The function of selenocysteine synthase and SELB in the synthesis and incorporation of selenocysteine.

The selAB operon codes for the proteins selenocysteine synthase and SELB which catalyse the synthesis and cotranslational insertion of selenocysteine into protein. This communication deals with the biochemical characterisation of these proteins and in particular with their specific interaction with the selenocysteine-incorporating tRNA(Sec). Selenocysteine synthase catalyses the synthesis of selenocysteyl-tRNA(Sec) from seryl-tRNA(Sec) in a pyridoxal phosphate-dependent reaction mechanism. The enzyme specifically recognizes the tRNA(Sec) molecule; a cooperative interaction between the tRNA binding site and the catalytically active pyridoxal phosphate site is suggested. SELB is an EF-Tu-like protein which specifically complexes selenocysteyl-tRNA(Sec). Interaction with the selenol group of the side chain of the aminoacylated residue is a prerequisite for the formation of a stable SELB.tRNA complex. Mechanistically, this provides the biochemical basis for the exclusive selection of selenocysteyl-tRNA(Sec) in the decoding step of a selenocysteine-specific UGA triplet.     

Biphasic changes in leukocytes induced by strenuous exercise

Seven healthy male volunteers participated in short- (STR, 1.7 km), middle- (MTR, 4.8 km) and long- (LTR, 10.5 km) term runs at a speed close to their maximum. A prompt mobilization of white cells, and lymphocytes in particular, appeared following the exercise. The initial increase in the number of lymphocytes was succeeded by a significant decrease [(P less than 0.03) lymphopenial], which on average was 32%-39% of the pre-exercise values in all groups. A close correlation was found between the initial increase in plasma cortisol concentration after exercise and the subsequent lymphopenia. A modest enhancement in the number of granulocytes immediately after the exercise was accompanied by a comprehensive increase in polymorphonuclear (PMN) elastase concentration accounting for 78.6%, SEM 16.3%, 140.7%, SEM 31.8% and 241.3%, SEM 48.1% in the STR, MTR and LTR groups. No correlation was found between granulocyte number and the plasma PMN elastase concentration. A delayed granulocytosis was noted in all subjects, reaching a peak between 2 and 4 h after the exercise. The magnitude of the granulocytosis varied among subjects and peak values of the number of circulating granulocytes were found to be 5.7 x 10(9) cells.l-1, SEM 0.5, 6.7 x 10(9) cells.l-1, SEM 0.6 and 8.8 x 10(9) cells.l-1, SEM 0.5 in STR, MTR and LTR respectively, whereas the mean baseline value was 3.6 x 10(9) cells.l-1, SEM 0.4. The neutrophilic granulocytosis was not accompanied by a corresponding enhancement in PMN elastase concentration. The plasma cortisol concentration reached a peak 30 min after exercise and declined below the control level in 4 h. Neither the initial increase, nor the subsequent decrease in plasma cortisol concentration were found to be essential for the magnitude of the delayed leukocytosis.     

Physiological effects of micropauses in isometric handgrip exercise

The physiological response to continuous and intermittent handgrip exercise was evaluated. Three experiments were performed until exhaustion at 25% of maximal voluntary contraction (MVC): experiment 1, continuous handgrip (CH) (n = 8); experiment 2, intermittent handgrip with 10-s rest pause every 3 min (IH) (n = 8); and experiment 3, as IH but with electrical stimulation (ES) of the forearm extensors in the pauses (IHES) (n = 4). Before, during, and after exercise, recordings were made of heart rate (HR), arterial blood pressure (BP), exercising forearm blood flow, and concentrations of potassium [K+] and lactate [La-] in venous blood from both arms. The electromyogram (EMG) of the exercising forearm extensors and perceived exertion were monitored during exercise. Before and up to 24 h after exercise, observations were made of MVC, of force response to electrical stimulation and of the EMG response to a 10-s test contraction (handgrip) at 25% of the initial MVC. Maximal endurance time (tlim) was significantly longer in IH (23.1 min) than in CH (16.2 min). The ES had no significant effect on tlim. During exercise, no significant differences were seen between CH and IH in blood flow, venous [K+] and [La-], or EMG response. The HR and BP increased at the same rate in CH and IH but, because of the longer duration of IH, the levels at exhaustion were higher in this protocol. The subjects reported less subjective fatigue in IH. During recovery, return to normal MVC was slower after CH (24 h) than after IH (4 h).(ABSTRACT TRUNCATED AT 250 WORDS)     

Enzyme activities in Chrysosporium strains

390 strains of Chrysosporium were screened for their ability to produce enzymes. All strains produced: catalase, phosphatase, lipase, amylase, DNAse and phosphoamidase. No strains showed: valine arylamidase, oxidase, -galactosidase, urease, pectolase, protease nor RNAse.     

Training effects of cross-country skiing and running on maximal aerobic cycle performance and on blood lipids

Two experiments were carried out to compare the cardiorespiratory and metabolic effects of cross-country skiing and running training during two successive winters. Forty-year-old men were randomly assigned into skiing (n = 15 in study 1, n = 16 in study 2), running (n = 16 in study 1 and n = 16 in study 2) and control (n = 17 in study 1 and n = 16 in study 2) groups. Three subjects dropped out of the programme. The training lasted 9-10 weeks with 40-min exercise sessions three times each week. The training intensity was controlled at 75%-85% of the maximal oxygen consumption (VO2max) using portable heart rate metres and the mean heart rate was 156-157 beats.min-1 in the training groups. In the pooled data of the two studies the mean increase in the VO2max (in ml.min-1.kg-1) on a cycle ergometer was 17% for the skiing group, 13% for the running group and 2% for the control group. The increase in VO2max was highly significant in the combined exercise group compared to the control group but did not differ significantly between the skiing and running groups. The fasting serum concentrations of lipoproteins and insulin did not change significantly in any of the groups. These results suggested that training by cross-country skiing and running of the same duration and intensity at each session for 9-10 weeks improved equally the cardiorespiratory fitness of untrained middle-aged men.     

Calcitonin gene-related peptide and its mRNA in pulmonary neuroendocrine cells and ganglia.

The occurrence of calcitonin gene-related peptide (CGRP) and it's mRNA was studied in lungs of rats and piglets using in situ hybridization with two synthetic oligonucleotide probes followed by immunocytochemistry (ICC). CGRP mRNA was present in pulmonary neuroendocrine cells (PNEC) of both the solitary type and cluster type (neuroepithelial body; NEB) at all levels of the airway epithelium from bronchi to alveoli. The distribution of labelled cells was similar to that previously described with ICC. The 44-mer probe provided stronger hybridization signal than the 34-mer and the two combined increased labelling slightly. Formalin fixation reduced labelling and tended to increase background. Labelling for CGRP mRNA was evenly distributed over the cytoplasm, whereas CGRP-like immunoreactivity (LI) usually was of highest intensity toward the base of the PNEC, suggesting basal accumulation of synthesized peptide. CGRP-LI was also observed in occasional rat ganglia and in some, but not all, piglet ganglia. These local neurons may contribute to the CGRP fibers of airways and vasculature, and could theoretically bridge their dendrites and axons between NEB and the effector organ (e.g. artery or arteriole) thus accomplishing a function similar to the postulated axon reflex.     

The effect of different blood sampling sites and analyses on the relationship~ between exercise intensity and 4.0 mmol ·1? blood lactate concentration

The aim of the study was to examine whether the difference in lactate concentration in different blood fractions is of practical importance when using blood lactate as a test variable of aerobic endurance capacity. Ten male firefighters performed submaximally graded exercise on a cycle ergometer for 20-25 min. Venous and capillary blood samples were taken every 5 min for determination of haematocrit and lactate concentrations in plasma, venous and capillary blood. At the same time, expired air was collected in Douglas bags for determination of the oxygen consumption. A lactate concentration of 4.0 mmol.l-1 was used as the reference value to compare the oxygen consumption and exercise intensity when different types of blood specimen and sampling sites were used for lactate analysis. At this concentration the exercise intensity was 17% lower (P less than 0.01) when plasma lactate was compared to venous blood lactate, and 12% lower (P less than 0.05) when capillary blood lactate was used. Similar discrepancies were seen in oxygen consumption. The results illustrated the importance of standardizing sampling and handling of blood specimens for lactate determination to enable direct comparisons to be made among results obtained in different studies.     

Structure and expression of the human XPBC/ERCC-3 gene involved in DNA repair disorders xeroderma pigmentosum and Cockayne's syndrome

The human XPBC/ERCC-3 was cloned by virtue of its ability to correct the excision repair defect of UV-sensitive rodent mutants of complementation group 3. The gene appeared to be in addition implicated in the human, cancer prone repair disorder xeroderma pigmentosum group B, which is also associated with Cockayne's syndrome. Here we present the genomic architecture of the gene and its expression. The XPBC/ERCC-3 gene consists of at least 14 exons spread over approximately 45 kb. Notably, the donor splice site of the third exon contains a GC instead of the canonical GT dinucleotide. The promoter region, first exon and intron comprise a CpG island with several putative GC boxes. The promoter was confined to a region of 260 bp upstream of the presumed cap site and acts bidirectionally. Like the promoter of another excision repair gene, ERCC-1, it lacks classical promoter elements such as CAAT and TATA boxes, but it shares with ERCC-1 a hitherto unknown 12 nucleotide sequence element, preceding a polypyrimidine track. Despite the presence of (AU)-rich elements in the 3'-untranslated region, which are thought to be associated with short mRNA half-life actinomycin-D experiments indicate that the mRNA is very stable (t 1/2 greater than 3h). Southern blot analysis revealed the presence of XPBC/ERCC-3 cross-hybridizing fragments elsewhere in the genome, which may belong to a related gene.     

Distribution of salmon gonadotrophin releasing-hormone in the brain and pituitary of the sea bass (Dicentrarchus labrax)

Summary The distribution of salmon gonadotrophin-releasing hormone (sGnRH) was studied in the brain and pituitary of two-year-old immature sea bass (Dicentrarchus labrax) by means of an enzymoimmunoassay (EIA) for sGnRH and immunocytochemistry. The EIA for sGnRH is a competitive assay using a tracer made of sGnRH coupled to acetylcholinesterase from an electric eel. The separation of free and bound tracer is achieved by coating the plates with mouse anti-rabbit IgG monoclonal antibodies. Displacement curves generated by sGnRH and extracts from pituitary and different brain regions showed a good parallelism allowing the assay to be used for sGnRH measurements in this species. Although all parts of the brain contained measurable levels of sGnRH, the highest concentrations were found in the pituitary, the olfactory bulbs and the telencephalon. These data were confirmed by immunocytochemistry. Cell bodies were found in the olfactory bulbs, ventral telencephalon, preoptic region and mediobasal hypothalamus. Immunoreactive fibers could be observed in all parts of the brain including the optic tectum, the cerebellum (corpus and valvula), the vagal lobe, the medulla oblongata and the rostral spinal cord. In most cases, these fibers do not form well defined bundles; however, there was clearly a continuum of immunoreactive fibers, extending from the olfactory bulbs to the pituitary, and along which all the cell bodies described above were located. In the ventral telencephalon and the preoptic region, clear pictures of varicose positive fibers contacting immunoreactive perikarya could be observed. These data indicate that sGnRH is most likely an endogenous peptide in the brain of the sea bass, although the presence of other forms of GnRH cannot be excluded at this point. This study also demonstrates that the general organization of the GnRH systems in the sea bass is highly similar to what has been described in most freshwater teleost species, and provides basis for further studies on the neuroendocrine control of gonadotrophin release in this commercially important species.     

Variation of geosmin content in Anabaena cells and its relation to nitrogen utilization

The addition of the proper amount of ammonium to the culture medium containing nitrate as nitrogen source enhanced the growth rate of Anabaena viguieri. The amount of geosmin produced by these cells varied with the concentrations of ammonium added. A negative correlation between the amount of geosmin produced and of the growth rate of cells was revealed. This was also found in cells grown on various forms of nitrogen sources. Without supply of any nitrogen compound, this organism is capable of fixing gaseous nitrogen, and under these conditions the cells grew relatively slowly. However, they produced more geosmin (per unit protein mass) than cells grown in the presence of combined nitrogen. The isolation of heterocysts, in which nitrogen was fixed, showed that these cells produced higher amounts of geosmin than vegetative cells. The possible relation of nitrogen assimilation to the production of geosmin in the cells was discussed.