The gene for trypsin inhibitor CMe is regulated in trans by the lys 3a locus in the endosperm of barley (Hordeum vulgare L.)

Summary A cDNA encoding trypsin inhibitor CMe from barley endosperm has been cloned and characterized. The longest open reading frame of the cloned cDNA codes for a typical signal peptide of 24 residues followed by a sequence which is identical to the known amino acid sequence of the inhibitor, except for an Ile/Leu substitution at position 59. Southern blot analysis of wheat-barley addition lines has shown that chromosome 3H of barley carries the gene for CMe. This protein is present at less than 2%–3% of the wild-type amount in the mature endosperm of the mutant Risø 1508 with respect to Bomi barley, from which it has been derived, and the corresponding steady state levels of the CMe mRNA are about I%. One or two copies of the CMe gene (synonym Itc1) per haploid genome have been estimated both in the wild type and in the mutant, and DNA restriction patterns are identical in both stocks, so neither a change in copy number nor a major rearrangement of the structural gene account for the markedly decreased expression. The mutation at the lys 3a locus in Risø 1508 has been previously mapped in chromosome 7 (synonym 5H). A single dose of the wild-type allele at this locus (Lys 3a) restores the expression of gene CMe (allele CMe-1) in chromosome 3H to normal levels.     

Immunocytochemical and ultrastructural study of the frog (Rana pipiens) pars distalis with special reference to folliculo-stellate cell function during in vitro superfusion

Summary The colloidal gold immunocytochemical technique was used to determine the ultrastructural features of the glandular cells in the pituitaries of male frogs, Rana pipiens, both in vivo and after superfusion in vitro. Specific reactions to antisera against bullfrog gonadotropins, human prolactin, and synthetic 1–39 corticotropin allowed identification of the 3 corresponding types of glandular cells. No immunoreaction was obtained with antisera against human or ovine-growth hormone, human -thyrotropin hormone, and bovine S-100 protein. General morphological features of these immunocytochemically identified glandular cells were similar to those of equivalent cells previously described in other amphibian species. Non-glandular folliculo-stellate cells were distinctive. In freshly removed pituitaries, these folliculo-stellate cells contained lysosome-like structures, but did not show phagocytic vacuoles in the cytoplasm; they contained many mitochondria, and the Golgi complex and endoplasmic reticulum were relatively undeveloped. After 4 or 18 h of superfusion, some immunoreactive gonadotropic, prolactin, and corticotropic cells showed degeneration and destruction. In the same gland, folliculo-stellate cells retained a viable appearance, but showed phagocytic vacuoles containing secretory granule-like structures which were immunoreactive to gonadotropic, prolactin, and corticotropic antibodies. Some folliculo-stellate cells showed phagocytic vacuoles containing complete glandular cells. These results suggest that superfusion causes a destruction of some of the glandular cells, and that folliculo-stellate cells act as phagocytes when cellular debris or moribund cells are present in the intercellular space in the pituitary parenchyma.Supported by grant DCB 8710462 from the National Science Foundation, grant 2148-83 from the CAICYT (Spain) and the Junta de Andalucia (Spain)     

Regulation of reduced nitrogen assimilation in Rhodobacter capsulatus E1F1

The phototrophic bacterium Rhodobacter capsulatus E1F1 assimilates ammonia and other forms of reduced nitrogen either through the GS/GOGAT pathway or by the concerted action of l-alanine dehydrogenase and aminotransferases. These routes are light-independent and very responsive to the carbon and nitrogen sources used for cell growth. GS was most active in cells grown on nitrate or l-glutamate as nitrogen sources, whereas it was heavily adenylylated and siginificantly repressed by ammonium, glycine, l-alanine, l-aspartate, l-asparagine and l-glutamine, under which conditions specific aminotransferases were induced. GOGAT activity was kept at constitutive levels in cells grown on l-amino acids as nitrogen sources except on l-glutamine where it was significantly induced during the early phase of growth. In vitro, GOGAT activity was strongly inhibited by l-tyrosine and NADPH. In cells using l-asparagine or l-aspartate as nitrogen source, a concerted induction of l-aspartate aminotransferase and l-asparaginase was observed. Enzyme level enhancements in response to nitrogen source variation involved de novo protein synthesis and strongly correlated with the cell growth phase.Abbreviations ADH l-alanine dehydrogenase - AOAT l-alanine:2-oxoglutarate aminotransferase - Asnase l-asparaginase - GOAT Glycine: oxaloacetate aminotransferase - GOGAT Glutamate synthase - GOT l-aspartate: 2-oxoglutarate aminotransferase - GS Glutamine synthetase - HPLC High-Pressure Liquid Chromatography - MOPS 2-(N-morpholino)propanesulfonic acid - MSX l-methionine-d,l-sulfoximine     

A similar protein portion for two exoglucanases secreted by Saccharomyces cerevisiae

Exoglucanase (exo-1,3-β-D-glucan glycohydrolase, EC 3.2.1.56) activity secreted by Saccharomyces cerevisiae into the culture medium was separated by ion exchange chromatography into two glycoprotein isoenzymes which contributed 10% (exoglucanase I) and 90% (exoglucanase II) towards the total activity. Analysis of the “in vitro” deglycosylated products by polyacrylamide gel electrophoresis under native or denaturing conditions indicated that the protein portions of both exoglucanases exhibited identical mobility, each one consisting of two polypeptides with M _r of 47000 and 48000. The same profile was shown by the exoglucanase secreted in the presence of tunicamycin. Antibodies raised against the protein portion of exoglucanase II did react with both native exoglucanases and their deglycosylated products with a pattern indicative of immunological identity. Digestion of the “in vitro” deglycosylated products of both exoglucanases with Staphylococcus aureus V-8 protease or trypsin generated the same proteolytic fragments in each case. Only exoglucanase II was secreted by protoplasts. These and previously reported results indicate that the protein portions of both isoenzymes may be the product of the same gene (or a family of related genes), and that exoglucanase I is a product of enzyme II, modified by a process occurring beyond the permeability barrier of the cell.     

Size variation in Brachionus plicatilis resting eggs

The effect of temperature and salinity on resting egg size of two Brachionus plicatilis (Rotifers) clones was investigated. Clones were selected according to their different behaviour in laying resting eggs: one clone ejects them, whereas they remain inside the females body in the other clone. The difference in resting eggs size between the two clones is noticeable, although the difference is not as great as that between female body size. An important temperature-salinity interaction on resting egg size has been observed. The general inverse relationship between size and temperature is only true at lower temperatures. At high temperatures size varies around the mean although could be greater than at intermediate temperatures. This is more evident at the intermediate salinity tested which is considered to be the closest to the optimum in our experiments. This pattern of variation suggests that mean size is bigger than expected, in relation to temperature and salinity, when these factors have values close to the extremes of their range, normally found in nature, and to which adaptative mechanisms can evolve. Size is bigger at the salinity — temperature low - low and high - high combinations which are the most commonly found in the temperate environments.     

Evidence from algal bioassays of seasonal nutrient limitations in two English lakes

Comparative laboratory bioassays using Asterionella formosa and Rhodomonas lacustris as test organisms were performed from March to November 1987 on filtered water samples from two English lakes, to assess their potential fertility and to identify possible limiting nutrients. The relative growth responses (log₂ increments) per week, were measured after additions of P, Fe, Si, N, and K singly and in combinations in comparison with unenriched (control) samples. Phosphate appeared to be the major limiting element for both species throughout the year, except during the spring diatom maxima when silicon usually becomes limiting. On most occasions chelated iron increased the growth increments, particularly in combination with phosphate. In general, the bioassay results showed correspondence with the nutrient concentrations in the test waters, which showed low (< 1 µg l^–1) levels of soluble reactive phosphate during all or most of the year and depleted silicon levels in late spring. Comparison between relative (incremental) ratio and absolute (cell concentration) response was made.     

Dicarboxylic acid anhydrides as dissociating agents of protein-containing structures

Dissociation of protein-containing structures by modification of protein amino groups with dicarboxylic acid anhydrides is a mild procedure which, in some cases, offers advantages over treatment with alternative dissociating agents, such as urea, guanidine hydrochloride, detergents, high ionic strength, and extremes of pH: In addition to dissociating multimeric proteins and protein aggregates, dicarboxylic acid anhydrides are effective dissociating agents for membrane-bound proteins and nucleoprotein particles. With most dicarboxylic acid anhydrides reviewed, the introduced reagent residues can be eliminated under moderate acid conditions, which allows the purification of unmodified individual components, and the use of disassembly-reconstitution systems valuable for investigating the structural and functional roles played by the individual components of complex particles:Each reagent can be suitable for a particular purpose, depending on the required specificity of the modification and stability of the modified groups: The stability of the acylated amino groups ranges from the very stable succinylated amino groups to the very labile acylation obtained with dimethylmaleic anhydride: Between these extremes, the stability of the modified amino groups decreases stepwise in the following order: maleic, exo-cis-3,6-endoxo-⁴-tetrahydrophthalic, citraconic, and 3,4,5,6-tetrahydrophthalic anhydride. With respect to the selectivity of the produced modification, little or no modification of hydroxyamino acid and cysteine residues has been observed with dimethylmaleic, exo-cis-3,6-endoxo-⁴-tetrahydrophthalic, and 3,4,5,6-tetrahydrophthalic anhydrides: With the other reagents, the extent of modification of hydroxyamino acid residues increases in the order citraconic, maleic and succinic anhydride: Citraconic and maleic anhydrides can produce irreversible modification of cysteine residues, the reactivity of sulfhydryl groups being higher with maleic anhydride:     

Stereological study of gonadotropes in the frog,Rana pipiens,after GnRH stimulation in vitro

Summary Previous physiological results have indicated the existence of two releasable pools of gonadotropins in amphibian pituitaries: an acute releasable pool that appears independent of protein synthesis, and a storage pool involved in chronic release that depends on protein synthesis. To elucidate the ultrastructural localization of these pools and the morphological changes induced in gonadotrope cells after treatment with gonadotropin-releasing hormone, we carried out a morphometric study of immuno-identified gonadotrope cells using an in vitro superfusion system. Treatment with gonadotropin-releasing hormone induced a degranulation of small (110–255 nm) and medium (236–360 nm) secretory granules as well as hypertrophy of the endoplasmic reticulum and Golgi complex. Simultaneous incubation with gonadotropin-releasing hormone and cycloheximide inhibited the release of secretory granules although the endoplasmic reticulum and Golgi complex were hypertrophied. These morphological results strongly suggest: (1) that gonadotropin-releasing hormone induces degranulation and hypertrophy of the biosynthetic machinery in gonadotrope cells; and (2) that the activation of the endoplasmic reticulum and Golgi complex by stimulation with gonadotropin-releasing hormone is independent of protein synthesis, while the release of secretory granules is protein synthesis-dependent. In addition, the second or storage pool of gonadotropin is associated mainly with the small and medium secretory granules.     

Underwater observations of piranhas in western Brazil

Synopsis The behaviour of three piranha species,Serrasalmus marginatus, S. spilopleura, andPygocentrus nattereri, and their prey fishes was studied underwater in the Pantanal region, Mato Grosso, Brazil. General habits, predatory tactics, feeding behaviour, and social interactions while foraging, as well as defensive tactics of prey fishes were observed.S. marginatus is solitary whereas the other two species live in shoals; their agonistic behaviour varies accordingly, the simplest being displayed by the solitary species. Predatory tactics and feeding behaviour also vary:S. spilopleura shows the most varied diet and highly opportunistic feeding strategy, which includes aggressive mimicry. The solitaryS. marginatus, besides fin and scale-eating, occasionally cleans larger individuals ofP. nattereri. Several cichlid species display defensive tactics clearly related to piranha attacks: tail protecting, watching, and confronting the predator are the most commonly observed behaviours. Piranhas seem to strongly influence use of habitat, social structure, and foraging mode of the fish communities.