Regulation of asparaginase, glutamine synthetase, and glutamate dehydrogenase in response to medium nitrogen concentrations in a euryhaline chlamydomonas species

The ammonium assimilatory enzymes glutamine synthetase (EC 6.3.1.2) and glutamate dehydrogenase (EC 1.4.1.3) were investigated for a possible role in the regulation of asparaginase (EC 3.5.1.1) in a Chlamydomonas species isolated from a marine environment. Cells grown under nitrogen limitation (0.1 millimolar NH(4) (+), NO(3) (-), or l-asparagine) possessed 6 times the asparaginase activity and approximately one-half the protein of cells grown at high nitrogen levels (1.5 to 2.5 millimolar). Biosynthetic glutamine synthetase activity was 1.5 to 1.8 times greater in nitrogen-limited cells than cells grown at high levels of the three nitrogen sources.Conversely, glutamate dehydrogenase (both NADH- and NADPH-dependent activities) was greatest in cells grown at high levels of asparagine or ammonium, while nitrate-grown cells possessed little activity at all concentrations employed. For all three nitrogen sources, glutamate dehydrogenase activity was correlated to the residual ammonium concentration of the media after growth (r = 0.88 and 0.94 for NADH- and NADPH-dependent activities, respectively).These results suggest that glutamate dehydrogenase is regulated in response to ambient ammonium levels via a mechanism distinct from asparaginase or glutamine synthetase. Glutamine synthetase and asparaginase, apparently repressed by high levels of all three nitrogen sources, are perhaps regulated by a common mechanism responding to intracellular nitrogen depletion, as evidenced by low cellular protein content.     

Toward a biophysical theory of organogenesis: Birefringence observations on regenerating leaves in the succulent,Graptopetalum paraguayense E. Walther

Polarity shifts occur during organogenesis. The histological criterion for polarity is the direction of cell division. The biophysical criterion is the orientation of reinforcing cellulose microfibrils which lie normal to the organ axis and which determine the preferred growth direction. Using cell pattern to deduce cell lineage, and polarized light to study cellulose alignment, both aspects of polarity were examined in the epidermis of regenerating G. paraguayense. In this system new leaves and a stem arise from parallel cell files on a mature leaf. Large (90°) shifts in polarity occur in regions of the epidermis to give the new organs radial symmetry in the surface plane (files radiating from a pole). Study of the shifts in the epidermis showed that, during certain stages, shifts in the division direction are accompanied by shifts in the cellulose deposition direction, as expected. The new cellulose orientation is parallel to the new cross wall. During normal organ extension, however, shifts in division direction do not bring on changes in cellulose pattern. Thus the coupling between the two kinds of polarity is facultative. This variable relation is used in a biophysical model which can account for the reorganization of cell file pattern and cellulose reinforcement pattern into the radial symmetry of the new organ.     

Axonal Transport of the Ca2+-Dependent Protein Modulator of 3'':5''-Cyclic-AMP Phosphodiesterase in the Rabbit Visual System

Water-soluble proteins were extracted from individual retinas, optic nerves, combined optic tracts and lateral geniculate bodies, and superior colliculi of rabbits at 1, 3, and 18 days after injection of [3H]leucine into the right eye. The Ca2+-dependent protein modulator of 3':5'-cyclic-AMP phosphodiesterase (calmodulin) was isolated from these samples by a two-step polyacrylamide gel electrophoresis procedure. An analysis of the radioactivity incorporated into the total soluble proteins and the calmodulin revealed that most of the calmodulin was axonally transported at a slow rate (2--4 mm/day) and represented about 0.45% of the total transported soluble protein.     

Substrate Specificity of [3H] Muscimol Binding to a Particulate Fraction of a Neuron-enriched Culture of Embryonic Rat Brain

Abstract: The effects of some GABA analogues and some drugs on the binding of [³H]muscimol (3.08 nM) to thoroughly washed subcellular particles prepared from a neuron-enriched culture of embryonic rat brain were examined using Na+-free Tris-citrate medium and a centrifugation method. Competition for [³H]muscimol binding sites by excess(10^?5 M) unlabelled GABA provided estimates of “specific” binding. In accord with in vivo neuropharmacological studies on GABA receptors and with in vitro studies on cerebral membrane preparations, [³H]muscimol binding was potently inhibited by muscimol itself (IC₅₀, 2.5 nM), GABA (1C₅₀, 43 nM), isoguvacine (IC₅₀, 61 nM), and 3-aminopropanesulphonic acid (IC₅₀, 160 nM), and less potently inhibited by the GABA antagonist bicuculline methobromide (IC₅₀, 800 nM). δ- Aminovaleric acid (IC₅₀, 2.6 μM), the glycinelp-alanine antagonist strychnine (IC₅₀, 6.6 μM), and the predominantly glial GABA uptake inhibitors β-alanine (IC₅₀, 23 μM) and p-proline (IC₅₀, 66 μM) also inhibited [³H]muscimol binding. Other inhibitors of Na+-dependent GABA uptake, (±)-nipecotic acid, L- 2,4-diaminobutyric acid, and guvacine, as well as picrotoxinin, were relatively inactive as inhibitors of [³H]muscimol binding (IC₅₀≥ 1 mM). In addition to revealing that GABA receptors are present on neuronal membranes before the formation of most synapses, this binding of [³H]muscimol that occurs to neuronal, but not to glial, membranes might be useful as a “neuronal marker” and for the further characterization and isolation of GABA receptors.     

Use of lambda pMu bacteriophages to isolate lambda specialized transducing bacteriophages carrying genes for bacterial chemotaxis.

A general method for constructing lambda specialized transducing phages is described. The method, which is potentially applicable to any gene of Escherichia coli, is based on using Mu DNA homology to direct the integration of a lambda pMu phage near the genes whose transduction is desired. With this method we isolated a lambda transducing phage carrying all 10 genes in the che gene cluster (map location, 41.5 to 42.5 min). The products of the cheA and tar genes were identified by using transducing phages with amber mutations in these genes. It was established that tar codes for methyl-accepting chemotaxis protein II (molecular weight, 62,000) and that cheA codes for two polypeptides (molecular weights, 76,000 and 66,000). Possible origins of the two cheA polypeptides are discussed.     

Lipoxygenase activity in apples in relation to storage and physiological disorders

Different methods for the preparation of active lipoxygenase (LOX) extracts from apples were compared. Highest activities were obtained using a 0.25 M phosphate buffer (pH 7) containing 1% Triton X-100 and 10^?2 M metabisulphite as extraction solvent. LOX activity during storage was investigated in the core, flesh, and peel. Activity was always highest in the core and peel. On storage, activity was increased in each part of the fruit but especially in the core and peel. Increase in LOX preceded the browning of the core. LOX may be responsible for the browning and may be concerned in the induction of superficial scald.