TBCD Links Centriologenesis,Spindle Microtubule Dynamics,and Midbody Abscission in Human Cells

Microtubule-organizing centers recruit α- and β-tubulin polypeptides for microtubule nucleation. Tubulin synthesis is complex, requiring five specific cofactors, designated tubulin cofactors (TBCs) A–E, which contribute to various aspects of microtubule dynamics in vivo. Here, we show that tubulin cofactor D (TBCD) is concentrated at the centrosome and midbody, where it participates in centriologenesis, spindle organization, and cell abscission. TBCD exhibits a cell-cycle-specific pattern, localizing on the daughter centriole at G1 and on procentrioles by S, and disappearing from older centrioles at telophase as the protein is recruited to the midbody. Our data show that TBCD overexpression results in microtubule release from the centrosome and G1 arrest, whereas its depletion produces mitotic aberrations and incomplete microtubule retraction at the midbody during cytokinesis. TBCD is recruited to the centriole replication site at the onset of the centrosome duplication cycle. A role in centriologenesis is further supported in differentiating ciliated cells, where TBCD is organized into “centriolar rosettes”. These data suggest that TBCD participates in both canonical and de novo centriolar assembly pathways.     

Frequencies of simultaneous transformation with different T-DNAs and their relevance to the Agrobacterium/plant cell interaction

Summary We investigated whether the efficiency of transformation of plant cells by Agrobacterium tumefaciens during cocultivation is limited by the properties of the plant cells or by the infecting bacteria.Therefore, tobacco protoplasts were infected by cocultivation with two different agrobacteria strains carrying Ti plasmids with distinguishable T-DNAs. These T-DNAs cotransform plant cells at a frequency equal to the product of their independent transformation frequencies, which indicates that all plant cells are equally competent. On the other hand, when these T-DNAs are located on the same Ti plasmid vector within one bacterial strain, the cotransformation frequency is significantly higher than the product of the single transformation frequencies. We interpret these results to indicate that transformation is limited more by the establishment of effective bacteria/plant cell interaction than by (i) the process of DNA integration and (ii) by the number of plant cells capable of being transformed by Agrobacterium. We found that most plant cells are transformed by only one or a few agrobacteria. Analysis of the number of T-DNA copies in these clonally transformed lines indicates amplification of the original, infecting T-region copy.     

Structural characterization of cytochrome c peroxidase by resonance Raman scattering

Resonance Raman scattering studies are reported on freshly prepared and aged ferric, ligand-free ferrous, and CO-bound ferrous cytochrome c peroxidase. The ferric form of the fresh enzyme has a heme which is penta-coordinate high spin, independent of buffer over the pH range 4.3-7, as determined by well established Raman marker lines. The aged enzyme displays a mixture of spin and coordination states, but it can be stabilized in the penta-coordinate high spin form in the presence of phosphate. These results can be accounted for by considering the size of the channel (6 A wide, 11 A long) between the distal side of the heme and the outer surface of the protein. A phosphate ion may be accommodated in this channel resulting in the stabilization of the distal heme pocket. The ferrous cytochrome c peroxidase in both the ligand-free and CO-bound states has an acidic and an alkaline form. The acidic form has the characteristic spectral features of peroxidases: a high frequency iron-histidine stretching mode (248 cm-1), a high frequency Fe-CO stretching mode (537 cm-1), and a low frequency C-O stretching mode (1922 cm-1). At alkaline pH these frequencies become similar to those of hemoglobin and myoglobin, with the corresponding modes located at 227, 510, and 1948 cm-1, respectively. We attribute the acid/alkaline transition in the ferrous forms of cytochrome c peroxidase to a rearrangement mainly of the proximal side of the heme, culminating in a change of steric interactions between the proximal histidine and the heme or of the hydrogen bonding network involving the proximal histidine. The new data presented here reconcile many inconsistencies reported in the past.     

New Zealand machair vegetation

Abstract. Machair vegetation is reported for the first time from New Zealand. The habitat is similar to that of British machairs in climate, topography and generally in soil. pH and CaCO₃ content are much lower through most of the sequence, though this difference may partly reflect the greater disturbance of British machair. Sea machair is present, predominantly comprising native species. This grades into machair proper, which contains many species found also in British machair. The machair includes Ammophila-occupied hillocks, a feature typical of British machair. Machair marsh is also present.     

Lawns have vertical stratification

Abstract. Vertical stratification was examined in a closely-mown lawn, at both the beginning and end of the mowing/regrowth cycle. Two treatments were examined: Undisturbed, which had been maintained as lawn for 30 yr, and Mechanically Perturbed, on which succession was occurring, after complete removal of vegetation 9.5 months previously. There was significant vertical stratification at both stages of the mowing/regrowth cycle, i.e. even immediately after mowing to a height of 3 cm, and in both treatments. There was general consistency of the vertical role of species during the cycle and between treatments. Relative vertical positions were not closely correlated with absolute leaf heights. However, in no case was the vertical position of a pair of species significantly reversed during the cycle or between treatments. No mutually cyclic vertical relations between species were seen. It is concluded that vertical stratification occurs in even the shortest communities, it starts to develop quite early in succession, and it is dependent largely on the intrinsic properties of the species, their height and (at least in lawns) their recovery from defoliation.     

Characterization of a quail suspension cell line for production of a fusogenic oncolytic virus

The development of efficient processes for the production of oncolytic viruses (OV) plays a crucial role regarding the clinical success of virotherapy. Although many different OV platforms are currently under investigation, manufacturing of such viruses still mainly relies on static adherent cell cultures, which bear many challenges, particularly for fusogenic OVs. Availability of GMP-compliant continuous cell lines is limited, further complicating the development of commercially viable products. BHK21, AGE1. CR and HEK293 cells were previously identified as possible cell substrates for the recombinant vesicular stomatitis virus (rVSV)-based fusogenic OV, rVSV-NDV. Now, another promising cell substrate was identified, the CCX.E10 cell line, developed by Nuvonis Technologies. This suspension cell line is considered non-GMO as no foreign genes or viral sequences were used for its development. The CCX.E10 cells were thus thoroughly investigated as a potential candidate for OV production. Cell growth in the chemically defined medium in suspension resulted in concentrations up to 8.9 × 10⁶ cells/mL with a doubling time of 26.6 h in batch mode. Cultivation and production of rVSV-NDV, was demonstrated successfully for various cultivation systems (ambr15, shake flask, stirred tank reactor, and orbitally shaken bioreactor) at vessel scales ranging from 15 mL to 10 L. High infectious virus titers of up to 4.2 × 10⁸ TCID₅₀/mL were reached in orbitally shaken bioreactors and stirred tank reactors in batch mode, respectively. Our results suggest that CCX.E10 cells are a very promising option for industrial production of OVs, particularly for fusogenic VSV-based constructs.     

The evolutionary relationships of two families of cottoid fishes of Lake Baikal (East Siberia) as suggested by analysis of mitochondrial DNA

Fragments of mtDNA genes Cyt B, ATPase 6, and ATPase 8 of six cottoid fishes species of Lake Baikal (East Siberia) were amplified and sequenced. In addition mtDNAs of the same fish were subjected to restriction analysis. The data obtained were used to construct phylogenetic trees. The topology of the ATPase tree differs from those of the Res (restriction) and Cyt B trees. Clustering of species within the trees confirms the viewpoint of Taliev (1955, Baicalian Sculpins (Cottoidei)) according to which Baikalian cottoids originate from two ancestral forms. The times of branching obtained do not confirm the existing viewpoint according to which the two golomyankas (Comephorus baicalensis and Comephorus dybowskii) are pre-Baikal (Myocene) relicts: these two species may have originated 1.2–1.8 million years ago in Baikal, and they seem to represent an example of rapid morphological evolution which resulted in the formation of a new family. Correspondence to: S. Ja. Slobodyanuk     

Responses of barley,pea, and maize to inoculation with different vesicular-arbuscular mycorrhizal fungi in irradiated soil

Summary The efficiency of different VAM fungi was investigated by inoculating barley, pea, and maize with different VAM fungi in irradiated soil in pots buried in the field. VAM frequency, growth and nutrient uptake were measured. In barleyGlomus epigaeus (CA) andG. macrocarpus (CA) were the most efficient out of 11 tested species and increased yield of grain by 24% and 21%, though they were not significant according to oneway analysis of variance. In pea, yield of grain was significantly increased from 46% to 104% (mean=68%) by 7 out of 10 tested species and by 105% by application of P fertilizer. The most efficient species wereG. epigaeus (CA),G. mosseae (GB), andG. etunicatus (CA). In maizeG. mosseae (GB) andG. caledonius (DK) increased total yield significantly by 59% and 47% in one experiment and in another experiment yield of cob was increased by 68% byG. mosseae (GB), 72% byG. caledonius (DK), and by 153% by application of P fertilizer. This experiment demonstrated that responsiveness to inoculation by VAM fungi differed among plant species, and that efficiency of different VAM fungi differed.     

Inhibitory activity of palladium(II) and platinum(II) complexes with isoxazole and its derivatives

The investigation of the inhibitory activity on the membrane bonded ATP-ase of the M(L)₂X₂ complexes [where M = Pd(II), Pt(II); L = isoxazole(isox), 3,5-dimethylisoxazole(3,5-diMeisox), 3-methyl,5-phenylisoxazole(3-Me,5-Phisox), 3,5-diphenylisoxazole-(3,5-diPhisox), and 4-amino-3,5-dimethylisoxazole(4-ADI); X = Cl, Br] is reported. These results show that the complexes with isox and its methyl and phenyl derivatives have a much stronger inhibitory effect than the corresponding free ligands; in the 4-ADI compounds this activity drops. The Pd(II) complexes have a greater effect than the Pt(II) derivatives. The interaction occurs with SH groups and probably also with other active centers of the enzyme. These conclusions have been correlated with the E.S.C.A. spectra. These measurements show that the electron density of the complexes on the central metal ion and N_ring atom or N_ring and N-hexocyclic atoms on passing from chloride to bromide derivatives changes slightly.     

Amphipod crustaceans as an important component of zoobenthos of the shallow Antarctic sublittoral

Benthic quantitative samples were taken in 1988 in the soft bottom sublittoral of Admiralty Bay (King George Island, South Shetlands) using a Tvärminne-type bottom sampler and SCUBA-diving technique at 7 successive stations situated at depths from 4 to 30 m.Dominant animal groups in terms of abundance were Amphipoda, Polychaeta and Bivalvia, whereas in terms of biomass Echinoidea were also dominant. Amphipod crustaceans clearly dominated the zoobenthos at depths from 10 to 25 m (the numerical share surpaising 60%) with maximal abundance of abt. 17 000 ind m^–2; in terms of biomass at specific depths amphipods occupied the 1st, 2nd or 3rd place with maximal biomass of abt. 100 g m^–2 where the maximal total biomass of zoobenthos reached 260 g m^–2 (10 m).Amphipoda were the most diversified group with some 35 taxa belonging to 14 families. Most species belonged to Eusiridae s.l. and Lysianassidae s.l. Dominant forms were Pontogeneiella brevicornis, Prostebbingia gracilis, Schraderia gracilis, Hippomedon kergueleni, Orchomenella cf. ultima, Cardenio paurodactylus and Paraphoxus rotundifrons.