Calcineurin activation is only one calcium-dependent step in cytotoxic T lymphocyte granule exocytosis

We have tested the idea that calcineurin, a calcium-dependent phosphatase that is critical for activating cytokine gene expression in helper T cells, plays a role in lytic granule exocytosis in cytotoxic T lymphocytes (CTLs). We used TALL-104 human leukemic CTLs as a model. Our results confirm an earlier report (Dutz, J. P., Fruman, D. A., Burakoff, S. J., and Bierer, B. E. (1993) J. Immunol. 150, 2591-2598) that immunosuppressive drugs inhibit exocytosis in CTLs stimulated either via the T cell receptor (TCR) or via TCR-independent soluble agents. Of the two recently reported alternate targets of immunosuppressive drugs (Matsuda, S., Shibasaki, F., Takehana, K., Mori, H., Nishida, E., and Koyasu, S. (2000) EMBO Rep. 1, 428-434 and Matsuda, S., and Koyasu, S. (2000) Immunopharmacology 47, 119-125), JNK is not required for lytic granule exocytosis, but we were not able to exclude a role for P38. Exocytosis could be inhibited by expressing GFP fused to a C-terminal fragment of CAIN (cabin 1), but not by expressing VIVIT-GFP. Finally, expressing either full-length or truncated constitutively active mutant calcineurin A enhanced lytic granule exocytosis. However, the mutant calcineurin was unable to support exocytosis when cells were stimulated in the absence of Ca2+ influx. Taken together, our results support the idea that activation of calcineurin is required for lytic granule exocytosis but suggest that it is not the sole Ca2+-dependent step.     

Stability and instability processes in the calli of Fagopyrum tataricum that have different morphogenic potentials

Plant Cell, Tissue and Organ Culture (PCTOC) - The morphogenic callus (MC) of Fagopyrum tataricum contains a large amount of flavonoids, especially rutin, and exhibits a high level of antioxidant...     

Application of the salt stress to the protoplast cultures of the carrot (Daucus carota L.) and evaluation of the response of regenerants to soil salinity

This is the first study to generate carrot plants for enhanced salinity tolerance using a single-cell in vitro system. Protoplasts of three carrot accessions were exposed to treatment by seven different concentrations of NaCl (10–400 mM). Salt concentrations higher than 50 mM decreased plating efficiency and those of 200–400 mM of NaCl completely arrested mitotic divisions of cultured cells. The protoplast-derived plants from the control and 50–100 mM NaCl treatment were subjected to an 8-week salt stress in greenhouse conditions induced by salinized soil (EC 3 and 6 mS cm^?1). 50 mM NaCl stress applied in vitro induced polyploidy among regenerated plants. The regenerants obtained from the 50 and 100 mM NaCl-treated protoplast cultures grown in saline soil had a higher survival rate compared to the regenerants from the control cultures. The salt-stressed plants accumulated anthocyanins in petioles and produced denser hairs on leaves and petioles in comparison to the control plants. Salt stress influenced pollen viability and seed setting of obtained regenerants. The results suggest that salt stress applied in vitro in protoplast cultures creates variation which allows alleviating the negative effects of salt stress on the development and reproduction of the carrot.

     

The influence of American mink odour on the spatial distribution and behaviour of water voles

The perception and assessment of predation risk often cause changes in the activities of animals and induce behavioural responses that may in turn affect their movements and distribution. To simulate high predation risk in a midfield pond riparian habitat, we used fresh faeces from ranch American mink Neovison vison and recorded behavioural responses of water voles Arvicola amphibius. In areas where mink odour was deployed, the numbers of captured vole individuals and their trappability were significantly lower than in control areas. Several voles migrated from the zones with deployed mink faeces to the areas without faeces, thus proving that increased predation risk affects the distribution of individuals in a population. The response to mink odour was much more pronounced in females than in males; in areas with deployed mink faeces, not a single female was trapped. We conclude that although American mink is a non‐native, invasive predator, water voles respond to mink odour by reducing their activity and/or by avoiding places with higher predation risk.     

Lipoxygenase interactions with natural flavonoid, quercetin, reveal a complex with protocatechuic acid in its X-ray structure at 2.1 A resolution

PUFA metabolites have a profound effect on inflammatory diseases and cancer progression. Blocking their production by inhibiting PUFA metabolizing enzymes (dioxygenases: cyclooxygenases and LOXs) might be a successful way to control and relieve such problems, if we learn to better understand their actions at a molecular level. Compounds with strong antioxidative and free radical scavenging properties, such as polyphenols, could be effective in blocking PUFA activities, and natural flavonoids possess such qualities. Quercetin belongs to the group of natural catecholic compounds and is known as a potent, competitive inhibitor of LOX. Structural analysis reveals that quercetin entrapped within LOX undergoes degradation, and the resulting compound has been identified by X-ray analysis as protocatechuic acid (3,4-dihydroxybenzoic acid) positioned near the iron site. Its C3-OH group points toward His523, C4-OH forms a hydrogen bond with O=C from the enzyme's C-terminus, and the carboxylic group is incorporated into the hydrogen bonding network of the active-site neighborhood via Gln514. This unexpected result, together with our previous observations concerning other polyphenols, yields new evidence about the metabolism of natural flavonoids. These compounds might be vulnerable to the co-oxidase activity of LOX, leading to enzyme-stimulated oxidative degradation, which results in an inhibitor of a lower molecular weight.     

Impaired nuclear localization of vitamin D receptor in leukemia cells resistant to calcitriol-induced differentiation

Calcitriol, the hormonal form of vitamin D₃, induces differentiation of monocytic leukemia cell lines in vitro, without inducing cytotoxicity of the cells. Besides this broad in vitro activity, a clinical implementation of calcitriol, or its analogs, as agents for differentiation therapy has been unsuccessful until now. A better understanding of cellular activities of calcitriol necessary for completion of cell differentiation program could help find better solutions for differentiation therapy of myeloid leukemias. In this paper we describe work carried on subline of acute monocytic leukemia, THP-1 resistant to calcitriol induced differentiation. This resistance correlates with impaired nuclear localization of vitamin D receptor, but not with its total expression in the cells. It also correlates with the resistance to calcitriol-induced growth inhibition, and to phorbol myristate acetate (PMA)-induced cell differentiation.