Tetrazolium Reduction by Guard Cells in Abaxial Epidermis of Vicia faba: Blue Light Stimulation of a Plasmalemma Redox System

The stomata in the abaxial epidermis of Vicia faba were examined for the location of redox systems using tetrazolium salts. Three distinct redox systems could be demonstrated: chloroplast, mitochondrial, and plasmalemma. The chloroplast activity required light and NADP. Mitochondrial activity required added NADH and was suppressed by preincubation with KCN. The plasmalemma redox system in guard cells also required NADH, but was insensitive to KCN and was stimulated by blue light. The involvement of an NADH dehydrogenase in the blue light stimulated redox system in guard cells was suggested by the sensitivity to plantanetin, an inhibitor of NADH dehydrogenase. The redox system of mitochondria was the most active followed by that of plasmalemma. The activity of chloroplasts was the least among the three redox systems. The plasmalemma mediated tetrazolium reduction was stimulated by exogenous flavins and suppressed by Kl or phenylacetate, inhibitors of flavin excitation. We therefore conclude that an NADH-dependent, flavin mediated electron transport system, sensitive to blue light, operates in the plasmalemma of guard cells.     

Queen succession in the primitively eusocial tropical waspRopalidia marginata (Lep.) (Hymenoptera: Vespidae)

Females of the primitively eusocial wasp Ropalidia marginata can be classified into three behavioral groups: Sitters, Fighters, and Foragers. It has been speculated that both Sitters and Fighters may be hopeful queens and that the Foragers may have little or no opportunities for direct reproduction. Here we show that in 9 of 12 queen-removal experiments where such a behavioral differentiation could be discerned, the individual that became a queen (the potential queen) was a Sitter in 6 cases, a Fighter in 2 cases, and a Forager in only 1 case. Although potential queens spent significantly more time absent from the nest and showed significantly higher rates of dominance behavior compared to the mean values for nonqueens in their colonies, they were intermediate with respect to all behaviors and age when compared to the range of values for nonqueens in their colonies. Potential queens were not necessarily the highest-ranking individuals among the nonqueens. The pattern of queen succession in this species appears to be quite different from the temperate pattern, where an old and active forager of high dominance rank is the potential queen. Although somewhat similar to the tropical pattern of a relatively younger female that has performed relatively little foraging being the potential queen, it is perhaps more accurate to describe the potential queens of R. marginata as unspecialized intermediates.     

Structure and function of disk aggregates of the coat protein of tobacco mosaic virus

Experiments have been carried out on the coat protein of tobacco mosaic virus (TMVP) to test for the occurrence of the previously postulated RNA-induced direct switching, during in vitro assembly of tobacco mosaic virus (TMV), of the subunit packing from the cylindrical bilayer disk to the virus helical arrangement. No evidence was found for such RNA-induced switching and no evidence for the direct participation of the bilayer disk in either the nucleation or elongation phases of the in vitro virus assembly. Instead, virus assembly proceeds by an initiation step involving the binding of the RNA to the previously characterized two-plus turn helical aggregate that is formed from small oligomers of subunits. However, a bilayer disk, which has been characterized in high ionic strength crystals, has been observed in low ionic strength virus assembly solutions only as a transient species upon depolymerization of dimers of bilayer disks formed in solution at high ionic strength, and not as an equilibrium species of TMVP.     

Photosynthetic Units and Carbon Assimilation in Leaves of Grain Sorghum under Different Light Intensities

The Cyt f and P700 contents in leaves of three Sorghum, varietieswere measured, in relation to their carbon assimilation, underdifferent light intensities during growth. At the maximum irradiationused (1,800 µE m^–2 s^–1) the ratio of P700to Cyt f was close to unity, whereas under low irradiation (450µE m^–2 s^–1) the ratio of P700 to Cyt f rangedfrom two to three. A strikingly positive correlation existedbetween the P700 contents of the leaves and their rates of carbondioxide fixation, dry matter production and Cyt f contents,only when the plants were grown under high light intensities.The P700 content of the leaves in plants grown under low irradiationwas unrelated to the contents of Cyt f. Thus, at a high lightintensity there is a close relationship between the Cyt f andP700 levels, but at low intensities the amounts of electroncarriers and the reaction centre are independent. (Received March 7, 1983; Accepted August 24, 1983)     

Beneficial interaction between photosynthesis and respiration in mesophyll protoplasts of pea during short light-dark cycles

The respiratory uptake or photosynthetic evolution of oxygen by mesophyll protoplasts of pea ( Pisum sativum L. cv. Arkel) were monitored during successive short. (3–5 min) cycles of darkness and illumination. The rate of respiration was nearly doubled after 3–4 short periods of illumination while there was a 15–20% enhancement in photosynthesis with cycles of illumination and darkness preceding illumination. Such interaction between photosynthesis and respiration was statistically significant when bicarbonate was present in the reaction medium. The inhibitors of photosynthesis [3(3,4–dichlorophenyl)-l,l-dimethylurea (DCMU), glyceraldehyde] decreased respiration after periods of illumination, whereas inhibitors of respiratory electron transport (Rotenone, antimycin A, NaN₃) suppressed photosynthesis, as well. We suggest that a rapid beneficial interaction exists between photosynthesis and respiration in protoplasts, even during short cycles of light and darkness.     

l-[3H]Nitroarginine and l-[3H]Arginine Uptake into Rat Cerebellar Synaptosomes: Kinetics and Pharmacology

Abstract: Characteristics of the transport of the nitric oxide synthase substrate l -arginine and its inhibitor, N ^G-nitro- l -arginine ( l -NOARG), into rat cerebellar synaptosomes were studied. Uptake of both l -arginine and l -NOARG was linear with increasing amount of protein (up to 40 µg) and time of incubation (up to 5 min) at 37°C. Uptake of both compounds reached a steady state by 20 min. Maximal uptake of l -NOARG (650 pmol/mg of protein) was three to four times higher than that of l -arginine (170 pmol/mg of protein). l -NOARG uptake showed biphasic kinetics ( K _{m 1} = 0.72 m M , V _{max 1} = 0.98 nmol/min/mg of protein; K _{m 2} = 2.57 m M , V _{max 2} = 16.25 nmol/min/mg of protein). l -Arginine uptake was monophasic with a K _m of 106 µ M and a V _max of 0.33 nmol/min/mg of protein. l -NOARG uptake was selectively inhibited by l -NOARG, N ^G-nitro- l -arginine methyl ester, and branched-chain and aromatic amino acids. l -Alanine and l -serine also inhibited l -NOARG uptake but with less potency. Uptake of l -arginine was selectively inhibited by N ^G-monomethyl- l -arginine acetate and basic amino acids. These studies suggest that in rat cerebellar synaptosomes, l -NOARG is transported by the neutral amino acid carrier systems T and L with high affinity, whereas l -arginine is transported by the basic amino acid carrier system y⁺ with high affinity. These data indicate that the concentration of competing amino acids is an important factor in determining the rates of uptake of l -NOARG and l -arginine into synaptosomes and, in this way, may control the activity of nitric oxide synthase.     

Molecular biology of C4 phosphoenolpyruvate carboxylase: Structure,regulation and genetic engineering

Three to four families of nuclear genes encode different isoforms of phosphoenolpyruvate (PEP) carboxylase (PEPC): C₄-specific, C₃ or etiolated, CAM and root forms. C₄ leaf PEPC is encoded by a single gene (ppc) in sorghum and maize, but multiple genes in the C₄-dicot Flaveria trinervia. Selective expression of ppc in only C₄-mesophyll cells is proposed to be due to nuclear factors, DNA methylation and a distinct gene promoter. Deduced amino acid sequences of C₄-PEPC pinpoint the phosphorylatable serine near the N-terminus, C₄-specific valine and serine residues near the C-terminus, conserved cysteine, lysine and histidine residues and PEP binding/catalytic sites. During the PEPC reaction, PEP and bicarbonate are first converted into carboxyphosphate and the enolate of pyruvate. Carboxyphosphate decomposes within the active site into Pi and CO₂, the latter combining with the enolate to form oxalacetate. Besides carboxylation, PEPC catalyzes a HCO₃ ^--dependent hydrolysis of PEP to yield pyruvate and Pi. Post-translational regulation of PEPC occurs by a phosphorylation/dephosphorylation cascade in vivo and by reversible enzyme oligomerization in vitro. The interrelation between phosphorylation and oligomerization of the enzyme is not clear. PEPC-protein kinase (PEPC-PK), the enzyme responsible for phosphorylation of PEPC, has been studied extensively while only limited information is available on the protein phosphatase 2A capable of dephosphorylating PEPC. The C₄ ppc was cloned and expressed in Escherichia coli as well as tobacco. The transformed E. coli produced a functional/phosphorylatable C₄ PEPC and the transgenic tobacco plants expressed both C₃ and C₄ isoforms. Site-directed mutagenesis of ppc indicates the importance of His¹³⁸, His⁵⁷⁹ and Arg⁵⁸⁷ in catalysis and/or substrate-binding by the E. coli enzyme, Ser⁸ in the regulation of sorghum PEPC. Important areas for further research on C₄ PEPC are: mechanism of transduction of light signal during photoactivation of PEPC-PK and PEPC in leaves, extensive use of site-directed mutagenesis to precisely identify other key amino acid residues, changes in quarternary structure of PEPC in vivo, a high-resolution crystal structure, and hormonal regulation of PEPC expression.Abbreviations OAA oxalacetate - PEP phosphoenolpyruvate - PEPC PEP carboxylase - PEPC-PK PEPC-protein kinase - PPDK pyruvate, orthophosphate dikinase - Rubisco ribulose 1,5-bis-phosphate carboxylase/oxygenase - CAM Crassulacean acid metabolism     

Suppression of Oxygen Evolving System in Spinach Chloroplast Membranes due to Release of Manganese on Exposure to Osmotic Stress in vitro in Presence of Magnesium

When spinach chloroplast membranes were exposed to osmotic stress in vitro, by incubation in 1.0 M sorbitol + 10 mM MgCl₂ their oxygen evolving system was suppressed. The possible reasons for such inactivation of PS II mediated oxygen evolution were examined. There were conformational changes in the chloroplast membranes, as indicated by their absorption spectra. The pattern of sensitivity to DCMU was not altered. The sensitivity of PS II to water stress remained, even after a pre-wash treatment with NaCI (which removed 18 and 24 kD proteins) but not when the thylakoids were pretreated with NH₂0H or CaCl₂ (removed manganese and 33 kD). The manganese content of thylakoid membranes was markedly reduced under osmotic stress in presence of magnesium. We suggest that exposure of chloroplasts to 1.0 M sorbitol in presence of Mg₂₊ released manganese from thylakoid membranes, thereby leading to a suppression in oxygen evolution.     

Uptake and metabolism of glutamate and aspartate by astroglial and neuronal preparations of rat cerebellum

Astrocytes, neuronal perikarya and synaptosomes were prepared from rat cerebellum. Kinetics of high and low affinity uptake systems of glutamate and aspartate, nominal rates of¹⁴CO₂ production from [U–¹⁴C]glutamate, [U–¹⁴C]aspartate and [1–¹⁴C]glutamate and activities of enzymes of glutamate metabolism were studied in these preparations. The rate of uptake and the nomial rate of production of¹⁴CO₂ from these amino acids was higher in the astroglia than neuronal perikarya and synaptosomes. Activities of glutamine synthetase and glutamate dehydrogenase were higher in astrocytes than in neuronal perikarya and synaptosomes. Activities of glutaminase and glutamic acid decarboxylase were observed to be highest in neuronal perikarya and synaptosomes respectively. These results are in agreement with the postulates of theory of metabolic compartmentation of glutamate while others (presence of glutaminase in astrocytes and glutamine synthetase in synaptosomes) are not. Results of this study also indicated that (i) at high extracellular concentrations, glutamate/aspartate uptake may be predominantly into astrocytes while at low extracellular concentrations, it would be into neurons (ii) production of -ketoglutarate from glutamate is chiefly by way of transamination but not by oxidative deamination in these three preparations and (iii) there are topographical differences glutamate metabolism within the neurons.