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71.
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Recent findings in our laboratory suggested that in citrus cells the salt induction of phospholipid hydroperoxide glutathione
peroxidase, an enzyme active in cellular antioxidant defense, is mediated by the accumulation of hydroperoxides. Production
of hydroperoxides occurs as a result of non-enzymatic auto-oxidation or via the action of lipoxygenases (LOXs). In an attempt
to resolve the role of LOX activity in the accumulation of peroxides we analyzed the expression of this protein under stress
conditions and in cells of Citrus sinensis L. differing in sensitivity to salt. Lipoxygenase expression was induced very rapidly only in the salt-tolerant cells and
in a transient manner. The induction was specific to salt stress and did not occur with other osmotic-stress-inducing agents,
such as polyethylene glycol or mannitol, or under hot or cold conditions, or in the presence of abscisic acid. The induction
was eliminated by the antioxidants dithiothreitol and kaempferol, thus once more establishing a correlation between salt and
oxidative stresses. Analyses of both in vitro and in vivo products of LOX revealed a specific 9-LOX activity, and a very fast
reduction of the hydroperoxides to the corresponding hydroxy derivatives. This suggests that one of the metabolites further
downstream in the reductase pathway may play a key role in triggering defense responses against salt stress.
Received: 3 February 2000 / Accepted: 13 June 2000 相似文献
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Anna Kogan Garik Y Gdalevsky Rivka Cohen-Luria Yehuda Goldgur Robert S Phillips Abraham H Parola Orna Almog 《BMC structural biology》2009,9(1):65-12
Background
Oligomeric enzymes can undergo a reversible loss of activity at low temperatures. One such enzyme is tryptophanase (Trpase) from Escherichia coli. Trpase is a pyridoxal phosphate (PLP)-dependent tetrameric enzyme with a Mw of 210 kD. PLP is covalently bound through an enamine bond to Lys270 at the active site. The incubation of holo E. coli Trpases at 2°C for 20 h results in breaking this enamine bond and PLP release, as well as a reversible loss of activity and dissociation into dimers. This sequence of events is termed cold lability and its understanding bears relevance to protein stability and shelf life. 相似文献76.
Serratia liquefaciens provided better protection of carnations from infection by Fusarium oxysporum f. sp. dianthi, than did Hafnia alvei. The protection occurred when the bacterial isolates were applied to the cuttings before rooting, but not when applied to the root system of rooted cuttings. S. liquefaciens was recovered from carnation stem segments along the stem up to the top after 60 days but after 120 days they were recovered only up to 2.5cm. Zones of inhibition of Fusarium-conidia sprayed on agar plates previously incubated with stem segments appeared around the bacterial colonies of S. liquifaciens after additional incubation for 24—48h. 相似文献
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Stefan Zdraljevic Drew Wagner Kevin Cheng Laura Ruohonen Jussi J?ntti Merja Penttil? Orna Resnekov C. Gustavo Pesce 《Applied and environmental microbiology》2013,79(24):7569-7582
Organic acids derived from engineered microbes can replace fossil-derived chemicals in many applications. Fungal hosts are preferred for organic acid production because they tolerate lignocellulosic hydrolysates and low pH, allowing economic production and recovery of the free acid. However, cell death caused by cytosolic acidification constrains productivity. Cytosolic acidification affects cells asynchronously, suggesting that there is an underlying cell-to-cell heterogeneity in acid productivity and/or in resistance to toxicity. We used fluorescence microscopy to investigate the relationship between enzyme concentration, cytosolic pH, and viability at the single-cell level in Saccharomyces cerevisiae engineered to synthesize xylonic acid. We found that cultures producing xylonic acid accumulate cells with cytosolic pH below 5 (referred to here as “acidified”). Using live-cell time courses, we found that the probability of acidification was related to the initial levels of xylose dehydrogenase and sharply increased from 0.2 to 0.8 with just a 60% increase in enzyme abundance (Hill coefficient, >6). This “switch-like” relationship likely results from an enzyme level threshold above which the produced acid overwhelms the cell''s pH buffering capacity. Consistent with this hypothesis, we showed that expression of xylose dehydrogenase from a chromosomal locus yields ∼20 times fewer acidified cells and ∼2-fold more xylonic acid relative to expression of the enzyme from a plasmid with variable copy number. These results suggest that strategies that further reduce cell-to-cell heterogeneity in enzyme levels could result in additional gains in xylonic acid productivity. Our results demonstrate a generalizable approach that takes advantage of the cell-to-cell variation of a clonal population to uncover causal relationships in the toxicity of engineered pathways. 相似文献
79.
Edith Hochhauser Orna Avlas Reut Fallach Larissa Bachmetov Romy Zemel Orit Pappo Asher Shainberg Ziv Ben Ari 《PloS one》2013,8(8)
Background
Toll-like receptors (TLRs) are expressed in immune cells and hepatocytes. We examined whether hepatic Toll-like receptor 4 (TLR4) is involved in the acute hepatic injury caused by the administration of lipopolysaccharide (LPS) (septic shock model).Methods
Wild type (WT), TLR4-deficient and chimera mice underwent myeloablative bone marrow transplantation to dissociate between TLR4 expression in the liver or in the immune-hematopoietic system. Mice were injected with LPS and sacrificed 4 hours later.Results
Compared to TLR4 deficient mice, WT mice challenged with LPS displayed increased serum liver enzymes and hepatic cellular inflammatory infiltrate together with increased serum and hepatic levels of interleukin 1β (IL-1β), tumor necrosis factor α (TNFα) ,Up-regulation of hepatic mRNA encoding TLR4, IκB and c-jun expressions. TLR4 mutant mice transplanted with WT bone marrow were more protected than WT chimeric mice bearing TLR4 mutant hemopoietic cells from LPS, as seen by IL-1β and TNFα levels. We then used hepatocytes (Huh7) and macrophages from monocytic cell lines to detect TLR mRNA expression. Macrophages expressed a significantly higher level of TLR4 mRNA and TLR2 (more than 3000- and 8000-fold respectively) compared with the hepatocyte cell line. LPS administration induced TLR4 activation in a hepatocyte cell line in a dose dependent manner while TLR2 mRNA hardly changed.Conclusions
These results suggest that TLR4 activation of hepatocytes participate in the immediate response to LPS induced hepatic injury. However, in this response, the contribution of TLR4 on bone marrow derived cells is more significant than those of the hepatocytes. The absence of the TLR4 gene plays a pivotal role in reducing hepatic LPS induced injury. 相似文献80.
Differentiation-Induced Internal Translation of c-sis mRNA: Analysis of the cis Elements and Their Differentiation-Linked Binding to the hnRNP C Protein
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Osnat Sella Gabi Gerlitz Shu-Yun Le Orna Elroy-Stein 《Molecular and cellular biology》1999,19(8):5429-5440
In previous reports we showed that the long 5' untranslated region (5' UTR) of c-sis, the gene encoding the B chain of platelet-derived growth factor, has translational modulating activity due to its differentiation-activated internal ribosomal entry site (D-IRES). Here we show that the 5' UTR contains three regions with a computer-predicted Y-shaped structure upstream of an AUG codon, each of which can confer some degree of internal translation by itself. In nondifferentiated cells, the entire 5' UTR is required for maximal basal IRES activity. The elements required for the differentiation-sensing ability (i.e., D-IRES) were mapped to a 630-nucleotide fragment within the central portion of the 5' UTR. Even though the region responsible for IRES activation is smaller, the full-length 5' UTR is capable of mediating the maximal translation efficiency in differentiated cells, since only the entire 5' UTR is able to confer the maximal basal IRES activity. Interestingly, a 43-kDa protein, identified as hnRNP C, binds in a differentiation-induced manner to the differentiation-sensing region. Using UV cross-linking experiments, we show that while hnRNP C is mainly a nuclear protein, its binding activity to the D-IRES is mostly nuclear in nondifferentiated cells, whereas in differentiated cells such binding activity is associated with the ribosomal fraction. Since the c-sis 5' UTR is a translational modulator in response to cellular changes, it seems that the large number of cross-talking structural entities and the interactions with regulated trans-acting factors are important for the strength of modulation in response to cellular changes. These characteristics may constitute the major difference between strong IRESs, such as those seen in some viruses, and IRESs that serve as translational modulators in response to developmental signals, such as that of c-sis. 相似文献