Ca2+,Mg2+-ATPase of microsomal membranes from bovine aortic smooth muscle. Identification and characterization of an acid-stable phosphorylated intermediate of the Ca2+,Mg2+-ATPase

An acid-stable phosphoprotein was formed in a microsomal membrane fraction isolated from bovine aortic smooth muscle in the presence of Mg2+ + ATP and Ca2+. The microsomes also showed Ca2+ uptake activity. The Ca2+ dependence of phosphoprotein formation and of Ca2+ uptake occurred over the same range of Ca2+ concentration (1-10 microM), and resembled similar findings from rabbit skeletal microsomes. The molecular weight of the phosphorylated protein, estimated by SDS-gel electrophoresis, was approximately 105,000. The phosphoprotein was labile at alkaline pH, and its decomposition was accelerated by hydroxylamine. Half-maximum incorporation of 32P in the presence of 10 microM Ca2+ occurred at 60 nM ATP. The calcium-dependent phosphoprotein formation was not affected by 5 mM NaN3, but was inhibited in a dose-dependent fashion by ADP with a 50% inhibition occurring at 180 microM. Fifty mM MgCl2 was required for the maximal phosphorylation. The rate of phosphoprotein decomposition after adding 2 mM EGTA was accelerated by varying the Mg2+ concentration from 10 microM to 3 mM. Alkaline pH (9.0) slowed the rate of phosphoprotein decay. Optimal Ca2+-dependent phosphoprotein occurred at 15 degrees C over a broad pH range (6.4 to 9.0). The activation energy of EGTA-induced phosphoprotein decomposition was 25.6 kcal/mol between 0 and 16 degrees C and 14.6 kcal/mol between 16 and 30 degrees C. The phosphoprotein formed by aortic microsomes was thus quite similar to the acid-stable phosphorylated intermediate of the Ca2+-transport ATPase of sarcoplasmic reticulum from skeletal and cardiac muscle. These data suggest that the Ca2+-dependent phosphoprotein is a reaction intermediate of the Ca2+,Mg2+-ATPase of the aortic microsomes.     

Zymographic demonstration of lactate and malate dehydrogenases isoenzymes in the rodent salivary glands

Summary Analysis of lactate and malate dehydrogenase zymograms of rodent salivary glands showed species and organ specific patterns.Lactate dehydrogenase isoenzyme patterns occupied the middle positions in relation to those of skeletal and heart muscle. Activities of the major salivary glands were in the order submaxillary gland>parotid>sublingual gland. Zymogram of the mouse and rat showed LDH₄ and LDH₅ high activity patterns, while that of the rabbit was the fast moving active one. Hamster salivary gland exhibited a neutral type of the former and the latter.Malate dehydrogenase isoenzyme exhibited very similar patterns for the mouse, rat and hamster. Malate dehydrogenase zymogram of rabbit showed 3 active bands, which was different from the other rodents.     

Influence of overwinteringDaphnia on spring zooplankton communities: An experimental study

To evaluate the influence of overwintering individuals of zooplankton on spring zooplankton communities, the dynamics of zooplankton communities with or without overwintering individuals were observed in experimental ponds from fall to spring. An insecticide, carbaryl, was used to regulate the overwintering individuals. In ponds which received insecticide applications in November or January, all cladoceran and rotiferan species were eliminated by the treatments and did not reappear until late March or early April, even when the chemical disappeared rapidly. The low water temperature may delayed the establishment of the populations from resting eggs. In these ponds, populations of various cladoceran and rotiferan species, which seemed to be originated from resting eggs, were built up in the spring. In control ponds,Daphnia ambigua orD. longispina overwintered as juveniles and adults and established a large spring population earlier than other cladocerans and rotifers overwintering as resting eggs. The latter zooplankters did not increase in the spring probably because their growth was suppressed by the precedingDaphnia species through competition. In nature, even if the number of overwintering individuals is small, they may have a potential to build up a large population earlier than the individuals hatching from resting eggs. As a result, the species which have overwintered as individuals seem to predominate in the spring and have a large influence on the spring zooplankton community.     

Production ecology of phyto- and zooplankton in a eutrophic pond dominated byChaoborus flavicans (Diptera: Chaobolidae)

Primary production of phytoplankton and secondary production of a daphnid and a chaoborid were studied in a small eutrophic pond. The gross primary production of phytoplankton was 290 gC m⁻² per 9 months during April–December. Regression analysis showed that the gross primary production was related to the incident solar radiation and the chlorophylla concentration and not to either total phosphorus or total inorganic nitrogen concentration. The mean chlorophylla concentration (14.2 mg m⁻³), however, was about half the expected value upon phosphorus loading of this pond. The mean zooplankton biomass was 1.60 g dry weight m⁻², of whichDaphnia rosea and cyclopoid copepods amounted to 0.69 g dry weight m⁻² and 0.61 g dry weight m⁻², respectively. The production ofD. rosea was high during May–July and October and the level for the whole 9 months was 22.6 g dry weight m⁻².Chaoborus flavicans produced 10 complete and one incomplete cohorts per year. Two consecutive cohorts overlapped during the growing season. The maximum density, the mean biomass, and the production were 19,100 m⁻², 0.81 g dry weight m⁻², and 11.7 g dry weight m⁻²yr⁻¹, respectively. As no fish was present in this pond, the emerging biomass amounted to 69% of larval production. The production ofC. flavicans larvae was high in comparison with zooplankton production during August–September, when the larvae possibly fed not only on zooplankton but also algae.     

Immunogold Localization of Ribulose-1,5-Bisphosphate Carboxylase with Reference to Pyrenoid Morphology in Chloroplasts of Synchronized Euglena gracilis Cells

Euglena gracilis strain (Z) cells were synchronized under photoautotrophic conditions using a 14 hour light:10 hour dark regimen. The cells grew during the light period (growth phase) and divided during the following 10 hour period either in the dark or in the light (division phase). Changes in morphology of the pyrenoid and in the distribution of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) within the chloroplasts were followed by immunoelectron microscopy during the growth and division phases of Euglena cells. Epon-embedded sections were labeled with an antibody to the holoenzyme followed by protein A-gold. The immunoreactive proteins were concentrated in the pyrenoid, and less densely distributed in the stroma during the growth phase. During the division phase, the pyrenoid could not be detected and the gold particles were dispersed throughout the stroma. Toward the end of the division phase, the pyrenoid began to form in the center of a chloroplast, and the immunoreactive proteins started to concentrate over that rudimentary pyrenoid. During the growth phase, small areas rich in gold particles, called `satellite pyrenoid,' were observed, in addition to the main pyrenoid. From a comparison of photosynthetic CO₂-fixation with the total carboxylase activity of Rubisco extracted from Euglena cells in the growth phase, it is suggested that the carboxylase in the pyrenoid functions in CO₂-fixation in photosynthesis.     

The Thermostable Direct Hemolysin Gene (tdh) of Vibrio hollisae Is Dissimilar in Prevalence to and Phylogenetically Distant from the tdh Genes of Other Vibrios: Implications in the Horizontal Transfer of the tdh Gene

Vibrio hollisae strains isolated recently from patients in various locations were examined for the presence of the thermostable direct hemolysin gene (tdh) using nucleic acid hybridization and polymerase chain reaction assays. The results were consistent with the previous finding that all strains of V. hollisae carry the tdh gene. In contrast, the tdh gene has been detected in a minority of strains for other Vibrio species (V. parahaemolyticus, V. cholerae non-O1, and V. mimicus). Detailed phylogenetic analysis showed that the tdh genes of the non-V. hollisae species were very closely related to each other and that the tdh gene of V. hollisae was distantly related to the tdh genes of the non-V. hollisae species. These results and the proposed insertion sequence-mediated tdh transfer mechanism suggest that the tdh gene may have been maintained stably in V. hollisae and that the tdh genes of the non-V. hollisae species may have been involved in recent horizontal transfer.     

Suppression of gravitropic response of primary roots by submergence

Primary roots of six plant species were placed horizontally either in humid air or under water, and their growth and gravitropic responses were examined. In air, all the roots showed a normal gravitropic curvature. Under water without aeration, roots of rice (Oryza sativa L.), oat (Avena sativa L.), azuki bean (Vigna angularis Ohwi et Ohashi), and cress (Lepidium sativum L.) curved downward at almost same rate as in air, whereas the curvature of roots of maize (Zea mays L.) and pea (Pisum sativum L.) was strongly suppressed. Submergence did not cause a decrease in growth rate of these roots. When roots of maize and pea were placed horizontally under water without aeration and then rotated in three dimensions on a clinostat in air, they showed a significant curvature, suggesting that the step suppressed by submergence is not graviperception but the subsequent signal transmission or differential growth process. Constant bubbling of air through the water partly restored the gravitropic curvature of maize roots and completely restored that of pea roots. The curvature of pea roots was also partly restored by the addition of an inhibitor of ethylene biosynthesis, aminooxyacetic acid. In air, ethylene suppressed the gravitropic curvature of roots of maize and pea. Furthermore, the level of ethylene in the intercellular space of the roots was increased by submergence. These results suggest that the accumulation of ethylene in the tissue is at least partly involved in suppression of transmission of the gravity signal or of differential growth in maize and pea roots under conditions of submergence.Abbreviations AOA aminooxyacetic acid - 3-D three-dimensional Dedicated to Professor Andreas Sievers on the occasion of his retirementWe thank Professor H. Suge and Drs. H. Takahashi and H. Kataoka, Tohoku University and Dr. T. Suzuki, Yamagata University, for helpful suggestions. The present study was supported in part by a Grant for Basic Research in Space Station Utilization from the Institute of Space and Astronautical Science, Japan.     

Efficiency of serum copper/zinc ratio for differential diagnosis of patients with and without lung cancer

We examined serum copper (Cu), serum zinc (Zn), and the serum copper/zinc ratio (Cu/Zn) in 162 patients. All of them were seen to have an abnormal shadow in the chest X-ray films, that is, 109 patients with lung cancer (LC) and 53 patients with no lung cancer (NLC). The mean Cu and Cu/Zn in LC patients were significantly higher than those in NLC patients (p<0.05). In LC patients, Cu and Cu/Zn were higher and Zn was lower in advanced tumors than early ones. There was a significantly clear relation between Cu or Cu/Zn and the tumor (T) stages. When the relative risk (RR) of LC was estimated, it was seen that the higher Cu and Cu/Zn became, the higher RR became. Furthermore, we showed the sensitivity of the receiver operator characteristic of the test (ROC) curve for Cu, Cu/Zn, and carcinoembryonic antigen (CEA) to diagnose LC, as explained in a paragraph of methods.The determinations of Cu, Zn, and Cu/Zn are simple and inexpensive. They also appear to have a great diagnostic value in determining the local invasion of LC and as a screening test in the high-risk patients for LC.     

Detection and Typing of Human Rotavirus in Reference to Repeated Acute Gastroenteritis in Infants

Stool specimens from infants who visited a clinic because of acute gastroenteritis were tested for the presence of human rotavirus. Among the samples obtained were specimens taken from seven patients who had visited the clinic at two different times. In six of these seven children, human rotavirus (HRV) was detected in only one of the specimens taken (i.e. during only one of the two visits). One patient was shown to have excreted HRV twice; in both cases the HRV was serotyped to be type 1. The present results indicate that the symptomatic reinfection of HRV was not a widely occurring phenomenon in the group of infants tested.