Triiodothyronine Is a High-Affinity Inhibitor of Amino Acid Transport System L1 in Cultured Astrocytes

Abstract: The relationship between the transport of thyroid hormones and that of amino acids was examined by measuring the uptake of amino acids that are characteristic substrates of systems L, A, and N, and the effect of 3,3',5-triiodo-L-thyronine (T₃) on this uptake, in cultured astrocytes. Tryptophan and leucine uptakes were rapid, Na⁺-independent, and efficiently inhibited by T₃ (half-inhibition at ∼ 2 μ M ). Two Na⁺-independent L-like systems (L₁ and L₂), common to leucine and aromatic amino acids, were characterized kinetically. System L₂ had a low affinity for leucine and tryptophan ( K _m= 0.3–0.9 m M ). The high-affinity system L₁ ( K _m∼ 10 μ M for both amino acids) was competitively inhibited by T₃ with a K _i of 2–3 μ M (close to the T₃ transport K _m). Several T₃ analogues inhibited system L₁ and the T₃ transport system similarly. Glutamine uptake and α-(methylamino)isobutyric acid uptake were, respectively, two and 200 times lower than tryptophan and leucine uptakes. T₃ had little effect on the uptakes of glutamine and α-(methylamino)isobutyric acid. The results indicate that the T₃ transport system and system L₁ are related.     

The effects of initial length and body curvature on shrinkage of juvenile Atlantic salmon during freezing

Fork length was measured in two groups of salmon parr (32–139 mm, frozen in a straight posture and frozen in a curved posture) before (L₁) and after (L₂) freezing and thawing. All the fish shrank. The decrease in length was significantly greater in the curved fish than the straight fish. The absolute reduction in length (L₁–L₂) was related directly to L₁, whereas the percentage reduction in length [(L₁–L₂)/L₁× 100] was related inversely to L₁.     

A MICRO-SCALE PROCEDURE FOR THE PREPARATION OF SUBCELLULAR FRACTIONS FROM INDIVIDUAL AUTONOMIC GANGLIA

Abstract— A microscale modification for the preparation of subcellular fractions employing milligram and submilligram amounts of neuronal tissue (brain nuclei and autonomic ganglia) is described.
Electron microscope characterization and enzymic studies were carried out on the six subcellular fractions of sympathetic ganglia of cat thus prepared.
The synaptosomal preparations obtained from individual ganglia were poorer in their nerve ending content than those obtained from brain by previous investigators. The highest RSA for AChE was found in layer L₂ which was rich in membranes and vesicle components. ChAc activity was also highly concentrated in layers L₂ and L₃ (membranes, nerve ending-like particles, mitochondria and 'ghosts'). MAO activity was particularly high in the layers L₄ and L₅ which contained a large number of mitochondria. Layer L₁ (membrane fragments) and particularly layer L₆ which contained mainly collagen fibres, were low in activity of all three enzymes.
After preganglionic denervation, both ChAc and AChE activities were significantly reduced in the purest nerve ending fraction, L₃ while MAO activity was practically unchanged.     

Evaluation of Serangium n. sp. (Col., Coccinellidae), a predator of Bemisia tabaci (Hom., Aleyrodidae) on cassava

Abstract  The potential of a new, previously unidentified Serangium species (Col., Coccinellidae) to control the high Bemisia tabaci (Gennadius) (Hom., Aleyrodidae) populations on cassava was evaluated. Field and laboratory studies were carried out to determine the abundance and feeding capacity of this Serangium species feeding on B. tabaci on cassava. Serangium nymphs and adults were most abundant in cassava fields late in the season, rising sharply from 5 months after planting (MAP) to a peak at 7–8 MAP. Pre-imaginal development averaged 21.2 days and was longest in eggs and shortest in the L₁ instar. Mean total prey consumption of immature Serangium increased with the stage of development with the lowest consumption in the L₁ instar and highest in the L₄ instar. Mean daily consumption was lowest on the first day after hatching in the L₁ instar and rose to a peak on the 13th day after hatching in the L₄ instar. Each Serangium larva consumed a mean of over 1000 nymphs during its entire development. These results have demonstrated the potential of this Serangium species to control B. tabaci populations on cassava.     

Differences in the Cyclic AMP-Dependent Phosphorylation of Plasma Membrane Proteins of Differentiated and Undifferentiated L6 Myogenic Cells

Differences in the cyclic AMP-dependent plasma membrane phosphorylation system of undifferentiated and differentiated L₆ myogenic cells have been detected. Endogenous plasma membrane protein phosphorylation in undifferentiated L₆ myoblasts was stimulated more than three fold by 5 × 10⁻⁵ M cyclic AMP, whereas no statistically significant cyclic AMP-dependent phosphorylation of endogenous plasma membrane proteins was observed in differentiated L₆ cells. In undifferentiated cells cyclic AMP promoted the phosphorylation of several proteins, the most prominent of which had a molecular weight of 110,000. In differentiated cells cyclic AMP did not selectively promote the phosphorylation of specific plasma membrane proteins. Both differentiated and undifferentiated L₆ cells, however, contain a cyclic AMP-dependent protein kinase capable of catalyzing the phosphorylation of exogenous substrates, such as histone f₂b. Therefore, the data show that differentiation in L₆ cells is associated with a selective change in the activity of a plasma membrane cyclic AMP-dependent protein kinase which employs endogenous membrane proteins as substrate.     

Duration times of the immature stages of Cacopsylla pyri L. (Hom., Psyllidae), estimated under field conditions, and their relationship to ambient temperature

The duration of the immature stages of Cacopsylla pyri L. was studied under field conditions by artificially infesting pear branches on several dates during the year. The duration of the egg stage decreased from winter to summer, as the season progresses and temperature rises, and slightly increased in September. It ranged from 27.4 to 6.7 days. The same trend was observed in the duration of the first three larval stages (L₁₋₃) which ranged from 18.8 to 10.3 days. For eggs deposited during the period February–August the duration of the last two larval stages (L₄₋₅) ranged from 17.5 to 12.1 days. However, the duration of L₄₋₅ developed from eggs deposited in September and which give rise to winter-form adults were the longest observed. The rate of egg development was related to average ambient temperature with a highly significant linear relationship. This relationship indicates that the egg stage requires a constant number of 158.9 (SD = 5.0) of day-degrees above an average temperature of 2.31°C to complete development. The rate of development both of L₁₋₃ and L₄₋₅ were related to average ambient temperature with curvilinear relationships. These relationships indicate a proportional increase in the developmental rate as temperature rises between 10–22°C. At the higher average temperatures that occurred in the summer experiments (24–27°C) the acceleration of development of L₁₋₃ is reduced and the developmental rate of L₄₋₅ decreases. The developmental rates of L₄₋₅ developed from eggs deposited in September did not follow the established relationship with temperature and they were lower than those in the other periods of the year with the same average temperature.     

Physiological function of water channels as affected by salinity in roots of paprika pepper

The sap flow (Jv) and the osmotic pressure-dependent hydraulic conductance (L₀) of detached exuding root systems from paprika pepper plants (cv. Albar) were measured. Plants stressed with NaCl (30 m M ) and with six times the macronutrients of the Hoagland nutrient solution (6×HNS) were compared with controls grown in complete Hoagland nutrient solution. Jv of +NaCl and +6×HNS plants decreased markedly, but recovered to values similar to those of controls after removal of the treatments. Hydraulic conductance L₀ was always less in NaCl plants than in controls and 6×HNS. A total increase in the ion concentration of the xylem (except Na⁺ and Cl⁻) was observed with both treatments. In control and 6×HNS plants, HgCl₂ treatment (50 μ M ) caused a sharp decline in L₀ to values similar to those of NaCl-stressed roots, but were restored by treating with 5 m M dithiothreitol (DTT). However, in NaCl roots only a slight effect of Hg²⁺ and DTT was observed. In each treatment, there was no difference in the flux of K⁺ into the xylem after HgCl₂ and DTT application. The results suggest that NaCl decreased L₀ of the roots by reducing either the activity or abundance of Hg-sensitive water channels. The putative reduction in water-channel function of NaCl-treated plants did not seem to be due to the osmotic effect.     

Nucleotide and deduced amino acid sequence of the Rhodobacter sphaeroides gene encoding form II ribulose-1,5-bisphosphate carboxylase/oxygenase and comparison with other deduced form I and II sequences

The nucleotide sequence for the Rhodobacter sphaeroides form II ribulose 1,5-bisphosphate carboxylase/oxygenase was determined. The deduced product is highly homologous with the form II-like enzyme of Rhodospirillum rubrum , but appears to be more distantly related to the large subunit of the L₈S₈ enzyme found in autotrophic bacteria, cyanobacteria and higher plants. Several regions highly conserved among L₈S₈ and L_X enzymes correspond with regions previously implicated in catalytic activity and subunit interactions. An imperfect palindrome and a stem loop structure were identified in the 5' and 3' flanking sequences, respectively, of R. sphaeroides rbpL .     

Cloning and expression in Escherichia coli of a species-specific Chlamydia trachomatis outer membrane antigen

Abstract A gene library of Chlamydia trachomatis serovar L₂ (strain 434) was constructed in Escherichia coli using plasmid pBR322. Amongst 200 recombinants we have identified and characterized a recombinant E. coli that expresses a protein antigen of M _r 74 000 similar in size to an outer membrane antigen produced by elementary bodies of C. trachomatis . Immunologically, the molecule synthesised by E. coli has the same specificity as the protein encoded by serovar L₂. A 1.8 kb DNA fragment from the recombinant insert, used as a hybridization probe, confirmed the species specificity of this clone at the gene level.     

Boric acid and salinity effects on maize roots. Response of aquaporins ZmPIP1 and ZmPIP2, and plasma membrane H+-ATPase, in relation to water and nutrient uptake

Under saline conditions, an optimal cell water balance, possibly mediated by aquaporins, is important to maintain the whole-plant water status. Furthermore, excessive accumulation of boric acid in the soil solution can be observed in saline soils. In this work, the interaction between salinity and excess boron with respect to the root hydraulic conductance (L₀), abundance of aquaporins (ZmPIP1 and ZmPIP2), ATPase activity and root sap nutrient content, in the highly boron- and salt-tolerant Zea mays L. cv. amylacea, was evaluated. A downregulation of root ZmPIP1 and ZmPIP2 aquaporin contents were observed in NaCl-treated plants in agreement with the L₀ measurements. However, in the H₃BO₃-treated plants differences in the ZmPIP1 and ZmPIP2 abundance were observed. The ATPase activity was related directly to the amount of ATPase protein and Na⁺ concentration in the roots, for which an increase in NaCl- and H₃BO₃+ NaCl-treated plants was observed with respect to untreated and H₃BO₃-treated plants. Although nutrient imbalance may result from the effect of salinity or H₃BO₃ alone, an ameliorative effect was observed when both treatments were applied together. In conclusion, our results suggest that under salt stress, the activity of specific membrane components can be influenced directly by boric acid, regulating the functions of certain aquaporin isoforms and ATPase as possible components of the salinity tolerance mechanism.     

Effect of light pulses in early scotophase on resetting of the night-measuring diapause clock of the Indian meal moth, Plodia interpunctella

Abstract.  The Indian meal moth, Plodia interpunctella Hübner (Lepidoptera: Pyralidae) may enter diapause in the last larval instar in response to the photoperiod during the preceding instars. An hourglass-type photoperiodic clock may measure night length for this purpose. The present study describes the resetting of the hourglass by light pulse(s) in the early scotophase and by scanning the subsequent clock phase by another light pulse (P). When the lights-off time of a first light pulse is fixed at 4 h after dusk under LD 4 : 20 h and LD 6 : 18 h photoperiods and its duration is increased from 1 to 3 h, the critical night length (CNL) from dawn is decreased, but that from dusk to P increases. A 3-h first light pulse efficiently resets the time measuring system. If this 3-h light pulse is split into two 1-h light pulses (L₁ and L₂) by 1 h of darkness, the dark-time measuring function appears to be impeded and CNL from P to dawn disappears, but that from L₂ to P is expressed. This indicates that the receptivity to light pulses varies among individual insects.     

Rapid isolation of genes from bacterial lambda libraries by direct polymerase chain reaction screening

Abstract Ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) was purified from an obligately autotrophic hydrogen-oxidizing bacterium, Hydrogenovibrio marinus MH-110. The protein has a M _r value of approximately 110 000, and is composed of two identical subunits of 55 000. To our knowledge, the existence of L₂-form RubisCO in a chemolithoautotrophic bacterium is first reported in this paper. The N-terminal amino acid sequence determination of the purified enzyme showed high homology with those of the L₂-form RubisCO of Rhodospirillum rubrum and the L _x -form RubisCO from Rhodobacter sphaeroides .     

Laboratory studies on the effects of pollen from Bt-maize on larvae of some butterfly species

Abstract:   Three lepidopteran species were tested to determine their susceptibility against the ingestion of pollen from genetically modified maize plants. To prove the existence of dose–response relations between the applied amount of pollen (Bt-maize) and the damages on the tested larvae, a method was developed which makes it possible to feed caterpillars with defined amounts of pollen. If their food plants were contaminated with pollen of a cultivar of the Bt-176 maize-line Pieris brassicae , Pieris rapae and Plutella xylostella -larvae fed less, grew more slowly and showed a higher mortality than caterpillars of an untreated control group. The 50% lethality (LD₅₀)-values were calculated for P.xylostella (L₄) with 19.2, for P. rapae (L₂) with 39.0 and also for P. brassicae (L₂) with 139.2 pollen of the transgenic maize Pactol CB. Studies with P. brassicae -caterpillars of different larval stages indicated, that older individuals showed a higher tolerance against pollen from Bt-maize than younger ones. It must be stated on the basis of the present studies, that ingestion of non-transgenic maize pollen has neither a positive nor a negative effect on caterpillars. It becomes clear from the information presented here that it is still difficult to make general statements about the endangering of butterflies, arising from cultivation of genetically modified maize lines. Further investigations on this issue are needed. Initially, the LD₅₀-values concerning the larvae of certain butterfly species have to be determined to anticipate the risks, and in addition the distances between habitats with caterpillar host plants and maize fields, and the abundance of these plants have to be considered.     

Hormonal regulation of pupation in the codling moth, Laspeyresia pomonella

ABSTRACT. According to the different reactions to the juvenoid Altosid®, the last larval instar (L₅) of Laspeyresia pomonella (L.) (Tortricidae) reared under 'long day' conditions (constant light) was subdivided into three sensitive phases: an additional larval instar, a larval–pupal intermediate, or a pupa. Under short day conditions, the prothoracotropic effect of juvenile hormone (JH) in L₅, which have a continuous high titre of JH during the whole instar, indicated that it is not a particular titre of JH but a rise in the titre that can induce the production of moulting hormone. Neck-ligation experiments showed that JH acts not directly on the prothoracic glands but via the head, probably via the neurosecretory system. The meaning of the JH-peak in mature L₅ reared under long days was determined either by injections with the anti-JH, precocene II, in combination with applications of Altosid, or by forcing precocene-treated larvae to a precocious moult by injecting them with ecdysterone. Precocene delayed, and JH accelerated pupation if administered 4.5 days after the L₅ -moult. JH was also found to stimulate the growth and differentiation of the imaginal discs. Moulting hormone in long-days reared insects was detected one day after the larvae had spun their cocoon, with a maximum on the second day after spinning. The hormone was also present in freshly moulted pupae. Neck-ligation of mature larvae indicated that the delay between activation of the prothoracic glands and the production of an effective amount of moulting hormone is less than one day.     

Biochemical characterization of pectate lyases produced by fluorescent pseudomonads associated with spoilage of fresh fruits and vegetables

An improved method for purification of pectate lyases (PLI and PLII) from culture fluids of Pseudomonas fluorescens CY091 and Ps. viridiflava PJ-08-6 by using a phosphocellulose cation exchanger was described. Analysis of purified PLI and PLII by sodium dodecyl sulphate-polyacrylamide and isoelectric focusing gel electrophoresis revealed that both enzymes had been purified to near homogeneity. Optimal Ca²⁺ concentration required for PLI and PLII activity was determined to be 0·5 mmol l⁻¹. The Ca²⁺ requirement could not be replaced by other metal cations such as Mg²⁺, Cu²⁺, Zn²⁺, Fe³⁺ and Co²⁺. Optimal pH for activity was determined to be between 8·5 and 9·0. The K _m values for sodium polygalacturonate were 1·28 and 1·11 mg ml⁻¹ for PLI and PLII, respectively. Both PLI and PLII were stable at low temperatures (25°C or below) for at least 1 month. However, at 37°C, the activity decreased 50% in 36 h. Optimal temperatures for activity were estimated to be 46° and 52°C for PLI and PLII, respectively. Thermal stability of both enzymes at elevated temperatures (48°C or higher) increased when CaCl₂ or a positively charged molecule such as polylysine was present, but decreased when polygalacturonate or a negatively charged molecule such as heparin was present. PLI and PLII exhibit differential degrees of sensitivity to group-specific inhibitors, including iodoacetic acid and diethylpyrocarbonate. This result suggests that both sulphydryl and imidazole groups are important for the catalytic function of PLI and PLII.     

Lysophospholipase L2 of Vibrio cholerae O1 affects cholera toxin production

Abstract The implication in cholera toxin (CT) production of the newly identified gene, lypA , that encodes the lysophospholipase L₂ of Vibrio cholerae , was investigated. Introduction of lypA into the V. cholerae O1 mutant (NF404), which has a Tn5-insertion in lypA and has lost CT as well as haemolysin production, restored the lysophospholipase activity and CT production but not the haemolytic activity. Inactivation of the lypA gene of the wild-type strain by chromosomal integration of a plasmid containing a portion of the lypA gene decreased the lysophospholipase L₂ activity and the production of CT but not the haemolytic activity. Furthermore, constructed mutants of El Tor-biotype and Classical-biotype strains which have a defective lypA failed to produce CT and exhibited decreased enterotoxicity in the ligated rabbit ileal loop test. These results suggest that lypA is possibly required for the expression of CT and may play a role in pathogenicity of V. cholerae .     

Growth response of pikeperch larvae in relation to body size and zooplankton abundance

The most critical period at onset of feeding in pikeperch Stizostedion lucioperca is short (<5 days at 20° C). The larvae are sensitive to prey density during the first week of exogenous feeding. First-feeding larvae of 6.5 mm total length ( L_T ) needed prey densities of >585 prey l⁻¹ to maintain mass (C_maint), whereas 5 days older larvae of 7 mm L _T C_maint= 55 prey l⁻¹ and for 11 mm L _T larvae C_maint<10 prey 1⁻¹. Changes in specific growth rates for larvae of 7 and 11 mm were similar to a type-II functional response curve reaching a specific growth rate of 26 and 30% day⁻¹, respectively, at 1000 prey l⁻¹, whereas the 6·5 mm larvae showed a linear growth response reaching a specific growth rate of only 9% day ⁻¹ at 1000 prey l⁻¹. The results suggest that prey density is a limiting factor, which might contribute to the high variation between year-class Strengths.     

Response of net photosynthesis in bean (Phaseolus vulgaris) leaves to the elevation of the partial pressures of oxygen and carbon dioxide

Bean ( Phaseolus vulgaris L. cv. Golden Saxa) plants were grown under low artificial light or under natural daylight. The rate of net photosynthesis (P_N) was measured at: CO₂ partial pressure, p(CO₂), of 0.03, 0.09 or 0.15 kPa; O₂ partial pressure, p(O₂), of 2, 21 or 31 kPa and at light intensities of 350 or 1000 μmol m⁻² s⁻¹ (photosynthetically active radiation). In plants which had been grown under natural light, stimulation of P_N at 21 kPa p(O₂) was found only at elevated p(CO₂) and high light. It is proposed that this phenomenon is dependent on a high capacity of the photosynthetic apparatus to regenerate ribulose 1.5-bisphosphate.     

Characterisation of a non-haemolytic enterotoxin complex from Bacillus cereus isolated after a foodborne outbreak

Abstract Three enterotoxic components have been isolated from a strain of Bacillus cereus which was involved in a large food poisoning outbreak in Norway in 1995. The components were purified by chromatography on three different columns. Three proteins of 39, 45 and 105 kDa, respectively, were found to be necessary for maximum cytotoxicity. The amino acid N-terminal sequences of the 39 and 45 kDa proteins were determined. The 45 kDa component was the same protein as the main antigen detected in the Bacillus Diarrhoeal Enterotoxin Visual Immunoassay (Tecra). The 39 kDa protein showed some similarity to the l₁ protein of haemolysin BL from B. cereus . Furthermore, the three toxic components were all recognised by a polyclonal antiserum reported to detect enterotoxin from B. cereus . The proteins were different from the B- and L₂-components of haemolysin BL, previously suggested to be a primary virulence factor, and had no detectable haemolytic activity.     

THE RELEASE OF COPPER-COMPLEXING LIGANDS BY THE BROWN ALGA FUCUS VESICULOSUS (PHAEOPHYCEAE) IN RESPONSE TO INCREASING TOTAL COPPER LEVELS

The growth of Fucus vesiculosus L. germlings in chemically defined culture media containing a range of Cu concentrations (20–1000 nM) was monitored simultaneously with measurement of the Cu speciation in the media by competitive equilibrium-adsorptive cathodic stripping voltammetry. Fucus vesiculosus germlings were found to exude Cu-complexing ligands with conditional stability constants of the order of 1.6 × 10¹¹. Ligand concentrations increased with increasing total dissolved Cu concentrations (Cu_T) until a concentration of 500–800 neq Cu·L⁻¹ was reached. Concentrations of the ligand exceeded Cu_T in treatments containing 20 and 100 nM Cu, were similar to Cu_T in the 500-nM Cu treatment, but were less than Cu_T in the 1000-nM treatment. Therefore, [Cu²⁺] were calculated to be at concentrations of 10⁻¹¹− 10⁻¹⁰ M in the 20- and 100-nM treatments, 10⁻⁹ M in the 500-nM treatment, and 10⁻⁷ M in the 1000-nM treatment. Growth rates were lowest at Cu²⁺ concentration > 10⁻⁹. These results are discussed within the context of the potential roles for exuded copper-complexing ligands.