heads and tails, while human, guinea pig, and rabbit spermatozoa were cleaved by trypsin only after incubation with 2-mercaptoethanol or dithiothreitol. Spermatozoa were also cleaved at the junction of the head and the tail by treatment with acid and base. Mitochondria were removed from intact spermatozoa or isolated tails by mechanical shear after treatment with 2-mercaptoethanol or dithiothreitol. The dissected components of spermatozoa were fractionated with good yield and high purity by density gradient centrifugation. Ultrastructural analysis indicates that proteolytic cleavage to yield separated heads and tails occurs at a specific location in the neck of the spermatozoon, leaving the basal plate attached to the head of the cell. In contrast, after acid cleavage the basal plate remains with the midpiece. Proteolytic treatment has no apparent effect on any other spermatozoan structures, whereas acid or base treatment results in damage to the plasma membrane, the acrosome, and other structures. The specificity of the proteolytic cleavage suggests that a particular protein or group of proteins may be responsible for the linkage between the sperm head and tail.     

Molecular docking using surface complementarity

A method is described to dock a ligand into a binding site in a protein on the basis of the complementarity of the inter-molecular atomic contacts. Docking is performed by maximization of a complementarity function that is dependent on atomic contact surface area and the chemical properties of the contacting atoms. The generality and simplicity of the complementarity function ensure that a wide range of chemical structures can be handled. The ligand and the protein are treated as rigid bodies, but displacement of a small number of residues lining the ligand binding site can be taken into account. The method can assist in the design of improved ligands by indicating what changes in complementarity may occur as a result of the substitution of an atom in the ligand. The capabilities of the method are demonstrated by application to 14 protein–ligand complexes of known crystal structure. © 1996 Wiley Liss, Inc.     

Floristic variation and willow carr development within a southwest England wetland

Abstract. Woodland colonization on wetlands is considered to have a detrimental effect on their ecological value, even though detailed analysis of this process is lacking. This paper provides an evaluation of the ecological changes resulting from succession of poor fen (base‐poor mire) to willow wet woodland on Goss Moor NNR in Cornwall, UK. Different ages of willow carr were associated with eight understorey communities. During willow colonization, in the ground flora, there was a progressive decrease in poor fen species and an associated increase in woodland species, which appeared to be related to an increase in canopy cover and therefore shade. The most diverse community was found to be the most recent willow and was dominated by poor fen species. The oldest willow was the second most diverse and was associated with a reduction in poor fen species and an increase in woodland species. Architectural features were used successfully to assess the general condition and structure of willow. Tree height and DBH were identified as useful parameters to accurately assess willow age in the field. The implications of active intervention to remove willow in order to conserve the full range of communities within the hydrosere are discussed.