Effects of the Phosphatase Inhibitors Calyculin A and Okadaic Acid on Acetylcholine Synthesis and Content of Rat Hippocampal Formation

Abstract: The biochemical mechanisms involved in the regulation of acetylcholine (ACh) turnover are poorly understood. In the experiments reported here, we examined whether inhibition of the serine/threonine phosphatases 1 and 2A by calyculin A or okadaic acid alters ACh synthesis by rat hippocampal preparations. With hippocampal slices, calyculin A (50 n M ) and okadaic acid (50 n M ) reduced significantly ( p < 0.01) the synthesis of [³H]ACh from [³H]choline. Both calyculin A and okadaic acid produced significant depletion of endogenous tissue ACh in a concentration-dependent manner ( p < 0.01). This depletion was not the result of a drug-induced increase of spontaneous ACh release, which was not changed significantly ( p > 0.7) by either drug. Choline acetyltransferase (ChAT) activity from tissue exposed to calyculin A or okadaic acid was reduced in a concentration-dependent manner ( p < 0.05), but these phosphatase inhibitors did not act directly on ChAT in vitro; i.e., enzymatic activity was not altered significantly ( p > 0.4) in the presence of calyculin A or okadaic acid. Both high-affinity and low-affinity [³H]choline uptake by hippocampal synaptosomes were reduced significantly in a concentration-dependent manner in the presence of calyculin A or okadaic acid; these agents reduced V _max values for high- and low-affinity choline uptake ( p < 0.01) with no significant change in K _m values ( p > 0.1), indicating a noncompetitive inhibition. Taken together, these data suggest that phosphatase activity plays a role in presynaptic central cholinergic nerve terminal function, in particular in the modulation of ACh synthesis.     

Effect of sulphate on photosynthesis in greenhouse–grown tomato plants

Sulphate accumulates in the rhizosphere of plants grown in hydroponic systems. To avoid such sulphate accumulation and promote the use of environmentally sound hydroponic systems, we examined the effects of four sulphate concentrations (0.1, 5,2, 10.4 and 20.8 m M ) on photosynthesis, ribulose-l,5-bisphosphate carboxylase/oxygenase (Rubisco, EC 4.1.1.39) activities and related physiological processes in greenhouse–grown tomato plants ( Lycopersicon esculentum Mill. cv. Trust). The lowest sulphate concentration (0.1 m M ) significantly decreased photosynthetic capacity (P_c) and Rubisco activities on a leaf area basis. This result was supported by our data for dry matter per plant, which was low for plants in the 0.1 m M treatment. The photosynthesis-related variables such as leaf conductance, chlorophyll and soluble protein were lowest for the 0.1 m M treatment. Both total Rubisco activity and the activated ratio were reduced with this treatment. However, Rubisco activities expressed per g of protein or per g of chlorophyll were not significantly affected. These results suggest that sulphur deficiency depressed P_c– by reducing the amount of both Rubisco and chlorophyll and by causing an inactivation of Rubisco. The ratio of organic sulphur vs organic nitrogen (S/N) in plants of the 0.1 m M treatment was far below the normal values. This low S/N ratio might be accountable for the negative effect of low sulphate on P_c and plant growth. P_c and dry matter were not affected until sulphate concentration in the nutrient solution reached a high level of 20.8 m M .     

The loss of culturability by Escherichia coli cells in seawater depends on availability of phosphate ions and phosphate transport systems

Using strains with or without the PhoE porin or different components of the phosphate regulon, we determined that maintenance of the culturability of Escherichia coli in seawater depended significantly on the presence of structures allowing access of phosphate ions to the periplasm, then to the cytoplasm of cells. Cells totally deprived of the two main phosphate transport systems (Pit, Pst) exhibited the highest loss of culturability. Most of this effect resulted from the loss of the high-affinity Pst system, and more specifically that of the periplasmic phosphate-binding protein PhoS. Survival was enhanced in seawater supplemented with phosphate (0.5 mm), whether or not these structures were present. From an ecological point of view, it is assumed that the presence of phosphate ions, even at low concentrations, can influence the behavior of E. coli cells in seawater. Offprint requests to: M.J. Gauthier     

First chromosome scale genomes of ithomiine butterflies (Nymphalidae: Ithomiini): Comparative models for mimicry genetic studies

The ithomiine butterflies (Nymphalidae: Danainae) represent the largest known radiation of Müllerian mimetic butterflies. They dominate by number the mimetic butterfly communities, which include species such as the iconic neotropical Heliconius genus. Recent studies on the ecology and genetics of speciation in Ithomiini have suggested that sexual pheromones, colour pattern and perhaps hostplant could drive reproductive isolation. However, no reference genome was available for Ithomiini, which has hindered further exploration on the genetic architecture of these candidate traits, and more generally on the genomic patterns of divergence. Here, we generated high-quality, chromosome-scale genome assemblies for two Melinaea species, M. marsaeus and M. menophilus, and a draft genome of the species Ithomia salapia. We obtained genomes with a size ranging from 396 to 503 Mb across the three species and scaffold N50 of 40.5 and 23.2 Mb for the two chromosome-scale assemblies. Using collinearity analyses we identified massive rearrangements between the two closely related Melinaea species. An annotation of transposable elements and gene content was performed, as well as a specialist annotation to target chemosensory genes, which is crucial for host plant detection and mate recognition in mimetic species. A comparative genomic approach revealed independent gene expansions in ithomiines and particularly in gustatory receptor genes. These first three genomes of ithomiine mimetic butterflies constitute a valuable addition and a welcome comparison to existing biological models such as Heliconius, and will enable further understanding of the mechanisms of adaptation in butterflies.     

Phylogenomic study of Amorphophallus (Alismatales; Araceae): When plastid DNA gene sequences help to resolve the backbone subgeneric delineation

Encompassing ca. 200 species distributed in paleotropical Africa and Asia, Amorphophallus is one of the largest genera of Araceae. In spite of the great economic interest in its glucomannan production, only a few studies have attempted to grasp the evolutionary history of this genus. In the current state of knowledge, four main clades, mostly linked to biogeographical delineation, have been identified from phylogenies based on a few genes. However, relationships among and within these clades still remain unclear, due to the rapid radiation that occurred during the early evolutionary history of the genus. Here, we generated genome skimming libraries for 43 specimens from 36 species distributed across the 4 clades, which allowed us to produce a phylogenetic matrix for a set of 71 plastid genes. Our phylogenies confirm the monophyly of these clades but show a new and well-resolved arrangement among these clades. Our analyses therefore provide a new scenario and timeline for the evolution of the main Amorphophallus clades, consistent with the morphological characteristics of the clades. The inferred scenario is also in agreement with climate dynamics and the onset of long-distance dispersal by the earliest migratory birds near the Oligocene/Miocene transition around 23 million years ago. Our study provides an up-to-date baseline to understand biogeographic and ecological processes that shaped the current diversity and distribution of Amorphophallus, paving the way for larger-scale phylogenomic studies based on plastid and nuclear genomes.     

Dominance competition through affiliation and support in Japanese macaques: An experimental study

It has been proposed that monkeys direct grooming to high-ranking individuals in an attempt to obtain agonistic support in return. But whether these two categories of interactions are causally related has proven difficult to establish. Part of the problem stems from the fact that in stable groups social relationships reflect an equilibrium state and that behaviors need only be performed at low rates and long intervals to maintain the current social structure. In theory, however, if affiliative and supportive interactions are indeed causally related, it should be possible to accentuate their temporal relation, hence their causal dynamics. For example, destabilizing dominance relations can be expected to induce competition for status and force individuals to deploy behavioral tactics for settling new rank relations. We experimentally induced rank reversals in a captive group of Japanese macaques (Macaca fuscata) composed of three matrilines (A-B-C rank order). A reversed C-A-B order composed of three individuals per matriline was maintained for 2 weeks. The results show the close temporal relation among (i) asserting one’s rank, (ii) competing for access to dominants through affiliation and interferences in affiliation, (iii) receiving support from dominants against lower-ranking individuals, and (iv) supporting dominants against subordinates. These findings are compatible with one version of the affiliation-for-support hypothesis, namely that monkeys affiliate with dominants as a way to assert their position in the hierarchy. In a functional perspective, mutual selfishness provides a better explanation than reciprocal altruism because the possibility that both groomers and supporters derive immediate net benefits cannot be excluded.     

The palatability of Arctic willow for greater snow geese: the role of nutrients and deterring factors

Temporary feeding on willow buds and leaves by nesting greater snow geese provided us with an opportunity to test the relative importance of nutrients and deterrents in affecting the palatability for geese of a food plant with a high phenol content. Protein, total phenol and fiber (neutral and acid detergent fiber, and lignin) were analyzed in closed and open buds and in rolled and open leaves. Geese feed on willows at the open-buds and rolled-leaf stages but not at the closed-bud and open-leaf stages. Protein content was higher in open buds and rolled leaves (25–27%) than in closed buds and open leaves (19–21%). Phenol content increased during leaf emergence but was already high (14%) in rolled leaves. All plant fibers were very high in closed buds but declined rapidly during leaf emergence. The increase in phenol: protein ratio appeared to be more important than phenol concentration alone in explaining the cessation of feeding by geese on willow leaves whereas the high fiber content of closed buds may explain why they were not eaten. Our results illustrate the value of a multifactorial approach in the study of the food selection process in herbivores.     

Marinobacter hydrocarbonoclasticus gen. nov., sp. nov., a new, extremely halotolerant, hydrocarbon-degrading marine bacterium.

On the basis of phenotypical characteristics and analysis of 16S rRNA sequence, a new species belonging to a new genus is described, and the name Marinobacter hydrocarbonoclasticus is proposed. This organism, isolated from Mediterranean seawater near a petroleum refinery, is a gram-negative, aerobic, rod-shaped bacterium. It grows at NaCl concentrations of 0.08 to 3.5 M and uses various hydrocarbons as the sole source of carbon and energy. Its DNA has a guanine-plus-cytosine content of 52.7 mol%. The 16S rRNA analysis shows a clear affiliation between M. hydrocarbonoclasticus and the gamma group of the phylum Proteobacteria. A close phylogenetic relationship appears among the species Marinomonas vaga, Oceanospirillum linum, Halomonas elongata, and Pseudomonas aeruginosa. Because of the impossibility of finding a single most closely related species, we suggest that this bacterium be assigned to a new genus, at least temporarily. The possibility of a revision of this status when new data appear is, however, not excluded. The type strain is M. hydrocarbonoclasticus SP.17 (= ATCC 49840).     

Inhibition of rat tissue kallikrein gene family members by rat kallikrein-binding protein and alpha 1-proteinase inhibitor.

The regulation of tissue kallikrein activity by plasma serine proteinase inhibitors (serpins) was investigated by measuring the association rate constants of six tissue-kallikrein family members isolated from the rat submandibular gland, with rat kallikrein-binding protein (rKBP) and alpha 1-proteinase inhibitor (alpha 1-PI). Both these serpins inhibited kallikreins rK2, rK7, rK8, rK9 and rK10 with association rate constants in the 10(3)-10(4) M-1.s-1 range, whereas only 'true' tissue kallikrein rK1 was not susceptible to alpha 1-PI. This results in slow inhibition of rK1 by plasma serpins, which could explain why this kallikrein is the only member of the gene family identified so far that induces a transient decrease in blood pressure when injected in minute amounts into the circulation.