Phosphoenolpyruvate carboxylase in Hydrilla plants with varying CO2 compensation points

Incubation of the submersed aquatic macrophyte, Hydrilla vertieillata Royle, for up to 4 weeks in growth chambers under winter-like or summer-like conditions produced high (130 to 150 μl CO₂/l) and low (6 to 8 μl CO₂/l) CO₂ compensation points (Γ), respectively. The activities of both ribulose bisphosphate (RuBP) and phosphoenolpyruvate (PEP) carboxylases increased upon incubation but the major increase was in the activity of PEP carboxylase under the summer-like conditions. This reduced the ratio of RuBP/PEP carboxylases from 2.6 in high Γ plants to 0.2 in low Γ plants. These ratios resemble the values in terrestrial C₃ and C₄ species, respectively. Kinetic measurements of the PEP carboxylase activity in high and low Γ plants indicated the Vmax was up to 3-fold greater in the low Γ plants. The Km (HCO₃ ^-) values were 0.33 and 0.22 mM for the high and low Γ plants, respectively. The Km (PEP) values for the high and low Γ plants were 0.23 and 0.40 mM, respectively; and PEP exhibited cooperative effects. Estimated Km (Mg²⁺) values were 0.10 and 0.22 mM for the high and low Γ plants, respectively. Malate inhibited both PEP carboxylase types similarly. The enzyme from low Γ plants was protected by malate from heat inactivation to a greater extent than the enzyme from high Γ plants. The results indicated that C₄ acid inhibition and protection were not reliable methods to distinguish C₃ and C₄ PEP carboxylases. The PEP carboxylase from low Γ plants was inhibited more by NaCl than that from high Γ plants. These analyses indicated that Hydrilla PEP carboxylases had intermediate characteristics between those of terrestrial C₃ and C₄ species with the low Γ enzyme being different from the high Γ enzyme, and closer to a C₄ type.     

Cloning, sequencing and characterization of the Saccharomyces cerevisiae URA7 gene encoding CTP synthetase

Summary The URA7 gene of Saccharomyces cerevisiae encodes CTP synthetase (EC 6.3.4.2) which catalyses the conversion of uridine 5-triphosphate to cytidine 5-triphosphate, the last step of the pyrimidine biosynthetic pathway. We have cloned and sequenced the URA 7 gene. The coding region is 1710 by long and the deduced protein sequence shows a strong degree of homology with bacterial and human CTP synthetases. Gene disruption shows that URA7 is not an essential gene: the level of the intracellular CTP pool is roughly the same in the deleted and the wild-type strains, suggesting that an alternative pathway for CTP synthesis exists in yeast. This could involve either a divergent duplicated gene or a different route beginning with the amination of uridine mono- or diphosphate.     

Adventitious bud regeneration from leaf expiants of the shrubby ornamental honeysuckle,Lonicera nitida Wils. cv. ‘Maigrün’: effects of thidiazuron and 2,3,5-triiodobenzoic acid

Different combinations of auxins and cytokinins were employed to assess the regeneration capacity from in vitro leaf explants of Lonicera nitida Wils. cv Maïgrün. A high frequency of rhizogenesis was noticed, with 2.3 M thidiazuron plus 2.9 M indole-3-acetic acid as the only hormonal combination to support caulogenic responses. Increasing thidiazuron concentration and/or suppressing auxin did not improve caulogenesis. Combining thidiazuron with 2,3,5-triiodobenzoic acid produced a dramatic increase in the percentage of caulogenic explants. A maximum of 74% of adventitious bud forming explants was obtained with 2.3 M thidiazuron plus 20 M 2,3,5-triiodobenzoic acid. Buds were often in a rosette form and were vitreous, so that shoot elongation was difficult to obtain. The effect of the duration of the 2,3,5-triiodobenzoic acid treatment on shoot elongation was investigated.Abbreviations BAP benzylaminopurine - 2,4-D 2,4-dichlorophenoxyacetic acid - GA3 gibberellic acid - IAA indole-3-acetic acid - IBA indole-3-butyric acid - 2 IP 2-isopentenyladenine - MS Murashige and Skoog - NAA -naphthylacetic acid - TDZ N-phenyl-N-1,2,3-thidiazol-5-ylurea (thidiazuron) - TIBA 2,3,5-triiodobenzoic acid - Z zeatin     

Proliferation,differentiation and ciliary beating of human respiratory ciliated cells in primary culture

Summary The growth, differentiation, ciliary beating pattern and frequency of human respiratory ciliated cells in primary culture were studied by scanning and transmission electron microscopy and by videomicroscopy. The epithelial cells were obtained as outgrowth from explants of adult nasal polyps. When the explants were grown on type-I and type-IV collagen substrates in a standard serum-free, hormone-supplemented medium, a high percentage of ciliated cells (range 29±5% to 37±6%) was present within 2 days of culture. After 5 days of culture, the percentage of ciliated cells near the explant was 51±5%. Most of the cultured ciliated cells (85%) were characterized by individual cilia showing a coordinated movement during the beat cycle and a beating frequency (13.3±1.3 Hz) similar to that reported in vivo. In the other 15% of the ciliated cells, the dyskinetic cilia were aggregated into clumps and characterized by a rigid and planar bending movement and a lower (P<0.01) beating frequency (10.7±1.4 Hz). It is suggested that the latter type of cell, already described during fetal development, might be an intermediate type of ciliated cell which appears temporarily during the surface respiratory epithelial differentiation.     

Cellular and biochemical indicators assessing the quality of a marine environment

Environmental stressors as well as the direct or combined effects of pollutants could be harmful to the populations living in a marine environment and the reproductive and nutritive processes could be impaired in a deteriorating environment. Sublethal effects of pollutants were studied in the blue mussel Mytilus edulis L., a good bioaccumulator of contaminants. Blue mussels of 3.5 cm were sampled on a rocky substrate at Pointe-Mitis (48° 40N, 68° 02W) along the coast of the St-Lawrence estuary. Mussels were placed in experimental tanks, fed, supplemented with mineral salts and continuous sea water flow and kept 72 h before the exposure to 0.01 µg l^–1 and 0.3 µg l^–1 methylmercury hydroxide in the presence or absence of selenium, at a concentration of 125 µg l^–1, a possible antagonist of methylmercury. The contamination protocol was performed during 45 days and a 14 day period of recuperation was allowed. The stress caused by the transplantation of mussels in the laboratory tanks and/or by the presence of pollutants was evaluated by a general indicator of stress developed in our laboratory, the measure of the lysosomal membrane fragility (LMF) of the digestive gland, according to the method developed by Moore (1976). The effects of contamination on metabolism were measured by the study of the variations of the malate dehydrogenase activity (MDH), a key enzyme of the aerobic metabolism. The first days of the contamination period led to an increased metabolism in the mantle and to a detoxifying mechanism in the hepatopancreas. At days 22 and 29 of the experiment, the affinity of the MDH was greatly decreased with both concentrations of methylmercury and selenium, suggesting a competitive inhibition of the enzymatic activity by the pollutants. LMF increased as the mussels were kept longer in the tanks. Methylmercury increased the stress undergone by the mussels. LMF gives information about the degree of stress of the organism while the biochemical indicator informs about the metabolic effects of sublethal concentrations of pollutants.     

Cellular Expression, Developmental Regulation, and Phylogenic Conservation of PEA-15, the Astrocytic Major Phosphoprotein and Protein Kinase C Substrate

Abstract: PEA-15 has recently been identified as a major phosphoprotein in astrocytes and an endogenous substrate for protein kinase C. This 15-kDa protein exists under three molecular forms, an unphosphorylated form, N, and two phosphorylated forms, Pa and Pb. ntisera were raised against synthetic peptides corresponding to the internal sequences of the mouse protein containing the two specific phosphorylation sites and affinity-purified antibodies were used for immunoblotting. PEA-15 was found mainly in the cytosol, but its protein kinase C-phosphorylated form, Pb, was also detectable in association with the membrane and remained with the fraction that contains stabilized microtubules. Abundant in astrocytes, particularly in the hippocampus, PEA-15 was also detected in all cultured brain cell types examined, indicating a more ubiquitous distribution of the protein, further demonstrated by its detection in the eye and in the lung. Parallel to the increase in expression levels, phosphorylation of PEA-15 also increased during development. This paralleled results obtained in primary cultures, where PEA-15 levels increase with cell maturation. Finally, physiological importance of PEA-15 phosphorylation was illustrated by immunoreactivity observed in brain homogenates of different mammals, birds, amphibians, and fish.     

Increased REM sleep associated with melatonin deficiency after pinealectomy: a case study

The objectives of the investigation were to assess hypersomnia, which progressively appeared in a young patient after a pinealectomy, chemotherapy, and radiotherapy for a typical germinoma, as well as the potential benefit of melatonin administration in the absence of its endogenous secretion. 24 h ambulatory polysomnography and the Multiple Sleep Latency Test (MSLT) were performed; in addition, daily plasma melatonin, cortisol, growth hormone, prolactin, and rectal temperature profiles were determined before and during melatonin treatment (one 2 mg capsule given nightly at 21:00 h for 4 weeks). MSLT showed abnormal sleep latency and two REM sleep onsets. Nighttime total sleep duration was lengthened, mainly as a consequence of an increased REM sleep duration. These parameters were slightly modified by melatonin replacement. Plasma melatonin levels, which were constantly nil in the basal condition, were increased to supraphysiological values with melatonin treatment. The plasma cortisol profile showed nycthemeral variation within the normal range, and the growth hormone profile showed supplementary diurnal peaks. Melatonin treatment did not modify the secretion of either hormone. The plasma prolactin profile did not display a physiological nocturnal increase in the basal condition; however, it did during melatonin treatment, with the rise coinciding with the nocturnal peak of melatonin concentration. A 24 h temperature rhythm of normal amplitude was persistent, though the mean level was decreased and the rhythm was dampened during melatonin treatment. The role of radiotherapy on the studied parameters cannot be excluded; the findings of this case study suggest that the observed hypersomnia is not the result of melatonin deficiency alone. Overall, melatonin treatment was well tolerated, but the benefit on the sleep abnormality, especially on daytime REM sleep, was minor, requiring the re-introduction of modafinil treatment.     

Changes in mammary uptake and metabolic fate of glucose with once-daily milking and feed restriction in dairy cows

The aim of this review is to better understand the regulation of milk yield in response to once-daily milking and feed restriction. Glucose is the principal precursor for the synthesis of lactose (a major osmotic agent in milk), and participates in determining the milk volume produced. When applying these two breeding factors, reductions in milk yield are associated with a reduction in milk lactose yield and in the arterial flow of glucose, due to a decrease in the mammary blood flow. The ability of the udder to extract glucose is altered with once-daily milking but not necessarily with feed restriction. Lactose synthesis is down-regulated in response to once-daily milking and feed restriction but the percentage of the extracted glucose which is converted into lactose is differently affected in response to treatments. No marked change is observed with once daily milking whereas this would be increased with feed restriction and in contrast, depressed with fasting.