The effect of cessation of growth on protein synthesis in a mutant of Chinese hamster ovary cells with a temperature-sensitive leucyl-tRNA synthetase

A temperature-sensitive mutant of Chinese hamster ovary cells with an altered leucyl-tRNA synthetase fails to grow and to incorporate amino acids into protein properly at or near the non-permissive temperature. This mutant was used to determine whether cessation of growth at the elevated temperature affected elongation factor EF-1, since the activity of EF-1 is markedly lower in non-growing cells in stationary phase than in rapidly-growing cells in exponential phase. Cell-free extracts prepared from cells maintained at 39°C for 24 h showed a marked decrease in the ability to translate natural mRNAs, compared to cells incubated at 34°C. However, the ability to translate poly(U), which requires elongation factor EF-1 (and EF-2), was not affected. Analyses of activities involved in the initiation of protein synthesis and in the activation of amino acids revealed that, with the exception of leucyl-tRNA synthetase, the rest of the components required for translation also appeared to be relatively stable even after 24 h at the elevated temperature. The effects of elevated temperature on cell-free extracts were also investigated. The results were similar to those obtained with intact cells; that is, except for leucyl-tRNA synthetase which was rapidly inactivated in vitro at 39°C, other aminoacyl-tRNA synthetases and translational components involved in chain initiation and elongation were relatively stable. Thus, no change in EF-1 activity was detected as a result of arrested cell growth, an inherent lability of the elongation factor, or metabolic degradation as a consequence of a rapid turnover rate in the absence of protein synthesis.     

Ascaris sp.: water loss during desiccation of embryonating eggs

The ability of embryonating eggs of Ascaris lumbricoides to avoid desiccation by reducing the loss of water through the egg shell was investigated. When exposed to desiccation the eggs lost water at a rate dependent upon the relative humidity and ambient temperature, eventually resulting in the collapse of the eggs and the death of the enclosed embryo. The eggs are small with a large surface to volume ratio. A low permeability to gaseous exchange thus restricts water loss while still ensuring an adequate supply of oxygen for embryonic development. Relative humidity did not appear to affect the rate of development. In eggs exposed to desiccation at various constant temperatures, the rate of water loss increased as an exponential function of increasing temperature. When eggs were exposed to various temperatures before exposure to desiccation at 22 C, the rate of water loss increased as a function of increasing pretreatment temperature. After exposure to 63–65 C, the ability of the egg shell to slow down the loss of water was destroyed. These phenomena suggest that there is not a simple “critical” or “transition” temperature, but a gradual melting of the complex mixture of components forming the lipid layer.     

Effect of temperature on locomotor activity in the goldfish (Carassius auratus) and the bluegill (Lepomis macrochirus): Presence of an ‘activity well’ in the region of the final preferendum

Goldfish (Carassius auratus) and bluegill sunfish (Lepomis macrochirus) were placed in aquaria where their locomotor activity was monitored by photocells, and tested at various acclimation temperatures over a range encompassing their final thermal preferenda. Activity was pooled over 24-hour periods to eliminate any circadian rhythm effects. Both species exhibited an activity well of reduced locomotor activity in the region of the final preferendum. Goldfish, tested either singly or in groups of 2–5 individuals, exhibited a social-interaction effect which became more pronounced at higher temperatures. These results are discussed in relation to a thermokinetic interpretation of thermo-regulatory behavior in fishes, and to the correspondence between thermal preferenda and thermal optima.     

Phenotypic changes in the chemistry of Aspergillus nidulans: Influence of culture conditions on mycelial composition

A quantitative study was made of macromolecular (nucleic acids, protein), carbohydrate and mineral (magnesium, potassium and phosphorus) components of Aspergillus nidulans in glucose limited chemostat cultures, under varying conditions of dilution rate, temperature, pH and NaCl concentration.The overall mineral content showed greatest variation in response to changes in culture salinity, which also affected the mycelial carbohydrate content. Concomitant and opposite changes in the conent of cations and carbohydrates under conditions of increasing salinity may be interpreted in terms of mycelial osmoregulation. Slight variations in DNA content but gross fluctuations in the level of RNA were noted under the different cultural conditions examined. Co-ordinate changes in RNA and Mg²⁺ contents were evident only under certain conditions: dilution rate from 0.05–0.07 h^-1 or temperature from 22–30° C. The constant molar stoichiometry between RNA and Mg²⁺ characteristic of unicellular microorganisms was not a feature of fungal growth. The protein content was most affected by shifts of temperature and reached minimal values at 25 and 50° C.The growth environment had a marked influence on the protein synthesising activity of RNA, which increased eightfold as the dilution rate was increased from 0.02–0.175 h^-1, doubled within the temperature range 20–30° C and fell by 50% between 40 and 50° C. These observations are discussed in the context of the constant ribosomal efficiency in protein synthesis hypothesis.     

Ultrastructure of cell wall and thylakoid membranes of the thermophilic cyanobacterium Synechococcus lividus under the influence of temperature shifts

The ultrastructure of the cell wall and the thylakoid membranes of the thermophilic cyanobacterium Synechococcus lividus was studied by freezefracture electron microscopy after temperature shifts. Different fracture faces of the outer, the cytoplasmic and the thylakoid membranes were demonstrated when the preparation-temperature was in the range of the optimal growth temperature at 52°C or after fixation at 52°C. In the outer membrane of the cell wall two fracture faces with holes and 7.5 nm intramembrane particles were detected. On both the outer (EF) and inner (PF) leaflet of the cytoplasmic membrane randomly distributed particles were demonstrated. The particle density on the PF-face was approx. three times that of the EF-face. The EF-face of the thylakoid membrane exposed rows of particles with an average diameter of 10 nm. The spacing between the particle rows was 35–50 nm. This regular particle arrangement on the EF-face was demonstrated only in a few cases. Mostly the intramembrane particles were distributed randomly on the thylakoid fracture faces. The particle density of thylakoids with a random distribution was approx. in the same range both on the EF-and PF-face. The EF-particles fall into four groups of 9,10,11, and 12.5 nm. The main particle class was the 10 nm class. The PF-face exposed smaller particles with two maxima at 8.5–9 nm and 10 nm. When Synechococcus lividus OH-53s was chilled to temperatures below 30–35°C before the freeze-etch preparation a phase transition took place after the temperature shift. On the fracture faces of the thylakoid and cytoplasmic membranes particle depleted areas occurred. The size of the areas were different in both membranes and dependent on the velocity of cooling. Contrary to Synechococcus lividus OH-53s in the mesophilic Synechococcus strain 6910 the phase transition point was 15°C. The lower phase transition point may be due to a higher content of unsaturated fatty acids.Dedicated to Prof. D. Peters (Hamburg) on the occasion of the 65th anniversary of his birthday     

Cell junctions in the epithelium of Bowman's capsule

Summary The intercellular connections between the epithelial cells of Bowman's capsule were investigated. It could be demonstrated that typical zonulae occludentes (tight junctions) are present in the species (rat, hamster, and Tupaia) studied. Freeze-fracturing shows a network of anastomizing strands; some species variations are described. In the rat two strands are common. In the golden hamster mostly two to four and occasionally five strands occur. In Tupaia regularly three tight junction strands are found and also gap junctions associated with the zonulae occludentes. In thin sections the goniometric analysis confirms the freeze-fracturing results and reveals attachment zones of macular shape, which are classified as intermediate junctions and desmosomes. The functional role of these cell junctions observed in the epithelium of Bowman's capsule is discussed.     

Leishmania tarentolae: streptomycin and chloramphenicol resistance of promastigotes.

Stable mutant strains of Leishmania tarentolae promastigotes resistant to chloramphenicol (CAP) were isolated by replica-plating techniques. In addition, cell lines stress-adapted to streptomycin and to high culture temperature (33 C) were obtained. Drug resistance was influenced by temperature, culture media, and plating technique. Inhibition of amino acid incorporation into protein occurs in CAP-resistant cells when exposed to 600 μg CAP/ml but this inhibition was 50–80% lower than that found in wild type sensitive cells. The primary site of CAP action appears to be inhibition of protein synthesis. CAP also adversely affected proline oxidation.     

Responses of Symbiodinium microadriaticum clade B to different environmental conditions

The effects of various environmental parameters on zooxanthellae isolated from the sea anemone Condylactis gigantea were studied under controlled conditions in the laboratory. We determined that the zooxanthellae, identified as Symbiodinium microadriaticum, (by Trench. B.) belong to clade B. These algae were exposed to a range of temperatures (17, 21.7, 26 °C), light intensities (25, 30, 45, 85 μmol q m⁻² s⁻¹) and nutrient regimes. While growth rate was relatively independent of treatment, respiration increased significantly with temperature. Both light and temperature did have a significant effect on photosynthetic parameters.The cultured zooxanthellae responded to the environment in ways similar to those of freshly isolated ones, and survived under a wide range of temperatures (17, 21.7, 26 °C).In general, clade B seems to be flexible and well adapted to the temperature range encountered in seawater throughout its wide global distribution.