Ultrastructural distribution of a MAP kinase and transcripts in quiescent and cycling plant cells and pollen grains.
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Mitogen-activated protein kinases (MAPKs) are components of a kinase module that plays a central role in the transduction of diverse extracellular stimuli, including mitogens, specific differentiation and developmental signals and stress treatments. This shows that reversible protein phosphorylation cascades play a pivotal role in signal transduction in animal cells and yeast, particularly the entry into mitosis of arrested cells. Homologues of MAPKs have been found and cloned in various plant species, but there have been no data about their in situ localization at the subcellular level and their expression in plant cells so far. In the present paper we report the first data on the ultrastructural in situ localization of MAPK and their mRNAs in various plant cells. Proliferating and quiescent meristematic plant cells were studied to evaluate whether changes in MAPK presence, distribution and expression accompany the entry into proliferation of dormant cells. Moreover, MAPK localization was analyzed in vacuolate microspores. Polyclonal antibodies against the deduced MAPK from the tobacco Ntf6 clone were able to recognize homologue epitopes by immunocytochemical techniques in the cell types studied. The pattern of protein distribution is similar in all the cases studied: it is localized in the cytoplasm and in the nucleus, mainly in the interchromatin region. The quantitative study of the density showed that MAPK labelling is more abundant in cycling than in quiescent cells, also suggesting that, in plants, MAPK pathways might play a role in cell proliferation. RNA probes for conserved regions of the catalytic domain of plant MAPK homologue genes were used to study MAPK expression in those plant cells. In situ hybridization (ISH) showed the presence of MAPK transcripts in the three plant cell types studied, but levels were very low in quiescent cells compared to those in cycling cells. The quantification of labelling density of ISH signals strongly suggests a higher level of MAPK expression in proliferating cells, but also some basal messenger presence and/or expression in the quiescent ones. Immunogold and ISH results show the presence and distribution of MAPK proteins and mRNAs in vacuolate microspores. This represents a very dynamic stage during pollen development in which the cell nucleus is being prepared for an asymmetrical mitotic division, giving rise to both the generative and the vegetative nuclei of the bicellular pollen grain. Taken together, the data indicate a role played by MAPK in the re-entry into proliferation in plant cells. (+info)
Photomophogenesis: Phytochrome takes a partner!
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How light signals are transduced by phytochromes is still poorly understood. Recent studies have provided evidence that a PAS domain protein, PIF3, physically interacts with phytochromes, plays a role in phytochrome signal transduction and might be a component of a novel signalling pathway in plants. (+info)
Reduced glutathione accelerates the oxidative damage produced by sodium n-propylthiosulfate, one of the causative agents of onion-induced hemolytic anemia in dogs.
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The oxidative effects of sodium n-propylthiosulfate, one of the causative agents of onion-induced hemolytic anemia in dogs, were investigated in vitro using three types of canine erythrocytes, which are differentiated by the concentration of reduced glutathione and the composition of intracellular cations. After incubation with sodium n-propylthiosulfate, the methemoglobin concentration and Heinz body count in all three types of erythrocytes increased and a decrease in the erythrocyte reduced glutathione concentration was then observed. The erythrocytes containing high concentrations of potassium and reduced glutathione (approximately five times the normal values) were more susceptible to oxidative damage by sodium n-propylthiosulfate than were the normal canine erythrocytes. The susceptibility of the erythrocytes containing high potassium and normal reduced glutathione concentrations was intermediate between those of erythrocytes containing high concentrations of potassium and reduced glutathione and normal canine erythrocytes. In addition, the depletion of erythrocyte reduced glutathione by 1-chloro-2, 4-dinitrobenzene resulted in a marked decrease in the oxidative injury induced by sodium n-propylthiosulfate in erythrocytes containing high concentrations of potassium and reduced glutathione. The generation of superoxide in erythrocytes containing high concentrations of potassium and reduced glutathione was 4.1 times higher than that in normal canine erythrocytes when the cells were incubated with sodium n-propylthiosulfate. These observations indicate that erythrocyte reduced glutathione, which is known as an antioxidant, accelerates the oxidative damage produced by sodium n-propylthiosulfate. (+info)
Rumen bacteria are involved in the onset of onion-induced hemolytic anemia in sheep.
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The mechanism of onion-induced hemolytic anemia in ruminants was investigated. The ether-extract obtained from the mixture of rumen fluid and onion juice incubated at 38.5 degrees C for 9 hr induced oxidative damage in sheep erythrocytes in vitro, indicating the production of certain oxidants in the mixture. The increase of the oxidative effect in the mixture was inhibited completely by the removal of rumen microorganisms and partly by treatment with antibiotics and by oxygen gas. The sheep fed onions (50 g/kg body weight/day) for 15 days developed more severe Heinz body hemolytic anemia than did the sheep fed the equivalent amount of onions with 5 g/day ampicillin sodium salt. The results indicated that certain rumen bacteria appear to be involved in the onset of onion-induced hemolytic anemia in sheep. (+info)
Evidence for functional conservation of a mammalian homologue of the light-responsive plant protein COP1.
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Identified in Arabidopsis as a repressor of light-regulated development, the COP1 (constitutively photomorphogenic 1) protein is characterized by a RING-finger motif and a WD40 repeat domain [1]. The subcellular localization of COP1 is light-dependent. COP1 acts within the nucleus to repress photomorphogenic development, but light inactivates COP1 and diminishes its nuclear abundance [2]. Here, we report the identification of a mammalian COP1 homologue that contains all the structural features present in Arabidopsis COP1 (AtCOP1). When expressed in plant cells, a fusion protein comprising mammalian COP1 and beta-glucuronidase (GUS) responded to light by changing its subcellular localization pattern in a manner similar to AtCOP1. Whereas the mammalian COP1 was unable to rescue the defects of Arabidopsis cop1 mutants, expression of the amino-terminal half of mammalian COP1 in Arabidopsis interfered with endogenous COP1 function, resulting in a hyperphotomorphogenic phenotype. Therefore, the regulatory modules in COP1 proteins that are responsible for the signal-dependent subcellular localization are functionally conserved between higher plants and mammals, suggesting that mammalian COP1 may share a common mode of action with its plant counterpart in regulating development and cellular signaling. (+info)
Specific interactions with TBP and TFIIB in vitro suggest that 14-3-3 proteins may participate in the regulation of transcription when part of a DNA binding complex.
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The 14-3-3 family of multifunctional proteins is highly conserved among animals, plants, and yeast. Several studies have shown that these proteins are associated with a G-box DNA binding complex and are present in the nucleus in several plant and animal species. In this study, 14-3-3 proteins are shown to bind the TATA box binding protein (TBP), transcription factor IIB (TFIIB), and the human TBP-associated factor hTAF(II)32 in vitro but not hTAF(II)55. The interactions with TBP and TFIIB were highly specific, requiring amino acid residues in the box 1 domain of the 14-3-3 protein. These interactions do not require formation of the 14-3-3 dimer and are not dependent on known 14-3-3 recognition motifs containing phosphoserine. The 14-3-3-TFIIB interaction appears to occur within the same domain of TFIIB that binds the human herpes simplex virus transcriptional activator VP16, because VP16 and 14-3-3 were able to compete for interaction with TFIIB in vitro. In a plant transient expression system, 14-3-3 was able to activate GAL4-dependent beta-glucuronidase reporter gene expression at low levels when translationally fused with the GAL4 DNA binding domain. The in vitro binding with general transcription factors TBP and TFIIB together with its nuclear location provide evidence supporting a role for 14-3-3 proteins as transcriptional activators or coactivators when part of a DNA binding complex. (+info)
An Arabidopsis cDNA encoding a DNA-binding protein that is highly similar to the DEAH family of RNA/DNA helicase genes.
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A cDNA encoding a putative RNA and/or DNA helicase has been isolated from Arabidopsis thaliana cDNA libraries. The cloned cDNA is 5166 bases long, and its largest open reading frame encodes 1538 amino acids. The central region of the predicted protein is homologous to a group of nucleic acid helicases from the DEAD/H family. However, the N- and C-terminal regions of the Arabidopsis cDNA product are distinct from these animal DEIH proteins. We have found that the C-terminal region contains three characteristic sequences: (i) two DNA-binding segments that form a probe helix (PH) involved in DNA recognition; (ii) an SV40-type nuclear localization signal; and (iii) 11 novel tandem-repeat sequences each consisting of about 28 amino acids. We have designated this cDNA as NIH (nuclear DEIH-boxhelicase). Functional character-ization of a recombinant fusion product containing the repeated region indicates that NIH may form homodimers, and that this is the active form in solution. Based on this information and the observation that the sequence homology is limited to the DEAH regions, we conclude that the biological roles of the plant helicase NIH differ from those of the animal DEIH family. (+info)
Fungal corneal ulcers of onion harvesters in southern Taiwan.
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Fungal corneal ulcers related to agriculture has been reported throughout the world, especially in tropical areas. Most of them were sporadic and had histories of ocular trauma or use of topical corticosteroids and topical antibiotics. Five onion harvesters had fungal corneal ulcers during the same harvest period in Southern Taiwan. The authors think that this is the first report of a group occurrence relating to agricultural workers. Although all of the patients improved after medical and surgical management, their vision was greatly decreased. It is suggested that the tropical climate, the harvest procedure, the characteristic monsoon, and lack of eye protection were involved. Therefore, the importance of the eye protection, hygiene education, and improving medical care to reduce the occurrence of fungal corneal ulcer in agriculture workers must be emphasised. (+info)