Project description:Background: Polycyclic aromatic hydrocarbons (PAHs) are toxic, widely-distributed, environmentally persistent, and carcinogenic byproducts of carbon-based fuel combustion. Previously, plant studies have shown that PAHs induce oxidative stress, reduce growth, and cause leaf deformation as well as tissue necrosis. To understand the transcriptional changes that occur during these processes, we performed microarray experiments on Arabidopsis thaliana L. under phenanthrene treatment, and compared the results to published Arabidopsis microarray data representing a variety of stress and hormone treatments. In addition, to probe hormonal aspects of PAH stress, we assayed transgenic ethylene-inducible reporter plants as well as ethylene pathway mutants under phenanthrene treatment. Results: Microarray results revealed numerous perturbations in signaling and metabolic pathways that regulate reactive oxygen species (ROS) and responses related to pathogen defense. A number of glutathione S-transferases that may tag xenobiotics for transport to the vacuole were upregulated. Comparative microarray analyses indicated that the phenanthrene response was closely related to other ROS conditions, including pathogen defense conditions. The ethylene-inducible transgenic reporters were activated by phenanthrene. Mutant experiments showed that PAH inhibits growth through an ethylene-independent pathway, as PAH-treated ethylene-insensitive etr1-4 mutants exhibited a greater growth reduction than WT. Further, phenanthrene-treated, constitutive ethylene signaling mutants had longer roots than the untreated control plants, indicating that the PAH inhibits parts of the ethylene signaling pathway. Conclusions: This study identified major physiological systems that participate in the PAH-induced stress response in Arabidopsis. At the transcriptional level, the results identify specific gene targets that will be valuable in finding lead compounds and engineering increased tolerance. Collectively, the results open a number of new avenues for researching and improving plant resilience and PAH phytoremediation.

Project description:Background: Polycyclic aromatic hydrocarbons (PAHs) are toxic, widely-distributed, environmentally persistent, and carcinogenic byproducts of carbon-based fuel combustion. Previously, plant studies have shown that PAHs induce oxidative stress, reduce growth, and cause leaf deformation as well as tissue necrosis. To understand the transcriptional changes that occur during these processes, we performed microarray experiments on Arabidopsis thaliana L. under phenanthrene treatment, and compared the results to published Arabidopsis microarray data representing a variety of stress and hormone treatments. In addition, to probe hormonal aspects of PAH stress, we assayed transgenic ethylene-inducible reporter plants as well as ethylene pathway mutants under phenanthrene treatment. Results: Microarray results revealed numerous perturbations in signaling and metabolic pathways that regulate reactive oxygen species (ROS) and responses related to pathogen defense. A number of glutathione S-transferases that may tag xenobiotics for transport to the vacuole were upregulated. Comparative microarray analyses indicated that the phenanthrene response was closely related to other ROS conditions, including pathogen defense conditions. The ethylene-inducible transgenic reporters were activated by phenanthrene. Mutant experiments showed that PAH inhibits growth through an ethylene-independent pathway, as PAH-treated ethylene-insensitive etr1-4 mutants exhibited a greater growth reduction than WT. Further, phenanthrene-treated, constitutive ethylene signaling mutants had longer roots than the untreated control plants, indicating that the PAH inhibits parts of the ethylene signaling pathway. Conclusions: This study identified major physiological systems that participate in the PAH-induced stress response in Arabidopsis. At the transcriptional level, the results identify specific gene targets that will be valuable in finding lead compounds and engineering increased tolerance. Collectively, the results open a number of new avenues for researching and improving plant resilience and PAH phytoremediation. Arabidopsis thaliana (ecotype Columbia) plants were long-day grown with +/- 0.25 mM phenanthrene in sterile plates at 23C for 21d before harvest. At least 20 plants were pooled prior to each mRNA extraction.

Project description:Ethylene induced hyponastic growth in Arabidopsis thaliana F.F. Millenaar L.A.C.J. Voesenek and A.J.M. Peeters Our aim is to identify genes involved in the ethylene induced hyponastic growth. Upon submergence some plant species like Rumex palustris changes its leaf angle (hyponastic growth) and shows enhanced petiole elongation to reach the water surface. In Rumex palustris the hyponastic growth is initiated by an increased concentration of ethylene due to physical entrapment and ongoing ethylene biosynthesis. A proteomics, genomics and genetical approach to improve our understanding of above described flooding-induced responses are not feasible in Rumex palustris since genomic information about this species is limited. However it is possible to use the model plant Arabidopsis thaliana as a tool in flooding research. Natural accessions (Be0 Col Cvi Kas Ler Nd Rld Shah and Ws) show considerable genetic variation in hyponastic growth upon exposure to ethylene Col exhibiting the largest effect (maximum rate after 3 hours) and Ler no effect whatsoever. Using a computer controlled digital camera the hyponastic growth is measured in great detail. Next to ethylene addition also a transfer to low light causes hyponastic growth. This seems to be an ethylene independent pathway because etr1 and ctr1 showed hyponastic growth after transfer to low light. Ethylene and low light showed additive effects in Col. It is likely that ethylene induces more changes in gene expression than only the ones involved in hyponastic growth. By subtracting changes in the Ler expression profile from changes in the Col expression profile we expect to find why Col and Ler respond differently on ethylene by finding specific ethylene induced genes that are involved in hyponastic growth. The expression profile of Col following transfer to low light will be substracted from Col following ethylene addition to distinguish between genes that are involved in hyponastic growth but are not specific for ethylene induced hyponastic growth. There are strong indications in Rumex palustris that other hormones i.e. auxin ABAand GA are involved in the ethylene induced hyponastic growth. Currently mutants in ethylene auxin and ABA biosynthesis and/or signal transduction are screened for hyponastic growth. Preliminary results showed that also in Arabidopsis these other hormones are involved in ethylene induced hyponastic growth. Beside the mutant approach we also started a proteomics and a PCR based differential screen approach. Together with the proposed transcriptome analysis we hope to find new genes involved in ethylene induced hyponastic growth.

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Arabidopsis thaliana

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