Quasiparticle chirality in epitaxial graphene probed at the nanometer scale.

Graphene exhibits unconventional two-dimensional electronic properties resulting from the symmetry of its quasiparticles, which leads to the concepts of pseudospin and electronic chirality. Here, we report that scanning tunneling microscopy can be used to probe these unique symmetry properties at the nanometer scale. They are reflected in the quantum interference pattern resulting from elastic scattering off impurities, and they can be directly read from its fast Fourier transform. Our data, complemented by theoretical calculations, demonstrate that the pseudospin and the electronic chirality in epitaxial graphene on SiC(0001) correspond to the ones predicted for ideal graphene.

[Inner ear damage due to leisure and broadband noise. An experimental study on initial and permanent functional and morphological damage]. / Innenohrschäden durch Freizeitlärm und Breitbandrauschen. Eine experimentelle Studie zu initialen und permanenten funktionellen und morphologischen Schäden.

The initial and permanent effects of leisure noise (toy pistols, rock music) compared to broadband noise were examined in 68 guinea pigs. Auditory threshold shifts at 1.5, 2, 3, 4, 6, 8, 12 und 16 kHz were registered before and immediately after exposure as well as on days 1, 2, 3, 5,7 and 21 post-exposure using the auditory brain stem response (ABR) technique. In order to examine cilia and hair cell damage in eight cochlear frequency regions (<0,4 kHz, 0,4-0,8 kHz, 0,8-1.5 kHz, 1.5-3 kHz, 3-5 kHz, 5-11.5 kHz, 11.5-26 kHz und >26 kHz), cytocochleograms were performed immediately after exposure and on days 1, 7 and 21.Frequency dependent functional or morphological damage was found which depended on the type of trauma tested. All results were highly significant ( P<0.001). The results show that partial recovery of hearing occurred within 3 days of acute acoustic trauma induced by toy pistols and within 1 day after exposure to rock music or broadband noise. There was no further recovery of hearing within the following 18 and 20 days, respectively. Furthermore, permanent threshold shifts after exposure to rock music or broadband noise were not associated with cilia and/or hair cell damage.

FliMax, a novel stimulus device for panoramic and highspeed presentation of behaviourally generated optic flow.

A high-speed panoramic visual stimulation device is introduced which is suitable to analyse visual interneurons during stimulation with rapid image displacements as experienced by fast moving animals. The responses of an identified motion sensitive neuron in the visual system of the blowfly to behaviourally generated image sequences are very complex and hard to predict from the established input circuitry of the neuron. This finding suggests that the computational significance of visual interneurons can only be assessed if they are characterised not only by conventional stimuli as are often used for systems analysis, but also by behaviourally relevant input.

A central role for cohesins in sister chromatid cohesion, formation of axial elements, and recombination during yeast meiosis.

A multisubunit complex, called cohesin, containing Smc1p, Smc3p, Scc1p, and Scc3p, is required for sister chromatid cohesion in mitotic cells. We show here that Smc3p and a meiotic version of Scc1p called Rec8p are required for cohesion between sister chromatids, for formation of axial elements, for reciprocal recombination, and for preventing hyperresection of double-strand breaks during meiosis. Both Rec8p and Smc3p colocalize with chromosome cores independently of synapsis during prophase I and largely disappear from chromosome arms after pachytene but persist in the neighborhood of centromeres until the onset of anaphase II. The eukaryotic cell's cohesion apparatus is required both for the repair of recombinogenic lesions and for chromosome segregation and therefore appears to lie at the heart of the meiotic process.

Binding and catalytic properties of the Cdc2 and Crp proteins of Dictyostelium.

Dictyostelium expresses at least two proteins of the cyclin-dependent kinase (Cdk) family, Cdc2 and Crp. Cdc2 levels remain relatively constant during differentiation, whereas the levels of Crp increase dramatically as differentiation progresses. Crp is highly related to the mammalian Cdk5, and p25 (a truncated form of p35, the activating subunit of Cdk5 from mammalian brain) stimulates the histone H1 kinase activity of GST-Crp by several fold. In contrast, p25 does not stimulate the histone H1 kinase activity of GST-Cdc2 or the Cdc2 activity present in cell extracts from vegetative Dictyostelium cells. GST-Cdc2, in vitro translated Cdc2 and Cdc2 from all stages of differentiation bind to p13suc1. In contrast, GST-Crp, in vitro translated Crp and the Crp protein present in cell extracts do not bind to p13suc1. We have confirmed a previous report by Arakane and Maeda [J. Plant Res. (1997) 110, 81-85] that there is a peak of p13suc1 bound histone H1 kinase activity during late development, but we found that there was no corresponding peak of p13suc1 bound Cdc2 protein that corresponds to this activity. Taken together, these data suggest that neither Cdc2 nor Crp is responsible for the late developmental peak of histone H1 kinase activity that binds to p13suc1.

An ESP1/PDS1 complex regulates loss of sister chromatid cohesion at the metaphase to anaphase transition in yeast.

Cohesion between sister chromatids during G2 and M phases depends on the "cohesin" protein Scc1p (Mcd1p). Loss of cohesion at the metaphase to anaphase transition is accompanied by Scc1p's dissociation from chromatids, which depends on proteolysis of Pds1p mediated by a ubiquitin protein ligase called the anaphase promoting complex (APC). We show that destruction of Pds1p is the APC's sole role in triggering Scc1p's dissociation from chromatids and that Pds1p forms a stable complex with a 180 kDa protein called Esp1p, which is essential for the dissociation of Scc1p from sister chromatids and for their separation. We propose that the APC promotes sister separation not by destroying cohesins but instead by liberating the "sister-separating" Esp1 protein from its inhibitor Pds1p.

Cohesins: chromosomal proteins that prevent premature separation of sister chromatids.

Cohesion between sister chromatids opposes the splitting force exerted by microtubules, and loss of this cohesion is responsible for the subsequent separation of sister chromatids during anaphase. We describe three chromosmal proteins that prevent premature separation of sister chromatids in yeast. Two, Smc1p and Smc3p, are members of the SMC family, which are putative ATPases with coiled-coil domains. A third protein, which we call Scc1p, binds to chromosomes during S phase, dissociates from them at the metaphase-to-anaphase transition, and is degraded by the anaphase promoting complex. Association of Scc1p with chromatin depends on Smc1p. Proteins homologous to Scc1p exist in a variety of eukaryotic organisms including humans. A common cohesion apparatus might be used by all eukaryotic cells during both mitosis and meiosis.

Mother cell-specific HO expression in budding yeast depends on the unconventional myosin myo4p and other cytoplasmic proteins.

Certain cell types give rise to progeny that adopt different patterns of gene expression in the absence of any differences in their environment. Cells of budding yeast give birth to mother and daughter cells that differ in that only mother cells express the HO endonuclease gene and thereby switch mating types. We describe the identification of five genes, called SHE1-SHE5, that encode cytoplasmic proteins required for mother-specific HO expression. She1p, which is identical to the minimyosin Myo4p, and She3p are not, however, mother-specific proteins. On the contrary, they accumulate in growing buds. She proteins might be required for the transport of factors that promote HO repression from the mother cell into its bud. In an accompanying paper, we show that SHE genes are needed for the accumulation in daughter nuclei of Ash1p, a repressor of HO.

Cyclin B and Cdc2 expression and Cd2 kinase activity during Dictyostelium differentiation.

Although Dictyostelium differentiation occurs in the absence of external nutrients, two periods of mitosis occur, one during early development and one during the formation of the migrating pseudoplasmodium. We showed previously that cyclin B mRNA levels vary in a cell cycle dependent manner during vegetative cell growth. In the present study, we report that cyclin B mRNA levels change dramatically during development, reaching a maximum at the tipped aggregate stage. However, amounts of cyclin B protein vary only slightly, peaking during early development and decreasing during late aggregation and pseudoplasmodial formation. Cdc2 protein levels also remain relatively constant during development. Cdc2-histone H1 kinase activity was considerably higher in vegetative cell extracts of transformants that expressed large amounts of truncated cyclin B protein in comparison to extracts of the parental Ax-2 cells. These results suggest that Cdc2 kinase activity is dependent upon the level of cyclin B in vegetative cells. This result is consistent with the idea that variations in the level of cyclin B during growth regulate the cell cycle. When Cdc2 histone H1 kinase activity was determined during development, it was also found that activity correlated reasonably well with the amount of cyclin B protein. Thus, there was an increase in Cdc2 histone H1 kinase activity early in development, and then levels decreased as development progressed. The increase in Cdc2 histone H1 kinase activity that occurs early in development following starvation may be important in accelerating G2-phase cells through into mitosis. There was no increase in Cdc2 histone H1 kinase that accompanied the previously reported late developmental mitosis.(ABSTRACT TRUNCATED AT 250 WORDS)

Genes involved in sister chromatid separation are needed for B-type cyclin proteolysis in budding yeast.

B-type cyclin destruction is necessary for exit from mitosis and the initiation of a new cell cycle. Through the isolation of mutants, we have identified three essential yeast genes, CDC16, CDC23, and CSE1, which are required for proteolysis of the B-type cyclin CLB2 but not of other unstable proteins. cdc23-1 mutants are defective in both entering and exiting anaphase. Their failure to exit anaphase can be explained by defective cyclin proteolysis. CDC23 is required at the metaphase/anaphase transition to separate sister chromatids, and we speculate that it might promote proteolysis of proteins that hold sister chromatids together. Proteolysis of CLB2 is initiated in early anaphase, but a fraction of CLB2 remains stable until anaphase is complete.

A Dictyostelium discoideum gene, which is highly related to mo15 from Xenopus, is expressed during growth but not during development.

We have isolated a cDNA from the cellular slime mold Dictyostelium discoideum encoding a protein that is 52% identical to the Xenopus Mo15 kinase and highly related to the equivalent proteins from human (52% identity), rice (52.7% identity), and yeast (47.6% identity). Mo15 is responsible for the activation of Cdc2 kinase and is itself a member of the large Cdc2-related family of protein kinases. The Dictyostelium protein is more related to the Xenopus Mo15 protein than it is to either the Dictyostelium Cdc2 or Crp proteins. Southern blot analysis of genomic V12-M2 DNA indicated that mo15 is present as a single copy gene that cross hybridizes with cdc2 at low stringency. Northern blot analysis of RNA from different stages of Dictyostelium development showed that mo15 is only expressed during vegetative cell growth.

Overexpression of a truncated cyclin B gene arrests Dictyostelium cell division during mitosis.

A cyclin gene has been isolated from Dictyostelium discoideum and the available evidence indicates that the gene encodes a B type cyclin. The cyclin box region of the protein encoded by the gene, clb1, has the highest degree of sequence identity with the B-type cyclins of other species. Levels of cyclin B mRNA and protein oscillate during the cell cycle with maximum accumulation of mRNA occurring prior to cell division and maximum levels of protein occurring during cell division. Overexpression of a N-terminally truncated cyclin B protein lacking the destruction box inhibits cell growth by arresting cell division during mitosis. The gene is present as a single copy in the Dictyostelium genome and there is no evidence for any other highly related cyclin B genes.

The isolation from a unicellular organism, Dictyostelium discoideum, of a highly-related cdc2 gene with characteristics of the PCTAIRE subfamily.

It has been suggested that cell-type determination in Dictyostelium discoideum is dependent on the position of a cell in the cell cycle at the time of starvation. In order to understand the molecular basis of this phenomenon, we initiated studies on the cell cycle and have recently described the isolation of a Dictyostelium gene encoding a homolog of the Cdc2 kinase. We have been unable to isolate additional cdc2 genes from Dictyostelium using polymerase chain reaction technology, but have isolated a gene that is highly related to cdc2. The encoded product is a protein of 33 kDa that shares over 60% identity to the cell-cycle-dependent Cdc2 kinases. However, despite this high level of identity, the gene is not capable of complementing the temperature-sensitive cdc28 mutant of Saccharomyces cerevisiae. Furthermore, the gene product shares some characteristics with the recently described PCTAIRE proteins; it contains a PCTAIRE motif instead of the Cdc2 kinase conserved PSTAIRE sequence, it does not possess the conserved GDSEID sequence that is involved in the activation of the enzyme and it has a Ser in the position equivalent to Thr-161. However, the Dictyostelium protein exhibits a slightly higher level of identity to the Cdc2 kinases than to the PCTAIRE proteins and is smaller than any of the PCTAIRE proteins thus far identified. Since the gene product has characteristics of both Cdc2 kinases and PCTAIRE proteins we have designated the gene product Crp (Cdc2-Related PCTAIRE) kinase. The gene is expressed as two transcripts of 1.5 and 1.8 kb and the expression is developmentally regulated with low levels of mRNA in vegetative cells and significantly higher levels throughout the remainder of the differentiation process. These results suggest the possibility that the gene product is involved in Dictyostelium differentiation rather than growth. This report is the first evidence for a highly-related cdc2 gene in unicellular eukaryote. It also demonstrates for the first time that a unicellular eukaryote expresses a protein containing the PCTAIRE sequence.

Differential recruitment of viral and allo-epitopes into the MHC class I antigen processing pathway of a novel mutant of Ltk- cells. HSV/MHC class I restriction/immune recognition/antigen processing/antigen presentation/influenza virus.

It is generally thought that the Ag processing pathways for endogenously synthesized proteins are the same for allo and viral Ag processing. However, this view does not take into consideration the diversity of specialized maturation and assembly pathways for viruses. In particular, viral assembly that takes place within intracellular membranes may require unique Ag processing steps. In this study we sought to assess this possibility. Hence, we describe the CTL response against a murine Ltk- cell derivative, gro29, which was previously shown to be defective in the propagation of herpes simplex virus type 1 (HSV-1). In HSV-1-infected gro29 cells, viral polypeptides are synthesized in normal amounts and viral assembly takes place. However, transport of the assembled particles is defective in these cells, resulting in the accumulation of noninfectious virus in cytoplasmic vesicles, and a reduction in the release of viral particles by at least 2000-fold. We show that the rate of transport of individual endogenous proteins through the organelles of the secretory pathway is also impaired, but only by roughly 50%, suggesting that the defect in this cell line affects the transport of particles to a greater extent than the transport of individual proteins. It is also shown that allogeneic and influenza A- specific CTL responses are indistinguishable between gro29 and Ltk- cells, as is the response against target cells pretreated with a influenza A derived synthetic peptide. By contrast, HSV-1-infected gro29 cells are approximately eightfold less sensitive than infected Ltk- cells to lysis by HSV-1-specific CTL. This illustrates that in contrast to the allogeneic and influenza specific responses, the recruitment of herpes virus-specific Ag into the Ag-processing pathway is dependent on a cellular function that is also required for viral maturation and egress. We believe that this is the first demonstration of this phenomenon.

Parental and professional perceptions of problems associated with long-term pediatric home tube feeding.

We surveyed 24 caregivers of children fed by gastrostomy tube to identify day-to-day problems encountered in home enteral nutrition. In a second survey, 21 respondents cross-validated the problem list by rating how often they experienced each problem and how difficult each problem was to manage. Problems related to social functioning, recreational activities, and family functioning were rated by caregivers as the most frequent and difficult. We asked 11 pediatric feeding specialists to rate the same problems based on their perceptions of the frequency and severity of problems experienced by caregivers. Moderate agreement was found between professional and caregiver ratings, although professionals generally rated problems as more common and difficult than did the caregivers. Additionally, positive relationships were found between caregiver ratings of common gastrostomy-tube problems and general levels of stress in the home. Data suggest that problems identified by caregivers as most common and difficult are often in the social rather than the medical realm. A broadly based family assessment that focuses on medical and social aspects of home enteral nutrition may maximize nutritional benefits for the patient as well as improve general family functioning and reduce stress in the home.

Toxin resistance and reduced secretion in a mouse L-cell mutant defective in herpes virus propagation.

The mouse L-cell mutant gro29 was selected originally for its inability to propagate herpes simplex virus; it shows severe defects in virus egress and the transport and processing of viral glycoproteins after infection. In this report, we show that uninfected gro29 cells display pleiotropic changes in protein secretion, oligosaccharide processing, and sensitivity to the toxins ricin and modeccin. Specifically, the rate of secretion of a nonglycosylated protein, human growth hormone, was reduced 70% in gro29 cells compared with the parental L cells. A direct measurement of the transport capacity of Golgi membranes in a cell-free assay suggests that gro29 cells contain less functional Golgi than parental cells. Despite this deficiency, N-linked oligosaccharides were processed efficiently in mutant cells, although there were differences in the structure of the mature forms. Lectin intoxication assays revealed that gro29 cells were cross-resistant to killing by the cytotoxic lectins ricin and modeccin, but not to wheat germ agglutinin, Ricinus communis agglutinin RCA120, or leucoagglutinin. Fluorescence labeling using fluorescein-conjugated lectins showed that uninfected gro29 cells expressed relatively few ricin-binding molecules, suggesting a possible mechanism for toxin resistance. These studies provide evidence that the processes of protein secretion, lectin intoxication, and herpes virus maturation and egress may share a common cellular component.

Isolation and characterization of a cdc 2 cDNA from Dictyostelium discoideum.

A cdc2 homologous sequence was amplified from Dictyostelium discoideum by the polymerase chain reaction and used to isolate several cDNA clones. The amino acid sequence encoded by these cDNAs exhibited approx. 60% identity to the Cdc2 proteins of other species. A cDNA containing the entire coding sequence complemented the temperature sensitive cdc28 mutant of Saccharomyces cerevisiae, although growth of the transformants was slow and limited. Southern blot analysis of restriction digests under high stringency conditions provided evidence that Dictyostelium contains a single cdc2 gene, although at lower stringency multiple fragments were detected, suggesting the existence of a cdc2 gene family. Northern blot analysis of RNA from different stages of Dictyostelium development showed that cdc2 mRNA levels increased during aggregation and then decreased to low levels by the pseudoplasmodial stage of development. By contrast, cdc2 mRNA levels remained relatively constant as cells passed from exponential growth to the stationary phase.