The pattern of proliferation of the neuroblasts in the wild-type embryo of Drosophila melanogaster

Summary The pattern of neuroblast divisions was studied in thoracic and abdominal neuromeres of wild-type Drosophila melanogaster embryos stained with a monoclonal antibody directed against a chromatin-associated antigen. Since fixed material was used, our conclusions are based upon the statistical evaluation of a large number of accurately staged embryos, covering the stages between the formation of the cephalic furrow up to shortened germ band. Our observations point to a rather stereotypic pattern of proliferation, consisting of several parasynchronous cycles of division. The data suggest that all SI neuroblasts divide at least eight times, all SII neuroblasts six or seven times and all SIII neuroblasts at least five times. This conclusion is based on the mapping of mitotic neuroblasts and is supported by the progressive reduction of the neuroblast volume and by the results of cell countings performed on embryos of increasing age. No conclusive evidence was obtained concerning the fate of the neuroblasts after their last mitosis, i.e. it cannot be decided whether the neuroblasts degenerate or become incorporated as inconspicuous cells in the larval ventral cord. The duration of the cycles of division of the neuroblasts was found to be 40–50 min each, while in the case of ganglion mother cells about 100 min are required to complete one cell cycle.     

Internal motions in yeast phenylalanine transfer RNA from 13C NMR relaxation rates of modified base methyl groups: a model-free approach

Internal motions at specific locations through yeast phenylalanine tRNA were measured by using nucleic acid biosynthetically enriched in 13C at modified base methyl groups. Carbon NMR spectra of isotopically enriched tRNA(Phe) reveal 12 individual peaks for 13 of the 14 methyl groups known to be present. The two methyls of N2,N2-dimethylguanosine (m22G-26) have indistinguishable resonances, whereas the fourteenth methyl bound to ring carbon-11 of the hypermodified nucleoside 3' adjacent to the anticodon, wyosine (Y-37), does not come from the [methyl-13C]methionine substrate. Assignments to individual nucleosides within the tRNA were made on the basis of chemical shifts of the mononucleosides [Agris, P. F., Kovacs, S. A. H., Smith, C., Kopper, R. A., & Schmidt, P. G. (1983) Biochemistry 22, 1402-1408; Smith, C., Schmidt, P. G., Petsch, J., & Agris, P. F. (1985) Biochemistry 24, 1434-1440] and correlation of 13C resonances with proton NMR chemical shifts via two-dimensional heteronuclear proton-carbon correlation spectroscopy [Agris, P. F., Sierzputowska-Gracz, H., & Smith, C. (1986) Biochemistry 25, 5126-5131]. Values of 13C longitudinal relaxation (T1) and the nuclear Overhauser enhancements (NOE) were determined at 22.5, 75.5, and 118 MHz for tRNA(Phe) in a physiological buffer solution with 10 mM MgCl2, at 22 degrees C. These data were used to extract two physical parameters that define the system with regard to fast internal motion: the generalized order parameters (S2) and effective correlation times (tau e) for internal motion of the C-H internuclear vectors.(ABSTRACT TRUNCATED AT 250 WORDS)     

The genotypic response of mouse embryos to multiple freezing variables

Four- and eight-cell embryos from 3 mouse genotypes were cryopreserved to study the relationship of genetics and freezing variables on post-thaw survival. Embryos from outbred N:NIH(S), N:NIH(S)-B and inbred (C57BL/6N) mice were exposed to 1 of 2 cryoprotectants (glycerol [GLYC] versus dimethyl sulfoxide [DMSO]) and stored in 1 of 2 containers (ampules [AMP] versus straws [STR]). Containerized embryos were cooled at a rate of 0.5 degrees C/min to a minimum of -35 degrees C, transferred into liquid nitrogen, and later thawed and cultured in vitro. Genotypic differences (p less than 0.05) were noted for 4 interrelated embryo characteristics including post-thaw survival (PTS), embryo degeneration rate (EDR), and quality grade (QG) and viability grade (VG) ratings. The PTS for outbred embryos was greater (p less than 0.05) in GLYC than DMSO, whereas inbred C57BL/6N embryos survived similarly (p greater than 0.05) in either cryoprotectant. Compared to DMSO counterparts, embryos from GLYC-treated outbred groups had improved QG and VG ratings and reduced EDR (p less than 0.05), but comparable results were observed between GLYC- AND DMSO-treated embryos in the C57BL/6N group. Between genotypes, type of container affected PTS and EDR (p less than 0.05) but not QG or VG ratings (p greater than 0.05). Within genotypes, PTS generally ranged 15 to 20% higher (p less than 0.01) in AMP than STR groups. Increased PTS was noted in outbred mouse x GLYC x AMP groups; however, based on the degrees of difference, the inbred C57BL/6N strain appeared less affected by this 3-way interaction.(ABSTRACT TRUNCATED AT 250 WORDS)     

Continuous production of L-phenylalanine by transamination

L-Phenylalanine was produced continuously from L-as-partate and phenylpyruvate by transaminase from a newly screened Pseudomonas putida strain. The process was carried out with an isolated enzyme in homogeneous phase in an enzyme membrane reactor and with immobilized whole cells in a stirred tank reactor, respectively. Due to the difference in transport resistance, the productivity of the free enzyme in homogeneous phase (72 mmol/L h) was about 3 times higher than the productivity achieved using immobilized cells. However, a better stability of the biocatalyst was observed with immobilized cells.     

Growth of Nitrobacter by dissimilatoric nitrate reduction

Abstract Eight strains of the genus Nitrobacter grew under anaerobic conditions in the presence of nitrate. The growth was inhibited by nitrate concentrations above 0.5 mM. By a special culture technique inhibition caused by nitrite was abolished. Nitrate oxidizing cells grew in gas tight culture flasks as a biofilm on a gas-permeable silicone tubing. The biofilm allowed nitrate-reducing cells to grow at a low nitrite concentration. These cells grew either actively motile in the anaerobic medium, or in anaerobic zones of the biofilm. They produced nitrite and ammonia. Nitrogen balance calculations established a loss of inorganic nitrogen for 5 of 8 strains. This implies that nitrate-reducing cells produced furthermore volatile nitrogen compounds. N₂O was detected by gas chromatography.     

The maize autonomous element Activator (Ac) shows a minimal germinal excision frequency of 0.2%–0.5% in transgenic Arabidopsis thaliana plants

Summary The autonomous mobile element Activator from Zea mays was introduced into Arabidopsis thaliana via Agrobacterium-mediated gene transfer. The use of a chimaeric construct, where the Ac element is located in the leader of the neomycin phosphotransferase (NPT II) gene, enabled the excision of Ac to be monitored by assaying for the reconstitution of NPT II gene activity. Using this approach, the transpositional activity of AC was initially studied in primary transformants. About 50% of the regenerating Ac transformants showed evidence for excision of the element. Reintegration of Ac was confirmed by Southern blot analysis. Transposition events are transmitted to the F1 generation with a minimal frequency of 0.3%. In a few exceptional cases they are detected in a high proportion of the F1 generation. Seedlings from the F2 and F3 generations were assayed for the rate of germinal excisions by scoring for kanamycin resistance. The minimal frequency of germinal excision events amounts to 0.2%–0.5% and hence allows the use of the Ac element for gene tagging purposes in A. thaliana.     

Studies of the oxysterol inhibition of tumor cell growth

The oxysterols 3 beta-hydroxy-5 alpha-cholest-8-en-11-one, 3 beta-hydroxy-5 alpha-cholest-8-en-7-one, 3 beta-hydroxy-5 alpha-cholest-8(14)-en-7-one, 3 beta-hydroxy-4,4'-dimethylcholest-5-ene-7 one, 4,4'-dimethylcholest-5-ene-3 beta, 7 alpha-diol, 4,4'-dimethylcholest-5-ene-3 beta, 7 beta-diol, lanost-8-ene-3 beta, 25-diol, 25-hydroxylanost-8-en-3-one, 9 alpha, 11 alpha-epoxy-5 alpha-cholest-7-en-3 beta-ol, 3 beta-hydroxycholest-5 alpha-en-22-one, and 3 beta-hydroxycholest-5-en-22-one oxime were evaluated with respect to their ability to inhibit cell growth. All of the sterols were found to possess cytotoxicity when incubated with hepatoma (HTC) and lymphoma (RDM-4) cells in culture at 10-30 microM concentrations.     

A mutant rat major histocompatibility haplotype showing a large deletion of class I sequences

The LEW.1LM1 inbred rat strain, which has been derived from a (LEW×LEW.1W) F₂ hybrid, carries a major histocompatibility (RT1) haplotype which is distinct from that of the LEW strain (RT1 ¹) in that certainRT1.C region-determined class I antigens are not expressed. Here we show that this phenotypic defect is due to genomic deletion of about 100 kb of theRT1.C region. Certain deleted DNA fragments have been cloned from the wild-type DNA into the EMBL4 vector. Five clones have been characterized and are shown to possess different restriction maps and to each carry a single stretch of class I cross-hybridizing sequences. Probes derived from the non-class I coding part of two clones detect fragments which are present in the wild-type but absent from thelm1 mutant. The type of deletion described here in the rat is discussed in the context ofH-2D/Q deletions in the mouse.     

Bioluminescence of the insect pathogen Xenorhabdus luminescens

Luminescence of batch cultures of Xenorhabdus luminescens was maximal when cultures approached stationary phase; the onset of in vivo luminescence coincided with a burst of synthesis of bacterial luciferase, the enzyme responsible for luminescence. Expression of luciferase was aldehyde limited at all stages of growth, although more so during the preinduction phase. Luciferase was purified from cultures of X. luminescens Hm to a specific activity of 4.6 x 10(13) guanta/s per mg of protein and found to be similar to other bacterial luciferases. The Xenorhabdus luciferase consisted of two subunits with approximate molecular masses of 39 and 42 kilodaltons. A third protein with a molecular mass of 24 kilodaltons copurified with luciferase, and in its presence, either NADH or NADPH was effective in stimulating luminescence, indicating that this protein is an NAD(P)H oxidoreductase. Luciferases from two other luminous bacteria, Vibrio harveyii (B392) and Vibrio cholerae (L85), were partially purified, and their subunits were separated in 5 M urea and tested for complementation with the subunits prepared from X. luminescens Hb. Positive complementation was seen with luciferase subunits among all three species. The slow decay kinetics of the Xenorhabdus luciferase were attributed to the alpha subunit.