The biosynthesis of a phytoalexin, beta-thujaplicin, in Cupressus lusitanica cell cultures can be stimulated by a yeast elicitor, H(2)O(2), or methyl jasmonate. Lipoxygenase activity was also stimulated by these treatments, suggesting that the oxidative burst and jasmonate pathway may mediate the elicitor-induced accumulation of beta-thujaplicin. The elicitor signalling pathway involved in beta-thujaplicin induction was further investigated using pharmacological and biochemical approaches. Treatment of the cells with calcium ionophore A23187 alone stimulated the production of beta-thujaplicin. A23187 also enhanced the elicitor-induced production of beta-thujaplicin. EGTA, LaCl(3), and verapamil pretreatments partially blocked A23187- or yeast elicitor-induced accumulation of beta-thujaplicin. These results suggest that Ca(2+) influx is required for elicitor-induced production of beta-thujaplicin. Treatment of cell cultures with mastoparan, melittin or cholera toxin alone or in combination with the elicitor stimulated the production of beta-thujaplicin or enhanced the elicitor-induced production of beta-thujaplicin. The G-protein inhibitor suramin inhibited the elicitor-induced production of beta-thujaplicin, suggesting that receptor-coupled G-proteins are likely to be involved in the elicitor-induced biosynthesis of beta-thujaplicin. Indeed, both GTP-binding activity and GTPase activity of the plasma membrane were stimulated by elicitor, and suramin and cholera toxin affected G-protein activities. In addition, all inhibitors of G-proteins and Ca(2+) flux suppressed elicitor-induced increases in lipoxygenase activity whereas activators of G-proteins and the Ca(2+) signalling pathway increased lipoxygenase activity. These observations suggest that Ca(2+) and G-proteins may mediate elicitor signals to the jasmonate pathway, and the jasmonate signalling pathway may then lead to the production of beta-thujaplicin. (+info)
Jasmonate and ethylene signalling and their interaction are integral parts of the elicitor signalling pathway leading to beta-thujaplicin biosynthesis in Cupressus lusitanica cell cultures.
Roles of jasmonate and ethylene signalling and their interaction in yeast elicitor-induced biosynthesis of a phytoalexin, beta-thujaplicin, were investigated in Cupressus lusitanica cell cultures. Yeast elicitor, methyl jasmonate, and ethylene all induce the production of beta-thujaplicin. Elicitor also stimulates the biosynthesis of jasmonate and ethylene before the induction of beta-thujaplicin accumulation. The elicitor-induced beta-thujaplicin accumulation can be partly blocked by inhibitors of jasmonate and ethylene biosynthesis or signal transduction. These results indicate that the jasmonate and ethylene signalling pathways are integral parts of the elicitor signal transduction leading to beta-thujaplicin accumulation. Methyl jasmonate treatment can induce ethylene production, whereas ethylene does not induce jasmonate biosynthesis; methyl jasmonate-induced beta-thujaplicin accumulation can be partly blocked by inhibitors of ethylene biosynthesis and signalling, while blocking jasmonate biosynthesis inhibits almost all ethylene-induced beta-thujaplicin accumulation. These results indicate that the ethylene and jasmonate pathways interact in mediating beta-thujaplicin production, with the jasmonate pathway working as a main control and the ethylene pathway as a fine modulator for beta-thujaplicin accumulation. Both the ethylene and jasmonate signalling pathways can be regulated upstream by Ca(2+). Ca(2+) influx negatively regulates ethylene production, and differentially regulates elicitor- or methyl jasmonate-stimulated ethylene production. (+info)
Cellular fine structures and histochemical reactions in the tissue of a cypress twig preserved in Baltic amber.
A twig of a cypress plant preserved for ca. 45 Myr in Baltic amber was analysed by light and electron microscopy. Cross-sections of the whole plant showed an almost intact tissue of the entire stem and leaves, revealing, to our knowledge, the oldest and most highly preserved tissue from an amber inclusion reported so far. The preparations are based on a new technique of internal imbedding, whereby the hollow spaces within the inclusion are filled with synthetic resin which stabilizes the cellular structures during the sectioning procedure. Cytological stains applied to the sections reacted with cell walls and nuclei. A strong green auto-fluorescence of the cuticle and the resin canals in the leaves was observed. Transmission electron micrographs revealed highly preserved fine structures of cell walls, membranes and organelles. The results were compared with taxonomically related recent Glyptostrobus and Juniperus plants. (+info)
Human CD1-restricted T cell recognition of lipids from pollens.
Plant pollens are an important source of environmental antigens that stimulate allergic responses. In addition to acting as vehicles for foreign protein antigens, they contain lipids that incorporate saturated and unsaturated fatty acids, which are necessary in the reproduction of higher plants. The CD1 family of nonpolymorphic major histocompatibility complex-related molecules is highly conserved in mammals, and has been shown to present microbial and self lipids to T cells. Here, we provide evidence that pollen lipids may be recognized as antigens by human T cells through a CD1-dependent pathway. Among phospholipids extracted from cypress grains, phosphatidyl-choline and phosphatidyl-ethanolamine were able to stimulate the proliferation of T cells from cypress-sensitive subjects. Recognition of phospholipids involved multiple cell types, mostly CD4(+) T cell receptor for antigen (TCR)alphabeta(+), some CD4(-)CD8(-) TCRgammadelta(+), but rarely Valpha24i(+) natural killer-T cells, and required CD1a(+) and CD1d(+) antigen presenting cell. The responding T cells secreted both interleukin (IL)-4 and interferon-gamma, in some cases IL-10 and transforming growth factor-beta, and could provide help for immunoglobulin E (IgE) production. Responses to pollen phospholipids were maximally evident in blood samples obtained from allergic subjects during pollinating season, uniformly absent in Mycobacterium tuberculosis-exposed health care workers, but occasionally seen in nonallergic subjects. Finally, allergic, but not normal subjects, displayed circulating specific IgE and cutaneous weal and flare reactions to phospholipids. (+info)
Streptomyces jietaisiensis sp. nov., isolated from soil in northern China.
An actinomycete, strain FXJ46(T), was isolated from cypress forest soil in northern China and shown to have chemotaxonomic and morphological properties consistent with streptomycetes. It developed greyish aerial mycelium and pinkish-brown substrate mycelium on oatmeal agar. Phylogenetic analyses based on an almost complete 16S rRNA gene sequence of the strain and on the 120 nucleotide variable gamma-region of this molecule showed that it formed a distinct (but closely associated) line with Streptomyces griseoaurantiacus DSM 40430(T) in Streptomyces trees. However, the DNA-DNA relatedness between the two strains was only 48.8%. A number of phenotypic properties also readily distinguished the isolate from S. griseoaurantiacus and related Streptomyces species with validly published names. It is proposed, therefore, that this organism be classified as a novel species of the genus Streptomyces, for which the name Streptomyces jietaisiensis sp. nov. is proposed. The type strain is FXJ46(T) (=AS 4.1859(T)=JCM 12279(T)). (+info)
In vitro and in vivo biological activities of old and fresh Cupressus arizonica pollen.
BACKGROUND: Respiratory allergy to the pollen of Cupressaceae is becoming more and more common every year in the Mediterranean area. OBJECTIVE: The purpose of this study was to see whether the allergenic potency of Cupressus arizonica pollen diminished after a 6-year period (1994-2000). MATERIALS AND METHODS: Among the Cupressaceae, we selected the pollen of C arizonica. The mode of sampling in 1994 and in 2000 was the same and the pollen was collected on the same tree and stored at room temperature. To compare its biological and allergenic activities data was collected with the following methods: cytohistology of Alexander, 2,3,5-triphenyltetrazolium chloride enzyme staining, skin testing, nasal provocation test, radioallergosorbent test (RAST), RAST inhibition, sodium dodecyl sulfate polyacrylamide gel electrophoresis, and immunoblotting to detect protein content. Thirty-eight patients with respiratory allergy to Cupressaceae were selected. RESULTS: We found no decrease in the allergenic potency of the pollen, but did find that viability and germinating power had disappeared completely after 30 to 40 days. Moreover, the amount of protein in the old pollen was half the amount found in the fresh one. Skin prick testing showed identical results with the old and the fresh pollens. CONCLUSIONS: The allergenic in vivo and in vitro activity of cypress pollen is retained for years after its collection. This activity seems to be independent of the viability of pollen grains and of the total protein content. This may explain the presence of clinical symptoms in patients out of the pollen season. (+info)
Identification of italian cypress (Cupressus sempervirens) pollen allergen Cup s 3 using homology and cross-reactivity.
BACKGROUND: The prevalence of seasonal allergic diseases of the upper airways is increasing in industrialized countries. The Cupressaceae are important causes of pollinosis, particularly in Europe. OBJECTIVE: To determine whether the pollen from Cupressus sempervirens (Italian cypress) contains a pathogenesis-related group 5 (PR-5) protein, similar to that found in other allergenic Cupressaceae pollens. METHODS: Messenger RNA was purified from Italian cypress pollen, and complementary DNA (cDNA) was synthesized. cDNAs for PR-5 proteins were amplified by polymerase chain reaction and extended by rapid amplification of cDNA ends methods. Recombinant Cup s 3 was expressed in Escherichia coli as a fusion protein. Inhibition enzyme-linked immunosorbent assays were used to test the allergenicity of Cup s 3. RESULTS: Three cDNAs were cloned. These clones had approximately 95% identity to Jun a 3 and Cup a 3. Recombinant Cup s 3.0102 maltose-binding protein inhibited the IgE from most patients from binding to an extract of Italian cypress. The extent of inhibition suggested that antibodies to Cup s 3 were a prominent component of the IgE response to Italian cypress pollen. CONCLUSION: Cup s 3, an allergen of Italian cypress pollen, was identified based on cross-reactivity and homology with other pollen PR-5 proteins, despite an apparently low level of protein expression. Variations in the content of Cup s 3 in the pollen from different regions or trees should be considered in the choice of extracts for diagnosis and specific immunotherapy for Italian cypress pollen hypersensitivity. (+info)
Antifungal effect of eugenol and nerolidol against Microsporum gypseum in a guinea pig model.
Essential oils have been widely used in anti-infectious application. In the present study, we elucidated the antifungal activities of eugenol and nerolidol isolated from Japanese cypress oil in a guinea pig model infected by Microsporum gypseum (M. gypseum). A minimal inhibitory concentration (MIC), skin lesion scoring, hair culture and histopathologic examination of skin tissues were performed to evaluate the antifungal effect of these oils. The MICs of eugenol, nerolidol and econazole (positive control) were 0.01-0.03% and 0.5-2% and 4-16 microg/ml, respectively. Based on these MICs, eugenol and nerolidol were adjusted to 10% concentration with a base of Vaseline petroleum jelly and were applied topically to the skin lesion infected with M. gypseum daily for 3 weeks. Both eugenol and nerolidol were clinically effective at improving the lesion during the first week of application, as determined by skin lesion scoring. Nerolidol improved the skin lesions infected by M. gypseum, but eugenol did not, as determined in the hair culture test. Histopathologic examination revealed that the eugenol- and nerolidol-treated groups had a lower degree of hyperkeratosis and inflammatory cell infiltration than the positive control. Taken together, these results suggest that eugenol and nerolidol could apply supplementary antifungal agents. (+info)