Role of auxin and gibberellin in the synthesis of Ascorbic Acid and growth of tissue explants

Coleoptile and root tips ofTriticum aestivum cv. Arnej 624 and those ofAvena sativa cv. Victory (Svalöf) as well as dry excised embryos ofTriticum aestivum cv. Rival (Svalöf) and those ofArachis hypogaea cv. 34 3A. H. were cultivated in media containing various concentrations of sucrose and growth regulators, like ascorbic acid, indole-3-acetic acid and gibberellin. Growth, differentiation and water uptake of the various explants were determined at regular time intervals. Further, the concentration of the endogenous ascorbic acid in mg./g. fresh weight, as well as the amount of this growth regulator utilized as per cent of the total were determined. Although all the three growth regulators promote growth in the explants, their effect is best felt when sucrose of a higher concentration (1.0 per cent) is added to the medium. In fact, the response to 1.0 per cent sucrose is sometimes as good as a combination of a growth regulator with sucrose, especially in the case of root explants. The results clearly indicate that the biosynthesis of ascorbic acid in the explants is catalyzed by the addition of indole-3-acetic acid as well as gibberellin. Simultaneously, the utilization of ascorbic acid is also appreciably increased by the presence of these growth regulators. Addition of 1.0 per cent sucrose to the medium containing the above mentioned growth regulators augments to a considerable extent not only the concentration of ascorbic acid, but also steps up its utilization. Enhancement of ascorbic acid as well as its increased utilization are correlated with rapid imbibition of water, growth and differentiation. The role of ascorbic acid in growth is discussed; and on the basis of the data presented here it is postulated that: (1) auxin and gibberellin function in the growth process by catalyzing the biosynthesis of ascorbic acid; and (2) that ascorbic acid not only participates in activation of various enzyme systems, but also stimulates the production of adenosine triphosphate by acting as an electron donor in photosynthetic phosphorylation as well as oxidative phosphorylation; (3) that the above action of ascorbic acid creates a favourable redox balance for synthesis of nucleic acids, proteins, enzymeproteins, and cell-wall constituents, thus enabling the processes of cell division and enlargement to proceed at a fast rate; and (4) that the relative rates of cell division and cell enlargement as well as “ageing” will determine the pattern of plant development.     

Localization of GABA-like immunoreactivity in the ectostriatum of domestic chicks: GABA immunocytochemistry combined with Golgi impregnation

Brain Cell Biology - The distribution of GABA-like immunoreactivity (GABA-LI) in the ectostriatal core (Ec) of domestic chicks (one to two days old) was investigated using (1) preembedding GABA...     

Novel inhibitors of enkephalin-degrading enzymes. III: 4-Carboxymethylamino-4-oxo-3 (phenylamino) butanoic acids as enkephalinase inhibitors.

4-Carboxymethylamino-4-oxo-3-(4'-aminophenylamino) butanoic acid (25), its ethyl ester (26) and the corresponding unsubstituted-aryl analogues (17) and (16) are fairly potent inhibitors of enkephalinase (neutral endopeptidase; EC 3.4.24.11), Ki = 0.14-0.39 microM, with weak inhibitory potency, Ki = 15-75 microM, towards aminopeptidase MII. In the mouse abdominal constriction test, the esters (26) and (16) showed systemic inhibitory (antinociceptive) activity with ED50 values 62 +/- 3.05 and 81 +/- 1.74 mg/kg respectively. In the mouse tail immersion test, both (26) and (16) exhibited antinociceptive activity when administered intracerebroventricularly and (26) also exhibited a systemic effect which was only partially reversed by naltrexone. The antinociceptive effect seen with (26) reflects its ranking in vitro as an inhibitor of enkephalinase (Ki = 0.14 microM) but it is possible that this effect is not totally opioid-mediated. Compounds (26) and (16) represent the first combined inhibitors of enkephalinase and aminopeptidase MII.     

Lineage, migration, and morphogenesis of longitudinal glia in the Drosophila CNS as revealed by a molecular lineage marker

Previous studies described three different classes of glial cells in the developing CNS of the early Drosophila embryo that prefigure and ensheath the major CNS axon tracts. Among these are 6 longitudinal glial cells on each side of each segment that overlie the longitudinal axon tracts. Here we use transformant lines carrying a P element containing a 130 bp sequence from the fushi tarazu gene in front of the lacZ reporter gene to direct beta-galactosidase expression in the longitudinal glia. Using this molecular lineage marker, we show that 1 of the "neuroblasts" in each hemisegment is actually a glioblast, which divides once symmetrically, in contrast to the typical asymmetric neuroblast divisions, producing 2 glial cells, which migrate medially and divide to generate the 6 longitudinal glial cells. As with neuroblasts, mutations in Notch and other neurogenic genes lead to supernumerary glioblasts. The results indicate that the glioblast is similar to other neuroblasts; however, the positionally specified fate of this blast cell is to generate a specific lineage of glia rather than a specific family of neurons.     

Physical and genetic mapping of the protein A gene in the chromosome of Staphylococcus aureus 8325-4

The gene coding for protein A (spa) has been mapped close to nov on the genetic map of the chromosome of Staphylococcus aureus 8325-4. A rapid mapping procedure has been developed which first allowed the region of the chromosome carrying the spa gene to be identified by blot +hybridization of large DNA fragments which had been separated by pulsed-field gel electrophoresis. Restriction endonuclease SmaI fragment G was shown to carry the spa gene. An insertion mutation in spa was constructed by in vitro insertion of a fragment of DNA expressing resistance to kanamycin and neomycin. A spa::Kan(r)Neo(r) mutation was isolated in S. aureus 8325-4 by allele replacement. This provided a selectable marker which allowed the spa gene to be mapped by transformation analysis.     

Patterns of costimulation of T cell clones by cross-linking CD3, CD4/CD8, and class I MHC molecules

The ability of mAb to class I MHC molecules, CD3, or CD4/CD8 to stimulate human T cell clones alone or in combination was examined. Cross-linking each of these surface Ag with appropriate mAb and goat anti-mouse Ig (GaMIg) resulted in a unique pattern of increase in intracellular free calcium ([Ca2+]i) and different degrees of functional activation. Cross-linking class I MHC molecules provided the most effective stimulus of IL-2 production and proliferation. Cross-linking more than one surface Ag induced a compound calcium signal with characteristics of each individual response. Cross-linking CD3 + HLA-A,B,C caused a rapid and prolonged increase in [Ca2+]i and synergistically increased IL-2 production and proliferation of all clones. Cross-linking CD3 + CD4/CD8 also generated a compound calcium signal and increased IL-2 production and DNA synthesis. Purposeful inclusion of CD3 was not required for costimulation as cross-linking HLA-A,B,C + CD4/CD8 also increased [Ca2+]i, IL-2 production, and proliferation. Cross-linking three surface Ag, CD3 + HLA-A,B,C + CD4/CD8, resulted in the greatest initial and sustained [Ca2+]i, IL-2 production, and DNA synthesis. Although there was a tendency for the various stimuli to increase both [Ca2+]i and functional responsiveness, neither the magnitude nor duration of the increased [Ca2+]i correlated with the amount of IL-2 produced or the ultimate proliferative response. To determine whether costimulation required that the various surface molecules were cross-linked together, experiments were carried out using isotype specific secondary antibodies. Augmentation of [Ca2+]i and costimulation of functional responses were noted when class I MHC molecules were cross-linked and CD3 was bound, but not cross-linked. Similarly, costimulation through CD3 and CD4/CD8 was observed when CD4/CD8 was cross-linked and the CD3 complex was engaged by an anti-CD3 mAb which was not further cross-linked. In contrast, costimulation by class I MHC molecules and CD4/CD8 was only observed when these molecules were cross-linked together. These data demonstrate that cross-linking class I MHC determinants or CD4/CD8 provides a direct signal to T cell clones that can be enhanced when CD3 is independently engaged. The results also indicate that T cell clones can be stimulated without engaging CD3 by the combination of signals delivered via class I MHC molecules and CD4/CD8, but only when these determinants were cross-linked together. These studies have demonstrated that these cell surface molecules differ in their capacity to deliver activation signals to T cell clones and also exhibit unique patterns of positive cooperativity in signaling potential.     

The precise radioimmunoassay of adenosine: minimization of sample collection artifacts and immunocrossreactivity.

Anti-adenosine antibodies were produced in rabbits immunized with N6-carboxymethyladenosine conjugated to methyl albumin. 125I-N6-Aminobenzyladenosine was synthesized and used as a high-specific-activity, high-affinity ligand. A radioimmunoassay (RIA) was developed that can detect 6.25 nM (312.5 fmol) of underivatized adenosine and cross-reacts less than 0.02% with adenine nucleotides and guanosine and not at all with 1 mM inosine. The sensitivity of the RIA can be increased to a detection limit of 0.125 nM (6.25 fmol) by derivitizing samples with benzyl bromide to form N6-benzyladenosine. The assay was adapted to an automated RIA procedure. Assay precision was increased by: (i) inhibiting slight adenosine deaminase activity present in anti-sera; (ii) treating buffers and albumin used in the RIA with charcoal to remove contaminating adenosine; and (iii) correcting for a small but variable component of immunoreactivity not attributable to adenosine. A second antibody prepared with a 2',3'-disuccinyladenosine-albumin conjugate was also found to detect some non-adenosine-mediated immunoreactivity in plasma samples. Immunointerference in human plasma was eliminated in samples treated with ZnSO4/Ba(OH)2 or partially purified over C18 Sep Paks to remove nucleotides and assayed after sample benzylation or succinylation. Human blood was mixed with a novel "stop" solution that was optimized to inhibit adenosine formation from AMP by greater than 99% and to inhibit adenosine uptake into red cells and degradation by greater than 94%. Human plasma/stop solution was assayed by RIA and HPLC with equivalent results.     

Effect of hypertension on elasticity and geometry of aortic tissue from dogs

Inflation-extension experiments were carried out on segments of the descending thoracic aortas from 4 normotensive and 4 hypertensive dogs rendered hypertensive using either unilateral or bilateral renal artery constriction. Intravascular pressures up to 200 mm Hg and axial forces up to 200 g were used. The external diameter of the segment and the distance between two longitudinally spaced gage marks were recorded photographically at each pressure-force level combination. Dimensions in the underformed configuration were measured at the end of the inflation-extension experiment. Data were analyzed for changes in geometry and force-deformation response. Results indicate that: 1. Under sustained hypertension the wall thickness in the underformed configuration increases with a concurrent reduction in the in-situ longitudinal extension ratio. 2. This dual tissue response accomplishes substantial reductions in the circumferential and longitudinal stresses from the levels that would be reached at equivalent pressures in the absence of these geometric changes. 3. At comparable intravascular pressures the extensibility in the circumferential direction is slightly greater for the hypertensive aortas as compared to normals. However, the stress-extension ratio relationship in the circumferential direction is similar in the two groups. 4. The stress-extension ratio relationship in the longitudinal direction indicates that the hypertensive aorta is stiffer than its normotensive counterpart.