A novel role for the TIR domain in association with pathogen-derived elicitors

Plant innate immunity is mediated by Resistance (R) proteins, which bear a striking resemblance to animal molecules of similar function. Tobacco N is a TIR-NB-LRR R gene that confers resistance to Tobacco mosaic virus, specifically the p50 helicase domain. An intriguing question is how plant R proteins recognize the presence of pathogen-derived Avirulence (Avr) elicitor proteins. We have used biochemical cell fraction and immunoprecipitation in addition to confocal fluorescence microscopy of living tissue to examine the association between N and p50. Surprisingly, both N and p50 are cytoplasmic and nuclear proteins, and N's nuclear localization is required for its function. We also demonstrate an in planta association between N and p50. Further, we show that N's TIR domain is critical for this association, and indeed, it alone can associate with p50. Our results differ from current models for plant innate immunity that propose detection is mediated solely through the LRR domains of these molecules. The data we present support an intricate process of pathogen elicitor recognition by R proteins involving multiple subcellular compartments and the formation of multiple protein complexes. © 2007 Burch-Smith et al

High light and temperature reduce photosynthetic efficiency through different mechanisms in the C4 model Setaria viridis.

Funder: start-up funding from Donald Danforth Plant Science CenterC4 plants frequently experience high light and high temperature conditions in the field, which reduce growth and yield. However, the mechanisms underlying these stress responses in C4 plants have been under-explored, especially the coordination between mesophyll (M) and bundle sheath (BS) cells. We investigated how the C4 model plant Setaria viridis responded to a four-hour high light or high temperature treatment at photosynthetic, transcriptomic, and ultrastructural levels. Although we observed a comparable reduction of photosynthetic efficiency in high light or high temperature treated leaves, detailed analysis of multi-level responses revealed important differences in key pathways and M/BS specificity responding to high light and high temperature. We provide a systematic analysis of high light and high temperature responses in S. viridis, reveal different acclimation strategies to these two stresses in C4 plants, discover unique light/temperature responses in C4 plants in comparison to C3 plants, and identify potential targets to improve abiotic stress tolerance in C4 crops

Griseofulvin stabilizes microtubule dynamics, activates p53 and inhibits the proliferation of MCF-7 cells synergistically with vinblastine

<p>Abstract</p> <p>Background</p> <p>Griseofulvin, an antifungal drug, has recently been shown to inhibit proliferation of various types of cancer cells and to inhibit tumor growth in athymic mice. Due to its low toxicity, griseofulvin has drawn considerable attention for its potential use in cancer chemotherapy. This work aims to understand how griseofulvin suppresses microtubule dynamics in living cells and sought to elucidate the antimitotic and antiproliferative action of the drug.</p> <p>Methods</p> <p>The effects of griseofulvin on the dynamics of individual microtubules in live MCF-7 cells were measured by confocal microscopy. Immunofluorescence microscopy, western blotting and flow cytometry were used to analyze the effects of griseofulvin on spindle microtubule organization, cell cycle progression and apoptosis. Further, interactions of purified tubulin with griseofulvin were studied <it>in vitro </it>by spectrophotometry and spectrofluorimetry. Docking analysis was performed using autodock4 and LigandFit module of Discovery Studio 2.1.</p> <p>Results</p> <p>Griseofulvin strongly suppressed the dynamic instability of individual microtubules in live MCF-7 cells by reducing the rate and extent of the growing and shortening phases. At or near half-maximal proliferation inhibitory concentration, griseofulvin dampened the dynamicity of microtubules in MCF-7 cells without significantly disrupting the microtubule network. Griseofulvin-induced mitotic arrest was associated with several mitotic abnormalities like misaligned chromosomes, multipolar spindles, misegregated chromosomes resulting in cells containing fragmented nuclei. These fragmented nuclei were found to contain increased concentration of p53. Using both computational and experimental approaches, we provided evidence suggesting that griseofulvin binds to tubulin in two different sites; one site overlaps with the paclitaxel binding site while the second site is located at the αβ intra-dimer interface. In combination studies, griseofulvin and vinblastine were found to exert synergistic effects against MCF-7 cell proliferation.</p> <p>Conclusions</p> <p>The study provided evidence suggesting that griseofulvin shares its binding site in tubulin with paclitaxel and kinetically suppresses microtubule dynamics in a similar manner. The results revealed the antimitotic mechanism of action of griseofulvin and provided evidence suggesting that griseofulvin alone and/or in combination with vinblastine may have promising role in breast cancer chemotherapy.</p

Distinct colonization patterns and cDNA-AFLP transcriptome profiles in compatible and incompatible interactions between melon and different races of Fusarium oxysporum f. sp. melonis

Background: Fusarium oxysporum f. sp. melonis Snyd. & Hans. (FOM) causes Fusarium wilt, the most important infectious disease of melon (Cucumis melo L.). The four known races of this pathogen can be distinguished only by infection on appropriate cultivars. No molecular tools are available that can discriminate among the races, and the molecular basis of compatibility and disease progression are poorly understood. Resistance to races 1 and 2 is controlled by a single dominant gene, whereas only partial polygenic resistance to race 1,2 has been described. We carried out a large-scale cDNA-AFLP analysis to identify host genes potentially related to resistance and susceptibility as well as fungal genes associated with the infection process. At the same time, a systematic reisolation procedure on infected stems allowed us to monitor fungal colonization in compatible and incompatible host-pathogen combinations. Results: Melon plants (cv. Charentais Fom-2), which are susceptible to race 1,2 and resistant to race 1, were artificially infected with a race 1 strain of FOM or one of two race 1,2 w strains. Host colonization of stems was assessed at 1, 2, 4, 8, 14, 16, 18 and 21 days post inoculation (dpi), and the fungus was reisolated from infected plants. Markedly different colonization patterns were observed in compatible and incompatible host-pathogen combinations. Five time points from the symptomless early stage (2 dpi) to obvious wilting symptoms (21 dpi) were considered for cDNA-AFLP analysis. After successful sequencing of 627 transcript-derived fragments (TDFs) differentially expressed in infected plants, homology searching retrieved 305 melon transcripts, 195 FOM transcripts expressed in planta and 127 orphan TDFs. RNA samples from FOM colonies of the three strains grown in vitro were also included in the analysis to facilitate the detection of in planta-specific transcripts and to identify TDFs differentially expressed among races/strains. Conclusion: Our data suggest that resistance against FOM in melon involves only limited transcriptional changes, and that wilting symptoms could derive, at least partially, from an active plant response. We discuss the pathogen-derived transcripts expressed in planta during the infection process and potentially related to virulence functions, as well as transcripts that are differentially expressed between the two FOM races grown in vitro. These transcripts provide candidate sequences that can be further tested for their ability to distinguish between races. Sequence data from this article have been deposited in GenBank, Accession Numbers: HO867279-HO867981

Creating Nutrient Sustainability Indicators For Dairies Nationwide

This paper was presented at the 2018 Cornell Nutrition Conference. For more information, please visit ansci.cals.cornell.edu/CNC.Many industries have, in recent years, focused on development of Sustainability Performance Indicators, reflecting the desire by food processing companies to implement a profitable agricultural supply chain that protects the environment through adoption of sustainable practices. Such indicators should be fairly easy to derive, be verifiable, be meaningful, and responsive to change in management over time. With the nutrient mass balance assessment, we present a farmer-focused approach of a sustainability stamp for three key nutrients (nitrogen, phosphorus, and potassium). We will present 10 years of results for New York dairies that participated in the development of this approach, and compare this to developments in other states and internationally.Department of Animal Scienc

Blast Interfacial Complex, a novel in planta structure that accumulates effector proteins of rice blast fungus Magnaporthe oryzae

International audienceThe hemibiotrophic fungus Magnaporthe oryzae causes the devastating rice blast disease using specialized intracellular invasive hyphae (IH) that successively invade living plant cells. IH differentiate from thin filamentous primary hyphae that form immediately af ter the appressorial penetration peg breaches the exterior plant surface. At this point, IH become enclosed in a generally tight-fitting, plant-derived Extra-Invasive Hyphal Membrane (EIHM). Blast effector proteins are secreted from IH to manipulate host responses, but the route of their delive ry across the EIHM to reach the host cytoplasm is not understood. Here we show that fusion proteins between blast effectors, AVR-Pita and PWL, and enhanced green fluorescent protein (EGFP) are secreted by the fungus into a previously unrecognized structure, the Blast Interfacial Complex (BIC). Live cell imaging showed that fusion proteins first appeared in primary BICs at the growing tips of primary hyphae, in the previously reported EIHM caps. At the point where primary hyphae differentiated into bulbous IH, the fluorescent BICs moved to the sides of enlarging IH and remained fluorescent at these same locations as long as IH grew within the cell. When the fungus moved into neighboring cells, fluorescence accumulated in secondary BICs at the tips of filamentous IH that grew immediately after crossing the plan t cell wall. Again, the fluorescent tip structures moved to the side when the thin hyphae thickened into IH. BICs reside between the fungal cell wall and the EIHM in dynamic association with plant cytoplasm as IH begin to grow. Correlative light and electron microscopy showed that BICs contained complex lamellar membranes and vesicles. BIC localization requires the N-terminal signal peptide of these effector proteins. We hypothesize that BI Cs are involved in delivering fungal effectors proteins inside plant cells

Dexamethasone-Loaded Block Copolymer Nanoparticles Induce Leukemia Cell Death and Enhance Therapeutic Efficacy: A Novel Application in Pediatric Nanomedicine

Nanotechnology approaches have tremendous potential for enhancing treatment efficacy with lower doses of chemotherapeutics. Nanoparticle (NP)-based drug delivery approaches are poorly developed for childhood leukemia. Dexamethasone (Dex) is one of the most common chemotherapeutic drugs used in the treatment of childhood leukemia. In this study, we encapsulated Dex in polymeric NPs and validated their antileukemic potential in vitro and in vivo. NPs with an average diameter of 110 nm were assembled from an amphiphilic block copolymer of poly­(ethylene glycol) (PEG) and poly­(ε-caprolactone) (PCL) bearing pendant cyclic ketals (ECT2). The blank NPs were nontoxic to cultured cells in vitro and to mice in vivo. Encapsulation of Dex into the NPs (Dex-NP) did not compromise the bioactivity of the drug. Dex-NPs induced glucocorticoid phosphorylation and showed cytotoxicity similar to the free Dex in leukemic cells. Studies using NPs labeled with fluorescent dyes revealed leukemic cell surface binding and internalization. In vivo biodistribution studies showed NP accumulation in the liver and spleen with subsequent clearance of the particles with time. In a preclinical model of leukemia, Dex-NPs significantly improved the quality of life and survival of mice as compared to the free drug. To our knowledge, this is the first report showing the efficacy of polymeric NPs to deliver Dex to potentially treat childhood leukemia and reveals that low doses of Dex should be sufficient for inducing cell death and improving survival