The role of microtubules in guard cell function. (1/22)

Guard cells are able to sense a multitude of environmental signals and appropriately adjust the stomatal pore to regulate gas exchange in and out of the leaf. The role of the microtubule cytoskeleton during these stomatal movements has been debated. To help resolve this debate, in vivo stomatal aperture assays with different microtubule inhibitors were performed. We observed that guard cells expressing the microtubule-binding green fluorescent fusion protein (green fluorescent protein::microtubule binding domain) fail to open for all major environmental triggers of stomatal opening. Furthermore, guard cells treated with the anti-microtubule drugs, propyzamide, oryzalin, and trifluralin also failed to open under the same environmental conditions. The inhibitory conditions caused by green fluorescent protein::microtubule binding domain and these anti-microtubule drugs could be reversed using the proton pump activator, fusicoccin. Therefore, we conclude that microtubules are involved in an upstream event prior to the ionic fluxes leading to stomatal opening. In a mechanistic manner, evidence is presented to implicate a microtubule-associated protein in this putative microtubule-based signal transduction event.  (+info)

Flow cytometric analysis and chromosome sorting of barley (hordeum vulgare L). (2/22)

Flow cytometric analysis was systematically performed to optimize the concentration and duration of hydroxyurea (DNA synthesis inhibitor) and trifluralin (metaphase blocking reagent) treatments for synchronizing the cell cycle and accumulating metaphase chromosomes in barley root tips. A high metaphase index (76.5% in the root tip meristematic area) was routinely achieved. Seedlings of about 1.0-cm length were treated with 1.25 mM hydroxyurea for 14 h to synchronize the root tip meristem cells at the S/G2 phase. After rinsing with hydroxyurea, the seedlings were incubated in a hydroxyurea-free solution for 2 h and were treated with 1 microM trifluralin for 4 h to accumulate mitotic cells in the metaphase. The consistent high metaphase index depended on the uniform germination of seeds prior to treatment. High-quality and high-quantity isolated metaphase chromosomes were suitable for flow cytometric analysis and sorting. Flow karyotypes of barley chromosomes were established via univariate and bivariate analysis. A variation of flow karyotypes was detected among barley lines. Two single chromosome types were identified and sorted. Bivariate analysis showed no variation among barley individual chromosomes in AT and GC content.  (+info)

Effect of nutritional and environmental conditions on the production and composition of rhamnolipids by P. aeruginosa UG2. (3/22)

The production of rhamnolipid biosurfactants by P. aeruginosa UG2 was examined under different culture conditions. Rhamnolipid yield was affected by the nature of the carbon sources, the nutrient concentrations, pH, and age of the culture. Hydrophobic substrates like corn oil, lard (rich in unsaturated and saturated fat), and long chain alcohols maximized biosurfactant production (100-165 mg/g substrate). Hydrophilic substrates like glucose, and succinic acid delivered poor yields (12-36 mg/g substrate). Rhamnolipid production was greater when N as (NH4)(2)SO4 and trace metals were added in several periodic doses rather than at the beginning of the process. Increased biosurfactant production was seen in cultures maintained at neutral pH relative to cultures allowed to develop acidic conditions (pH = 6.25). Although the level of rhamnolipid production was affected by culture conditions, the distribution of rhamnolipid subspecies did not vary between cultures. A dirhamnolipid species containing two 10 carbon alpha-hydroxy fatty acids [Rh2C10C10] was the most abundant in the mixtures (60.6 mol%), while the levels of the monorhamnolipid [RhC10C10] (20.7 mol%) and two dirhamnolipids [Rh2C10C12 and its dehydro variant Rh2C10C12-H2] (18.7 mol%) were similar. Biosurfactant mixtures produced with corn oil as sole carbon source solubilized the herbicide trifluralin [2,6-dinitro-N,N-dipropyl-4-(trifluoromethyl)benzamine] to a greater extent. This suggests that the presence of incompletely metabolized hydrophobic by-products acting as co-solvents can increase the solubilization capacity of biosurfactant mixtures.  (+info)

Transdermal penetration of atrazine, alachlor, and trifluralin: effect of formulation. (4/22)

Commercial formulations of herbicides contain surfactants and other compounds to increase absorption by targeted plants. These chemicals, however, are also potential penetration enhancers for mammalian skin. The effect of formulation on dermal absorption of the herbicides atrazine, alachlor, and trifluralin and their commercial formulations Aatrex, Lasso, and Treflan was determined. In vitro absorption studies were performed by placing hairless mouse skin in a Bronough flow-through diffusion system. Donor solution was spiked with (14)C-labeled herbicide, and its penetration through the skin was monitored in 90-min fractions. Results demonstrate that dermal penetration of commercially formulated compound was significantly greater (p < 0.05) than that of the pure compound at the same concentration. The physical properties of a herbicide predicted penetration (r(2) = 0.97-0.99) for commercial formulations but were not as effective at predicting absorption for the pure compounds (r(2) = 0.51-0.71). The solvents associated with the hydrophobic herbicide Treflan altered dermal penetration of the more hydrophilic herbicides Lasso and Aatrex. Furthermore, although the most hydrophobic compound had the least penetration, it accumulated in the stratum corneum at the greatest rate. These studies can have important implications on future experiments performed to predict percutaneous penetration of herbicides.  (+info)

Benzoate X receptors alpha and beta are pharmacologically distinct and do not function as xenobiotic receptors. (5/22)

The Xenopus benzoate nuclear hormone receptors, BXRalpha and BXRbeta, share 82% identity within their ligand-binding domains and are classified as members of the NR1I2 subfamily that includes the mammalian steroid and xenobiotic receptor, SXR/PXR. Although alkyl benzoates have been identified as endogenous ligands, the exact role of the benzoate receptors in amphibian physiology has not been established. In this report, we show that BXRalpha and BXRbeta are pharmacologically distinct from each other: BXRalpha is more promiscuous than BXRbeta with respect to both ligand specificity and co-activator recruitment. BXRalpha can be transactivated by a number of benzoate derivatives including 4-amino-butylbenzoate (4-ABB), 4-hydroxy-butylbenzoate (4-HBB), 3-hydroxy ethyl benzoate (3-HEB), and benzyl benzoate, but only 4-HBB acts as an agonist for both receptors. Furthermore, BXRalpha-specific agonists such as 4-ABB, chlorpyrifos, and trifluralin act as antagonists on BXRbeta. BXRs are widely distributed in adult tissues but do not show any enrichment in liver and intestine, major sites of SXR/PXR expression that are critical in xenobiotic metabolism. Neither BXR shows the broad specificity toward steroids or xenobiotics exhibited by SXR/PXR. Therefore, we conclude that the BXRs are pharmacologically distinct from each other and unlikely to serve as xenobiotic sensors.  (+info)

Chemosensory responses of a protozoan are modified by antitubulins. (6/22)

Modification of a behavioral response of a marine dinoflagellate to chemical cues is described. Negative response to choline was modified by the antitubulins vincristine, vinblastine, griseofulvin, and trifluralin, but not by colchicine. Positive responses to 3,4-dihydroxyphenylalanine were unaffected by these drugs.  (+info)

Isolation of a protein interacting with Vfphot1a in guard cells of Vicia faba. (7/22)

A recent study has demonstrated that phototropins act as blue light receptors in stomatal guard cells. However, the downstream components responsible for phototropin signaling are largely unknown. In this study, using a yeast two-hybrid system, we isolated a Vicia faba protein that has a high similarity to dynein light chain in the C terminus, which interacts with Vicia faba phototropin 1a (Vfphot1a). Protein-blot and two-hybrid analyses revealed that Vfphot1a interacting protein (VfPIP) bound to the N-terminal [corrected] region of Vfphot1a but did not bind to Vfphot1b. The interaction between VfPIP and Vfphot was indicated by a pull-down assay. Northern analysis revealed that the transcription level of VfPIP gene was more abundant in guard cells than in other tissues or cell types. The transiently expressed fusion protein of VfPIP-green fluorescent protein was localized on cortical microtubules in Vicia guard cells. Microtubule-depolymerizing herbicides partially inhibited both blue light-dependent H(+) pumping in Vicia guard cell protoplasts and stomatal opening in the Vicia epidermis. From these results, we conclude that VfPIP may act as a downstream component of phototropin (Vfphot1a) in blue light signaling in guard cells. The possible role of VfPIP in blue light signaling of guard cells is discussed.  (+info)

Membrane trafficking and osmotically induced volume changes in guard cells. (8/22)

Guard cells rapidly adjust their plasma membrane surface area while responding to osmotically induced volume changes. Previous studies have shown that this process is associated with membrane internalization and remobilization. To investigate how guard cells maintain membrane integrity during rapid volume changes, the effects of two membrane trafficking inhibitors on the response of intact guard cells of Vicia faba to osmotic treatments were studied. Using confocal microscopy and epidermal peels, the relationship between the area of a medial paradermal guard-cell section and guard-cell volume was determined. This allowed estimates of guard-cell volume to be made from single paradermal confocal images, and therefore allowed rapid determination of volume as cells responded to osmotic treatments. Volume changes in control cells showed exponential kinetics, and it was possible to calculate an apparent value for guard-cell hydraulic conductivity from these kinetics. Wortmannin and cytochalasin D inhibited the rate of volume loss following a 0-1.5 MPa osmotic treatment. Cytochalasin D also inhibited volume increases following a change from 1.5 MPa to 0 MPa, but wortmannin had no effect. Previous studies showing that treatment with arabinanase inhibits changes in guard-cell volume in response to osmotic treatments were confirmed. However, pressure volume curves show that the effects of arabinanase and the cytochalasin D were not due to changes in cell wall elasticity. It is suggested that arabinanase, cytochalasin D, and wortmannin cause reductions in the hydraulic conductivity of the plasma membrane, possibly via gating of aquaporins. A possible role for aquaporins in co-ordinating volume changes with membrane trafficking is discussed.  (+info)

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