Effect of short- and long-lasting chilling on pre-rRNA synthesis and transport in root meristematic cells of three soybean cultivars

<p>Autoradiographic studies of ³H-uridine incorporation (20-min incubation) and dynamics of radioactive particle translocation from nucleolus into cytoplasm (following 80-min postincubation in non-radioactive medium) in root meristematic cells of soybean have been carried out. The experiment was performed with plants subjected to 4-day acclimation in chilling or subjected to 2-hour cold stress and control plants. Three cultivars of soybean: Mazowia, Polan and Progres (cultivated in Poland) were used in the experiment.</p><p>It has been shown that in control conditions the greatest number of RNA precursor is incorporated into nucleoli after 20-min incubation. Following 80-min postincubation cytoplasm is the most radioactive area of the cell - this mainly testifies to dynamic translocation of radioactive ribosome subunits from nucleolus into cytoplasm.</p><p>In chilling conditions the reduction of ³H-uridine incorporation into cells occurs, as compared to control conditions. Plants subjected to a 4-day acclimation incorporate the radioactive precursor more intensively than plants subjected to cold stress.</p><p>Following 80-min postincubation - in the case of acclimated plants - the nucleolus is the most radioactive area of the cell, which testifies to accumulation of pre-rRNA in it. After the cold stress cytoplasm is more radioactive than the nucleolus. In all three cultivars the processes of synthesis and transport of pre-rRNA particles are similar, only their intensity is different.</p><p>Morphometric measurements of nucleoli in all cultivars subjected to 4-day chilling have shown that root cell nucleoli are larger than those in control. This phenomenon can be connected with stronger inhibition of rRNA transport than its synthesis.</p>

Concentrations of endogenous free amino acids in rabbit reticulocyte lysates and the effect of incubation

Amino acid analyses show that while the free amino acids found in the rabbit reticulocyte translation system do not increase during nuclease treatment or during prolonged storage, the endogenous levels of many amino acids are so high that the choice of a radioactive; precursor for a translation should be based not only on the abundance of the amino acid in the translation product but also on its concentration in the lysate preparation. It is shown that the variation of amino acid concentrations between different lysates is sufficiently small to allow one to use the concentrations reported in this study to calculate the amount of radioactive amino acid necessary for satisfactory incorporation.

Colonic glycoprotein secretion and calmodulin-acceptor proteins in the reserpine-treated rat

The rate of radioactive precursor-labeled colonic glycoprotein secretion in chronically reserpine-treated versus saline-injected control rats was examined. Everted colonic sacs prepared from reserpine-treated rats were found to be hypersecretory, exhibiting a basal rate of glycoprotein secretion that was threefold higher than control everted sacs. Furthermore, glycoprotein secretion in control tissue was stimulated by the secretagogue carbachol, and this stimulation was precluded by 10 microM trifluoperazine, the calmodulin antagonist. Reserpine-treated tissue, in contrast, was refractory to treatment with carbachol as well as trifluoperazine. While reserpine-treated and control colonic mucosae were demonstrated to contain equivalent levels of calmodulin via radioimmunoassay, reserpine-treated tissue was determined to lack two calmodulin-acceptor proteins with molecular weights of 29 and 47 kilodaltons. The data suggest that the mechanism by which reserpine elicits this cystic fibrosislike, hypersecretory state of glycoprotein secretion within the colonic mucosa entails the loss of calmodulin function in the regulation of this secretory process. We speculate that the biochemical defect present in cystic fibrosis could also entail such a loss of calmodulin function in the regulation of glycoprotein secretion.

The effect of cholesterol on ubiquinone and tetrahymanol biosynthesis in Tetrahymena pyriformis

The biosynthesis of ubiquinone-8 from radioactive mevalonate by cultures of Tetrahymena pyriformis is demonstrated. Under normal conditions the incorporation of this radioactive precursor into ubiquinone and the triterpenoid alcohol tetrahymanol reflects the amounts of these two compounds in the cell. Growth of T. pyriformis in the presence of cholesterol results in a complete inhibition of incorporation of radioactive mevalonate into tetrahymanol while there is a corresponding increase of radioactive incorporation into ubiquinone. This increased incorporation of mevalonic acid into ubiquinone must reflect a reduced level of mevalonic acid in the cell under these conditions and is not due to increased ubiquinone biosynthesis, indicating tight regulation of the pathway prior to mevalonate formation.

Ultraviolet-induced DNA excision repair in human B and T lymphocytes. III. Repair in lymphocytes from chronic lymphocytic leukaemia

There have been conflicting reports about the capacity of lymphocytes from individuals with chronic lymphocytic leukaemia (CLL) to undertake u.v.-induced DNA repair. We have examined repair in CLL cells and in control and age-matched purified B and T lymphocytes by a technique independent of incorporation of radioactive precursor, i.e. by the recovery of normal sedimentation behaviour of nucleoid bodies obtained from these cells by lysis in high salt and non-ionic detergent. Recovery of normal sedimentation is associated with restoration of DNA supercoiling. CLL cells were found to be as sensitive to u.v. and to repair at similar rates as age-matched B controls. They are considerably more sensitive than young B cells and repair less efficiently. Reasons for the reported discrepancies in CLL repair are discussed.

Cellular interaction between fixed and living cells. Transfer of radioactive materials from living cells to fixed cells.

Transfer of radioactive materials to fixed cells from an overlying layer of living cells has been examined to determine whether fixed cells can act as acceptors of glycosyltransferases of living cells. After the incubation of living cells were removed by EDTA treatment, and the radioactivity associated with the fixed cells was determined. Lipids, proteins and carbohydrates were found to be transfered from the living cells to the fixed cells. The amount of radioactivity transferred to the fixed cells was dependent on the number of both fixed and living cells and increased with the time of incubation. When fixed cells were treated with chloroform-methanol before the addition of living cells, the transfer of both lipids and proteins to the fixed cells decreased drastically, but only a slight decrease incarbohydrate transfer was observed. Most of the radioactive materials transferred from living cells labeled with glucosamine or fucose to chloroform-methanol-treated fixed cells were solubilized by trypsin but not by the detergents tested. Approximately 55% of the materials transferred from the cells labeled with glucosamine could be solubilized by hyaluronidase and chondroitinase, and the rest was solubilized by neuraminidase and a glycosidase mixture. The treatment of chloroform-methanol-extracted fixed cells with trypsin caused a significant decrease in the transfer from cells labeled with glucosamine. When nucleotide sugars were used as the radioactive precursor, no significant amount of radioactivity was transferred to the fixed cells.