Influence of plasma pedestal profiles on access to ELM-free regimes in ITER

The influence of current density and pressure gradient profiles in the pedestal on the access to the regimes free from edge localized modes (ELMs) like quiescent H-mode in ITER is investigated. Using the simulator of MHD modes localized near plasma boundary based on the KINX code, calculations of the ELM stability were performed for the ITER plasma in scenarios 2 and 4 under variations of density and temperature profiles with the self-consistent bootstrap current in the pedestal. Low pressure gradient values at the separatrix, the same position of the density and temperature pedestals and high poloidal beta values facilitate reaching high current density in the pedestal and a potential transition into the regime with saturated large scale kink modes. New version of the localized MHD mode simulator allows one to compute the growth rates of ideal peeling-ballooning modes with different toroidal mode numbers and to determine the stability region taking into account diamagnetic stabilization. The edge stability diagrams computations and sensitivity studies of the stability limits to the value of diamagnetic frequency show that diamagnetic stabilization of the modes with high toroidal mode numbers can help to access the quiescent H-mode even with high plasma density but only with low pressure gradient values at the separatrix. The limiting pressure at the top of the pedestal increases for higher plasma density. With flat density profile the access to the quiescent H-mode is closed even with diamagnetic stabilization taken into account, while toroidal mode numbers of the most unstable peeling-ballooning mode decrease from n = 10?40 to n = 3?20.     

Impacts of pellets injected from the low-field side on plasma in ITER

Impacts of pellets injected from the low-field side (LFS) on plasma in ITER are investigated using the 1.5D BALDUR integrated predictive modeling code. In these simulations, the pellet ablation is described using the neutral gas shielding (NGS) model. The pellet ablation model is coupled with the plasma core transport model, which is a combination of the MMM95 anomalous transport model and NCLASS neoclassical transport model. The boundary conditions are assumed to be at the top of the pedestal, in which the pedestal parameters are predicted using a pedestal model based on the theoretical-based pedestal width scaling (either magnetic and flow shear stabilization width scaling, or flow shear stabilization width scaling, or normalized poloidal pressure width scaling) and the infinite-n ballooning mode pressure gradient limit. These pedestal models depend sensitively on the density at the top of the pedestal, which can be strongly influenced by the injection of pellets. The combination of the MMM95 and NCLASS models, together with the pedestal and NGS models, is used to simulate the time evolution of the plasma current, ion and electron temperatures, and density profiles for ITER standard type-I ELMy H-mode discharges during the injection of LFS pellets. It is found that the injection of pellets results in a complicated plasma scenario, especially in the outer region of the plasma and the plasma conditions at the boundary in which the pellet has an impact on increasing the plasma edge density, but reducing the plasma edge temperature. The LFS pellet has a stronger impact on the edge as compared to the center. For fusion performance, the pellet can result in either enhancement or degradation, depending sensitively on the pellet parameters; such as the pellet size, pellet velocity, and pellet frequency. For example, when a series of deuterium pellets with a size of 0.5 cm, velocity of 1 km/s, and frequency of 2 Hz are injected into the ITER plasma from the LFS, the plasma performance, evaluated in terms of Q _fusion, can increase to 72% of that before the use of pellets. It is also found that the injection of pellets results in an increase in the ion and electron densities, but does not enhance the central plasma density. On the other hand, it results in the formation of another peak of the plasma density in the outer region near the plasma edge. The formation of the density peak results in the reduction of plasma transports near the edge by decreasing the contributions of ion-temperature-gradient and trapped electron modes, as well as kinetic ballooning modes.     

Nature of collagens synthesized by monkey periodontal-ligament fibroblasts in vitro.

1. First subcultures of fibroblast-like cells from adult monkey periodontal ligament were incubated in the presence of 14C-labelled amino acids and produced significant amounts of type-I and type-III collagens. 2. The proportion of type-III collagen produced was calculated on the basis of the recovery of procollagens from DEAE-cellulose chromatography to be approx. 20%, and at least 10% when analysed as collagens on CM-cellulose chromatography. 3. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of the procollagens, the collagens and their CNBr peptides was used to confirm the identity of the collagen types. 4. In serum-free media extensive conversion of type-I procollagen, but not of type-III procollagen, into collagen was observed, suggesting that a specific type-I procollagen peptidase was produced. 5. The pattern of collagen synthesis was not significantly different from that obtained with fibroblasts derived from skin corium of the same animals.     

Study of Turbulence in the Globus-M Tokamak Plasma during the Transition to the ELM-free H-mode

Plasma Physics Reports - Here we report the results of the turbulence study in the high-confinement mode (H-mode) with and without edge localized modes (ELMs). The study was performed by the...     

Multinucleate nature,and mating by use of isozyme analysis inPoria cocos

Double staining study of nuclei and cell walls inPoria cocos indicated that the hyphal cells were multinucleate and had no clamp connections. Isozyme analysis of alcohol dehydrogenase (ADH) in 52 natural isolates revealed that there were three types of banding patterns: type I, five bands; type II, one slow band; type III, one fast band. Regenerants expressing type-II or type-III ADH-isozyme pattern were obtained from type-I isolates via protoplast manipulation. When the type-II regenerants were mated with the type-III regenerants, hyphae of type-I phenotype appeared. These data indicated that these type-II and type-III regenerants derived from protoplasts of the type-I isolates were primary hyphae. These primary hyphal cells were also multinucleate. Inter-strain mating ofP. cocos was performed and confirmed by ADH-isozyme analysis. Confronting cultures of a type-III regenerant derived from protoplasts of a type-I isolate and a type-II regenerant derived from a type-II isolate resulted in type-I hyphae.     

Effects of colchicine on the ultrastructure of mouse taste buds

Summary Effect of colchicine on the ultrastructure of taste bud cells was studied in the mouse. In untreated mice microtubules were abundant throughout the entire cytoplasm of type-III cells, but only in the apical cytoplasm of type-I cells. After 2 h of colchicine treatment, no microtubules were observed in any taste bud cells; dense secretory granules in the apical cytoplasm of type-I cells mostly disappeared, and instead, numerous phagosomes appeared. It is suggested that colchicine causes an interruption of the transport of the secretory granules in type-I cells from the Golgi apparatus to the membrane of the apical surface, from which release occurs. In type-III cells, after 4 or 5 h of treatment, dense-cored vesicles scattered throughout the cytoplasm tended to increase in number; they were often observed to accumulate in the vicinity of the Golgi apparatus. Five hours after treatment with 5-hydroxy-l-tryptophan (5-HTP) following colchicine pretreatment, monoamine specific fluorescent cells and vesicles with highly electron-dense cores of type-III cells were still present. On the other hand, 5 h after 5-HTP treatment alone both fluorescent cells and vesicles with highly electron-dense cores had already disappeared. These observations suggest that the treatment with colchicine interrupts the transport of densecored vesicles of type-III cells to synaptic areas, in which those vesicles are presumed to discharge the neurotransmitter substance.     

Identification of "buried" lysine residues in two variants of chloramphenicol acetyltransferase specified by R-factors.

Two variants of chloramphenicol acetyltransferase which are specified by genes on plasmids found in Gram-negative bacteria were subjected to amidination with methyl acetimidate to determine the relative reactivity of surface lysine residues and to search for unreactive or "buried" amino groups which might contribute to stabilization of the native tetramers. Representative examples of the type-I and type-III variants of chloramphenicol acetyltransferase were found to have one lysine residue each in the native state which appears to be inaccessible to methyl acetimidate. The uniquely unreactive residue of the type-I protein is lysine-136, whereas the lysine that is "buried" in the type-III enzyme is provisonally assigned to residue 38 of the prototype sequence. It is suggested that the lysine residue in each case participates in the formation of an ion pair at the intersubunit interface and that the two amino groups in question occupy functionally equivalent positions in the quaternary structures of their respective enzyme variants. Lysine-136 of type-I enzyme is also uniquely unavailable for modification by citraconic anhydride, a reagent used to disrupt the quaternary structure of the native enzyme. Contrary to expectation, exhaustive citraconylation fails to dissociate the tetramer, but does destroy catalytic activity. Removal of citraconyl groups from modified chloramphenicol acetyltransferase is accompanied by a full region of catalytic activity. Analysis of the rate of hydrolysis of citraconyl groups from the modified tetramer by amidination of unblocked amino groups with methyl [14C]acetamidate reveals difference in lability for several of the ten modified lysine residues. Although the unique stability of the quaternary structure of chloramphenicol acetyltransferase may be due to strong hydrophobic interactions, it is argued that lysine-136 may contribute to stability via the formation of an ion pair at the subunit interface.     

Time-lapse tracking of barley androgenesis reveals position-determined cell death within pro-embryos

Following abiotic stress to induce barley (Hordeum vulgare L.) androgenesis, the development of 794 enlarged microspores in culture was monitored by time-lapse tracking. In total, 11% of the microspores tracked developed into embryo-like structures (type-I pathway), 36% formed multicellular structures (type-II pathway) and 53% of the microspores followed gametophytic divisions, accumulated starch and died in the first days of tracking (type-III pathway). Despite the microspore fate, enlarged microspores showed similar morphologies directly after stress treatment. Ultrastructural analysis, however, revealed two morphologically distinct cell types. Cells with a thin intine layer and an undifferentiated cytoplasm after stress treatment were associated with type-I and type-II pathways, whereas the presence of differentiated amyloplasts and a thick intine layer were associated with the type-III pathway. Tracking revealed that the first morphological change associated with embryogenic potential was a star-like morphology, which was a transitory stage between uninucleate vacuolated microspores after stress and the initiation of cell division. The difference between type-I and type-II pathways was observed during the time they displayed the star-like morphology. During the transition phase, embryo-like structures in the type-I pathway were always released out of the exine wall at the opposite side of the pollen germ pore, whereas in the type-II pathway multicellular structures were unable to break the exine and to release embryo-like structures. Moreover, by combining viability studies with cell tracking, we show that release of embryo-like structures was preceded by a decrease in viability of the cells positioned at the site of exine wall rupture. These cells were also positively stained by Sytox orange, a cell death indicator. Thereby, we demonstrate, for the first time, that a position-determined cell death process marks the transition from a multicellular structure into an embryo-like structure during barley androgenesis.     

Difference in thermal stability of type-I and type-III collagen from rat skin

Type-I and type-III collagens were obtained by differential salt fractionation of neutral-salt-soluble collagen from rat skin. Their thermal stabilities were determined by u.v. difference spectroscopy. The `melting' temperature (T_m) in 5mm-acetic acid of type-III collagen was almost 2°C above that of type-I collagen. Intramolecular covalent cross-linking had no effect on the thermal stability.     

Increased type-III/type-I collagen ratios in diabetic human conjunctival biopsies

The ratio of type-III to type-I collagen is measured in human conjunctival biopsies from control and diabetic subjects. The tissue is digested by CNBr and the resulting peptides are quantified by SDS polyacrylamide gel electrophoresis. The peptides used are alpha 1-(I)CB7 and alpha 1-(III)CB8. In control population, of type-III collagen slightly increases with age. In two diabetic populations, (juvenile onset diabetes and maturity onset diabetes), the percentage of type-III collagen is significantly higher than in age-matched control groups. These data plus those previously obtained on genetically diabetic mice indicate that diabetes mellitus affects the expression of interstitial collagen phenotype. Preliminary results on prediabetic subjects suggest the role of genetic factors in such alterations.     

The interaction of calmodulin with alternatively spliced isoforms of the type-I inositol trisphosphate receptor

A 592-amino acid segment of the regulatory domain of the neuronal type-I inositol 1,4,5-trisphosphate receptor (IP(3)R) isoform (type-I long, amino acids1314-1905) and the corresponding 552-amino acid alternatively spliced form present in peripheral tissues (type-I short, amino acids 1693-1733 deleted) were expressed as glutathione S-transferase fusion proteins. These domains encompass a putative calmodulin (CaM) binding domain and two protein kinase A phosphorylation sites. Both long and short fusion proteins retained the ability to bind CaM in a Ca(2+)-dependent manner as measured by CaM-Sepharose chromatography or a dansyl-CaM fluorescence assay. Both assays indicated that the short fusion protein bound twice the amount of CaM than the long form at saturating concentrations of CaM. In addition, the binding of the short form to CaM-Sepharose was inhibited by phosphorylation with protein kinase A, whereas the binding of the long form was unaffected. Full-length cDNAs encoding type-I long, type-I short, and type-III IP(3)R isoforms were expressed in COS cells, and the Ca(2+) sensitivity of [(3)H]IP(3) binding to permeabilized cells was measured. The type-I long isoform was more sensitive to Ca(2+) inhibition (IC(50) = 0.55 microM) than the type-I short (IC(50) = 5.7 microM) or the type-III isoform (IC(50) = 3 microM). In agreement with studies on the fusion proteins, the full-length type-I short bound more CaM-Sepharose, and this binding was inhibited to a greater extent by protein kinase A phosphorylation than the type-I long IP(3)R. Although type-III IP(3)Rs did not bind directly to CaM-Sepharose, hetero-oligomers of type-I/III IP(3)Rs retained the ability to interact with CaM. We conclude that the deletion of the SII splice site in the type-I IP(3)R results in the differential regulation of the alternatively spliced isoforms by Ca(2+), CaM, and protein kinase A.     

Characterization of pepsin-solubilized bovine heart-valve collagen.

Collagens extracted from heart valves by using limited pepsin digestion were fractionated by differential salt precipitation. Collagen types were identified by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, amino acid analysis and cleavage with CNBr. Heart-valve collagen was heterogeneous in nature, consisting of a mixture of type-I and type-III collagens. The identity of type-III collagen was established on the basis of (a) insolubility in 1.7 M-NaC1 at neutral pH, (b) behaviour of this collagen fraction on gel electrophoresis under reducing and non-reducing conditions, (c) amino acid analysis showing a hydroxyproline/proline ratio greater than 1, and (d) profile of CNBr peptides on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis showing a peak characteristic for type-III collagen containing peptides alpha1(III)CB8 and alpha1(III)CB3. In addition to types-I and -III collagen, a collagen polypeptide not previously described in heart valves was identified. This polypeptide represented approx. 30% of the collagen fraction precipitated at 4.0 M-NaCl, it migrated between beta- and alpha1-collagen chains on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and its electrophoretic behaviour was not affected by disulphide-bond reduction. All collagen fractions from the heart valves contained increased amounts of hydroxylysine when compared with type-I and -III collagens from other tissues. The presence of beta- and gamma-chains and higher aggregates in pepsin-solubilized collagen indicated that these collagens were highly cross-linked and suggested that some of these cross-links involved the triple-helical regions of the molecule. It is likely that the higher hydroxylysine content of heart-valve collagen is responsible for the high degree of intermolecular cross-linking and may be the result of an adaptive mechanism for the specialized function of these tissues.     

Isolation and identification of the major plasma fibronectin cDNA from Japanese catfish Silurus asotus

Major plasma fibronectin from Japanese catfish was isolated using affinity chromatography, and the fibronectin was digested with thermolysin. Peptide sequences of the fragments were obtained by peptide sequencer. Complete fibronectin cDNA was obtained from Japanese catfish liver cells using 5'-rapid amplification of cDNA end (RACE) and 3'-RACE based on the peptide sequences. It consists of a 6885 bp open reading frame, which is putatively translated to a protein of 2295 amino acids resides. The catfish fibronectin has 12 type-I modules, 2 type-II modules and 15 type-III modules, and variable sites V and lacks both EIIIA and EIIIB sites. Homology of the entire amino acids residues of catfish fibronectin with those of mammals (Homo sapiens, Rattus norvegicus, Bos taurus) is only 47-48% and 57% with that of Danio rerio. However, amino acid sequence of type-I module 3 and type-I module12 are highly conserved and homology exceeds 80% with corresponding regions of the mammals, Xenopus laevis and fish species (Silurus asotus and D. rerio). Phylogenetic analysis indicates that only type-I module 4 shows a different pattern of phylogenetic tree. One major fibronectin mRNA was detected in whole liver and hepatocytes by northern hybridization, however, five to six other bands were also detected in both samples.     

Inhibition of collagen-induced platelet aggregation by antibodies to distinct types of collagens.

Aggregation of platelets by fibrils formed from collagens type I, II and III could be inhibited by coating the fibrils with anti-collagen antibodies or Fab fragments. Similar results were obtained in a clot-retraction assay. Inhibition was achieved with stoichiometric amounts of antibodies and was specific for each type of collagen. Aggregation caused by a mixture of type-I and -III collagens could only be inhibited by a mixture of antibodies against both collagens. The data show that each interstitial collagen is capable of interacting with platelets and do not support the concept of an outstanding activity of type-III collagen.     

Iodothyronine 5-deiodinase in rat posterior pituitary.

We first describe the presence of iodothyronine 5-deiodinase (5D) in the neural lobe of rat pituitary. 6-n-Propyl-2-thiouracil (PTU), a specific inhibitor of type-I deiodinase, had no effect, showing that 5D in neurohypophysis is of type-III isozyme, which is specific for 5-deiodination and has been found only in the brain, placenta and skin. The presence of 5D (type-III) together with our previous report of 5'-deiodinase (type-I in euthyroidism and type-II in hypothyroidism) shows that the isozymes of deiodinases in the neurohypophysis are quite similar to those in the brain. These data suggest a previously unrecognized role of thyroid hormone in posterior pituitary physiology.     

Identification of sequence variations by PCR-RFLP and its application to the evaluation of cpDNA diversity in wild and cultivated soybeans

The diversity and maternal lineage in wild and cultivated soybeans have previously been assayed using restriction fragment length polymorphism (RFLP) and sequencing analyses of chloroplast DNA (cpDNA). Here we describe a method based on PCR-RFLP for the identification of nucleotides at four mutation sites in non-coding regions of cpDNA. Of the four sites, two were located in restriction enzyme sites and two were not. For the latter two sites, new primers were designed to artificially create restriction sites that spanned them. The PCR-RFLP method enabled us to identify nucleotides at each of the four mutation sites easily and reliably. Fifty-seven wild and sixty-seven cultivated soybeans of different origins and different cpDNA types (types I, II, and III) were assayed. All of the samples tested could be classified into four haplotypes. All of the type-I and -II accessions had the same nucleotides at each of the four mutation sites, while all of the type-III accessions, except for 3 wild ones, had nucleotides that were different from those of types I and II. A sequencing analysis revealed that the 3 wild accessions possessed other single-base variations in the non-coding regions of trnH-psbA and trnT-trnL. The results of this study suggest that the type-I and type-II chloroplast genomes form a group that is distinct from the type-III chloroplast genome. Received: 14 April 2000 / Accepted: 11 July 2000     

Diversity of trypsins in the Mediterranean corn borer Sesamia nonagrioides (Lepidoptera: Noctuidae), revealed by nucleic acid sequences and enzyme purification

The existence of a diverse trypsin gene family with a main role in the proteolytic digestion process has been proved in vertebrate and invertebrate organisms. In lepidopteran insects, a diversity of trypsin-like genes expressed in midgut has also been identified. Genomic DNA and cDNA trypsin-like sequences expressed in the Mediterranean corn Borer (MCB), Sesamia nonagrioides, midgut are reported in this paper. A phylogenetic analysis revealed that at least three types of trypsin-like enzymes putatively involved in digestion are conserved in MCB and other lepidopteran species. As expected, a diversity of sequences has been found, including four type-I (two subtypes), four type-II (two subtypes) and one type-III. In parallel, four different trypsins have been purified from midgut lumen of late instar MCB larvae. N-terminal sequencing and mass spectrometric analyses of purified trypsins have been performed in order to identify cDNAs coding for major trypsins among the diversity of trypsin-like sequences obtained. Thus, it is revealed that the four purified trypsins in MCB belong to the three well-defined phylogenetic groups of trypsin-like sequences detected in Lepidoptera. Major active trypsins present in late instar MCB lumen guts are trypsin-I (type-I), trypsin-IIA and trypsin-IIB (type-II), and trypsin-III (type-III). Trypsin-I, trypsin-IIA and trypsin-III showed preference for Arg over Lys, but responded differently to proteinaceous or synthetic inhibitors. As full-length cDNA clones coding for the purified trypsins were available, three-dimensional protein models were built in order to study the implication of specific residues on their response to inhibitors. Thus, it is predicted that Arg73, conserved in type-I lepidopteran trypsins, may favour reversible inhibition by the E-64. Indeed, the substitution of Val213Cys, unique for type-II lepidopteran trypsins, may be responsible for their specific inhibition by HgCl2. The implication of these results on the optimisation of the use of protease inhibitors for pest control, and on the identification of endoprotease-mediated resistance to Bacillus thuringiensis Cry-toxins is discussed.     

ATPase-Independent Type-III Protein Secretion in Salmonella enterica

Type-III protein secretion systems are utilized by gram-negative pathogens to secrete building blocks of the bacterial flagellum, virulence effectors from the cytoplasm into host cells, and structural subunits of the needle complex. The flagellar type-III secretion apparatus utilizes both the energy of the proton motive force and ATP hydrolysis to energize substrate unfolding and translocation. We report formation of functional flagella in the absence of type-III ATPase activity by mutations that increased the proton motive force and flagellar substrate levels. We additionally show that increased proton motive force bypassed the requirement of the Salmonella pathogenicity island 1 virulence-associated type-III ATPase for secretion. Our data support a role for type-III ATPases in enhancing secretion efficiency under limited secretion substrate concentrations and reveal the dispensability of ATPase activity in the type-III protein export process.