Modulation of the expression of interleukin 2 receptors of human thymocytes by recombinant interleukin 2

The effect of purified recombinant interleukin 2 on the expression of the receptors for interleukin 2 by human thymocytes was examined. Interleukin 2 augmented the expression of interleukin 2 receptors and interferon-gamma synthesis by thymocytes activated with concanavalin A, and it was required to maintain the growth of thymocytes in vitro and the expression of interleukin 2 receptors. The increase observed in the number of receptor bearing thymocytes and in the density of receptors due to interleukin 2 occurred within the first 2 days of culture. Dexamethasone inhibited the expression of interleukin 2 receptors, the synthesis of interferon-gamma, and the early proliferation and protein synthesis of lectin-activated thymocytes during the first 2 days of culture. The inhibitory effect of dexamethasone on the expression of interleukin 2 receptors and on the synthesis of interferon-gamma was reversed by interleukin 2, whereas its effect on proliferation and on protein synthesis during the first two days of culture was not reversed by interleukin 2. Interleukin 2 induced the proliferation of thymocytes in vitro, even in the absence of activation by lectin; however, the number of cells displaying receptors which could be detected with anti-Tac remained low throughout the first week of culture and interferon-gamma synthesis was not observed. Nonetheless, interleukin 2-induced proliferation was inhibited by anti-Tac on a dose dependent manner. The results of the study document that recombinant interleukin 2, like purified natural interleukin 2, is required for the expression of interleukin 2 receptors, for interferon-gamma synthesis, and for the growth of thymocytes in vitro.     

Stimulation of the hexose-monophosphate pentose pathway by delta 1-pyrroline-5-carboxylic acid in human fibroblasts.

L-pyrroline-5-carboxylic acid, an intermediate in the interconversions of glutamic acid, ornithine and proline, is a potent stimulator of the hexose-monophosphate pentose pathway in cultured human fibroblasts. These studies suggest that pyrroline-5-carboxylate reductase, which catalyzes the conversion of pyrroline-5-carboxylate to proline coupled with the oxidation of NADPH, provides the NADP for the observed activation of the hexose-monophosphate pentose pathway.     

Induction of rat jejunal epithelial cell expression of sucrase-isomaltase by glucocorticoids in primary cell culture and in vivo

The cell cycle phase that mediates the induction of intestinal sucrase-isomaltase (SI) expression by glucocorticoids was investigated by measuring migration rates of 3H-DNA-labeled and of SI-containing epithelial cells by autoradiography and indirect immunofluorescent staining after simultaneous administration of [3H]thymidine and cortisone to 12-d-old rat pups. By 24 and 48 h, lead 3H-DNA-labeled cells had migrated 7.8 and 12.4 cell positions higher on the villus than lead cells expressing SI. Cell migration rates from 12 to 24 h and 24 to 48 h were 0.68 and 0.97 cell position/h. Thus, commitment to SI expression occurred in cells 11.5-12.8 h after the S phase, which is calculated to be in the G1 phase. To determine whether committed cells need to replicate to express SI, cell differentiation was examined in primary cultures of crypt cells originating from corticosterone-treated rats. About two-thirds of cultured cells were retarded in the S phase after plating, as judged by no increase of DNA labeling indices, no change in epithelial cell number, and the absence of mitosis (less than 0.01%). The proportion of cells expressing SI increased from 0 to 6-8% between 12 and 24 h, and reached 48% 48 h after plating on collagen-coated dishes. SI expression did not occur in cells plated on glass or plastic surfaces. Pulse labeling with [35S]methionine confirmed that de novo synthesis of SI occurred in cell cultures. Thus, additional cell cycling of committed cells occurring in vivo is not obligatory for the expression of SI.     

Embryogenesis and plant regeneration from anther culture of bamboo (Sinocalamus latiflora (Munro) McClure)

Embryogenic callus was initiated from bamboo (Sinocalumus satiflora (Munro) McClure) anthers cultured on N₆ medium supplemented with 1 mg/l 2,4-D, 1 mg/l BA, 2 g/l charcoal, 0.8% agar (Sigma) and 9% sucrose. Anthers with microspores at miduninucleate to early-binucleate stages showed better rate of response for callus induction. Prolonged culture of these embryogenic calli on the original medium or subculture to an auxin-free medium resulted in embryoid formation and their subsequent germination to form rooted plantlets. Chromosome counts from root-tip cells of anther-derived plant indicated that they were haploid (N=36).Abbreviations N₆ Chu et al. (1975) - MS Murashige and Skoog (1962) - 2,4-D 2,4-dichlorophenoxyacetic acid - NAA -naphthaleneacetic acid - BA 6-benzylaminopurine     

Protein topography of the 40 S ribosomal subunit from Saccharomyces cerevisiae as shown by chemical cross-linking

Protein-protein cross-linking was used to examine the spatial arrangement of proteins within the 40 S ribosomal subunits of Saccharomyces cerevisiae. Purified ribosomal subunits were treated with either 2-iminothiolane or dimethyl 3,3'-dithiobispropionimidate under conditions such that the ribosomal particle was intact and that formation of 40 S subunit dimers was minimized. Proteins were extracted from the treated subunits and fractionated on Sephadex G-150 or by acid-urea-polyacrylamide gel electrophoresis. Cross-linked proteins in these fractions were analyzed by two-dimensional diagonal sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Constituent members of cross-linked pairs were radiolabeled with 125I and identified by two-dimensional gel electrophoresis and comparison with nonradioactive ribosomal protein markers. Forty-two pairs involving 25 of the 32 40 S subunit proteins were identified. Many proteins were detected in several cross-linked dimers. These proteins with multiple cross-links form foci for the construction of a schematic model of the spatial arrangement of proteins within the 40 S subunit.     

A kinetic description of antifreeze glycoprotein activity

The antifreeze glycoproteins (AFGP) of polar fish have the ability to depress the freezing temperature of water approximately 500 times the amount expected based on the number of AFGP molecules in solution; yet AFGP solutions have a purely colligative melting point depression. The difference of solution melting and freezing temperatures is the antifreeze activity of AFGP. One characteristic of AFGP activity that requires further examination is the effect of concentration on antifreeze activity, especially whether the activity saturates at high concentrations or the measured activity increases ad infinitum. This study first surveys the activity of the various antifreeze components from both Pagothenia borchgrevinki and the Arg-containing antifreeze glycoprotein from Eleginus gracilis (EgAF). It was found that all AFGP components examined have a plateau in activity at high concentration, but the actual value of the plateau activity differs between the different length AFGP components and between AFGP and EgAF. While the low molecular weight components of both AFGP and EgAF lose activity at deep supercooling, at high concentration activity is restored. The activity data is then shown to fit a reversible kinetic model of AFGP activity, and the coefficients obtained are used to compare the activity differences between AFGP components and between AFGP and EgAF. The model is also shown to describe the activity of the antifreeze protein of the fish Pseudopleuronectes americanus and the thermal hysteresis protein of the insect, Tenebrio molitor.     

Regulation by interleukin 2 of interleukin 2 receptors and gamma-interferon synthesis by human thymocytes: augmentation of interleukin 2 receptors by interleukin 2

The role of interleukin 2 (IL 2) on the expression of IL 2 receptors and on the synthesis of gamma-interferon (gamma-IFN) by human thymocytes was investigated. Human thymocytes isolated from specimens obtained during cardiac surgery of infants and children were induced with one or all of the following agents: IL 2, concanavalin A (Con A), and 12-O-tetradecanoylphorbol 13-acetate (TPA). The expression of IL 2 receptors and gamma-IFN titers were determined. The results indicate that thymocytes cultured in complete medium do not express receptors for IL 2, nor did IL 2 by itself induce the expression of IL 2 receptors. Con A induced the expression of IL 2 receptors by a moderate number of the thymocyte population and induced the synthesis of low amounts of gamma-IFN. Preincubation of thymocytes with TPA increased the response to Con A; both the number of thymocytes expressing receptors and the synthesis of gamma-IFN were increased. Addition of IL 2 to these cultures further augmented the expression of IL 2 receptors and gamma-IFN synthesis and resulted in the optimal expression of IL 2 receptors and maximal gamma-IFN synthesis. The expression of IL 2 receptors could be detected within 24 hr and preceded the induction of proliferation; it was therefore probably not due to the clonal expansion of a population of receptor-bearing thymocytes. Conversely, inhibition of IL 2 synthesis with dexamethasone (Dex) by thymocytes activated with Con A, or inhibition of the function of IL 2 receptors by anti-Tac, resulted in a decrease in the number of IL 2 receptor-bearing thymocytes activated with Con A, or inhibition of the function of IL 2 receptors by anti-Tac, resulted in a decrease in the number of IL 2 receptor-bearing thymocytes and of gamma-IFN synthesis. Thymocytes activated with TPA and Con A were more resistant to the inhibitory effects of Dex on the expression of IL 2 receptors than thymocytes activated with Con A alone. Maximal inhibition of the expression of IL 2 receptors and of gamma-IFN synthesis was achieved as a result of the synergistic effect of anti-Tac with Dex. Therefore, when IL 2 was prevented from binding to the receptors, and IL 2 synthesis was inhibited, the number of thymocytes expressing IL 2 receptors was sharply reduced and gamma-IFN synthesis was markedly inhibited.(ABSTRACT TRUNCATED AT 400 WORDS)     

Genetic engineering of coryneform bacteria

The coryneforms are a diverse group of bacteria which includes animal and plant pathogens as well as non-pathogenic bacteria. Although they are of significant economic and health importance, their genetics is poorly understood. The development of genetic engineering techniques for coryneforms and initial gene cloning studies are discussed.