Environmental stress responses in Lactobacillus: a review

Environmental stress responses in Lactobacillus, which have been investigated mainly by proteomics approaches, are reviewed. The physiological and molecular mechanisms of responses to heat, cold, acid, osmotic, oxygen, high pressure and starvation stresses are described. Specific examples of the repercussions of these effects in food processing are given. Molecular mechanisms of stress responses in lactobacilli and other bacteria are compared.     

Interaction of Osmotic Stress, Temperature, and Abscisic Acid in the Regulation of Gene Expression in Arabidopsis

The impact of simultaneous environmental stresses on plants and how they respond to combined stresses compared with single stresses is largely unclear. By using a transgene (RD29A-LUC) consisting of the firefly luciferase coding sequence (LUC) driven by the stress-responsive RD29A promoter, we investigated the interactive effects of temperature, osmotic stress, and the phytohormone abscisic acid (ABA) in the regulation of gene expression in Arabidopsis seedlings. Results indicated that both positive and negative interactions exist among the studied stress factors in regulating gene expression. At a normal growth temperature (22°C), osmotic stress and ABA act synergistically to induce the transgene expression. Low temperature inhibits the response to osmotic stress or to combined treatment of osmotic stress and ABA, whereas low temperature and ABA treatments are additive in inducing transgene expression. Although high temperature alone does not activate the transgene, it significantly amplifies the effects of ABA and osmotic stress. The effect of multiple stresses in the regulation of RD29A-LUC expression in signal transduction mutants was also studied. The results are discussed in the context of cold and osmotic stress signal transduction pathways.     

High Throughput Screening of Signal Transduction Mutants With Luciferase Imaging

A detailed procedure for high throughput genetic screening of hormone and environmental stress signal transduction mutants of Arabidopsis thaliana is described. The screen was carried out with mutagenized plants expressing the firefly luciferase reporter under control of a cold, osmotic stress, and absciscic acid responsive promoter. A thermoelectrically cooled CCD camera was used to detect luminescence emitted by the plants in response to stresses or ABA. Advantages of the screening procedure include high throughput, capability to identify low as well as high expression mutants and employment of a highly sensitive but affordable imaging system and software. This procedure can be used to study complex signal transduction networks in higher plants.     

A proteomics approach for identifying osmotic-stress-related proteins in rice

Osmotic stress can endanger the survival of plants. To investigate the mechanisms of how plants respond to osmotic stress, rice protein profiles from mannitol-treated plants, were monitored using a proteomics approach. Two-week-old rice seedlings were treated with 400mM mannitol for 48h. After separation of proteins from the basal part of leaf sheaths by two-dimensional polyacrylamide gel electrophoresis, 327 proteins were detected. The levels of 12 proteins increased and the levels of three proteins decreased with increasing concentration or duration, of mannitol treatment. Levels of a heat shock protein and a dnaK-type molecular chaperone were reduced under osmotic, cold, salt and drought stresses, and ABA treatment, whereas a 26S proteasome regulatory subunit was found to be responsive only to osmotic stress. Furthermore, proteins whose accumulation was sensitive to osmotic stress are present in an osmotic-tolerant cultivar. These results indicate that specific proteins expressed in the basal part of rice leaf sheaths show a coordinated response to cope with osmotic stress.     

Activation of the HOG pathway upon cold stress in Saccharomyces cerevisiae

When Saccharomyces cerevisiae cells are exposed to hyper-osmotic stress, the high-osmolarity glycerol response (HOG) pathway is activated to induce osmotic responses. The HOG pathway consists of two upstream osmosensing branches, the SLN1 and SHO1 branches, and a downstream MAP kinase cascade. Although the mechanisms by which these upstream branches transmit signals to the MAP kinase cascade are well understood, the mechanisms by which they sense and respond to osmotic changes are elusive. Here we show that the HOG pathway is activated in an SLN1 branch-dependent manner when cells are exposed to cold stress (0 degrees C treatment). Dimethyl sulfoxide (DMSO) treatment, which rigidifies the cell membrane, also activates the HOG pathway in both SLN1 branch- and SHO1 branch-dependent manners. Moreover, cold stress, as well as hyper-osmotic stress, exhibits a synergistic effect with DMSO treatment on HOG pathway activation. On the other hand, ethanol treatment, which fluidizes the cell membrane, partially represses the cold stress-induced HOG pathway activation. Our results suggest that both osmosensing branches respond to the rigidification of the cell membrane to activate the HOG pathway.     

Rapid determination of the damage to photosynthesis caused by salt and osmotic stresses using delayed fluorescence of chloroplasts

Chloroplasts are one of the most susceptible systems to salt and osmotic stresses. Based on quantitative measurements of delayed fluorescence (DF) of the chloroplasts, we have investigated the damage to photosynthesis caused by these two kinds of stresses in Arabidopsis seedlings by using a custom-built multi-channel biosensor. Results showed that the DF intensity and net photosynthesis rate (Pn) decreased in a similar way with increasing NaCl or sorbitol concentration. Incubation of the seedlings in 200 mM NaCl induced a rapid and reversible decline and subsequent slow and irreversible loss in both the DF intensity and Pn. The rapid decline was dominantly related to osmotic stress, whereas the slow declines in the DF intensity and Pn were specific to ionic stress and could be reversed to a similar extent by a Na+-channel blocker. The DF intensity and Pn also exhibited a similar response to irradiation light under NaCl or sorbitol stress. All results indicated that the DF intensity correlated well with Pn under salt and osmotic stresses. We thus conclude that DF is an excellent marker for detecting the damage to photosynthesis caused by these two stresses. The mechanism of the correlation between the DF intensity and Pn under salt and osmotic stresses was also analyzed in theory and investigated with experiments by measuring intercellular CO2 concetration (Ci), stomatal conductance (Gs), chlorophyll fluorescence parameter, and chlorophyll content. This proposed DF technique holds the potential to be a useful means for analyzing the dynamics of salt and osmotic stresses in vivo and elucidating the mechanism by which plants respond to stress.     

The Arabidopsis LOS5/ABA3 locus encodes a molybdenum cofactor sulfurase and modulates cold stress- and osmotic stress-responsive gene expression

To understand low temperature and osmotic stress signaling in plants, we isolated and characterized two allelic Arabidopsis mutants, los5-1 and los5-2, which are impaired in gene induction by cold and osmotic stresses. Expression of RD29A-LUC (the firefly luciferase reporter gene under the control of the stress-responsive RD29A promoter) in response to cold and salt/drought is reduced in the los5 mutants, but the response to abscisic acid (ABA) remains unaltered. RNA gel blot analysis indicates that the los5 mutation reduces the induction of several stress-responsive genes by cold and severely diminishes or even completely blocks the induction of RD29A, COR15, COR47, RD22, and P5CS by osmotic stresses. los5 mutant plants are compromised in their tolerance to freezing, salt, or drought stress. los5 plants are ABA deficient, as indicated by increased transpirational water loss and reduced accumulation of ABA under drought stress in the mutant. A comparison with another ABA-deficient mutant, aba1, reveals that the impaired low-temperature gene regulation is specific to the los5 mutation. Genetic tests suggest that los5 is allelic to aba3. Map-based cloning reveals that LOS5/ABA3 encodes a molybdenum cofactor (MoCo) sulfurase. MoCo sulfurase catalyzes the generation of the sulfurylated form of MoCo, a cofactor required by aldehyde oxidase that functions in the last step of ABA biosynthesis in plants. The LOS5/ABA3 gene is expressed ubiquitously in different plant parts, and the expression level increases in response to drought, salt, or ABA treatment. Our results show that LOS5/ABA3 is a key regulator of ABA biosynthesis, stress-responsive gene expression, and stress tolerance.     

Differential regulation of microRNAs in response to osmotic,salt and cold stresses in wheat

MicroRNAs (miRNAs) are tiny non-coding regulatory molecules that modulate plant’s gene expression either by cleaving or repressing their mRNA targets. To unravel the plant actions in response to various environmental factors, identification of stress related miRNAs is essential. For understanding the regulatory behaviour of various abiotic stresses and miRNAs in wheat genotype C-306, we examined expression profile of selected conserved miRNAs viz. miR159, miR164, miR168, miR172, miR393, miR397, miR529 and miR1029 tangled in adapting osmotic, salt and cold stresses. The investigation revealed that two miRNAs (miR168, miR397) were down-regulated and miR172 was up-regulated under all the stress conditions. However, miR164 and miR1029 were up-regulated under cold and osmotic stresses in contrast to salt stress. miR529 responded to cold alone and does not change under osmotic and salt stress. miR393 showed up-regulation under osmotic and salt, and down-regulation under cold stress indicating auxin based differential cold response. Variation in expression level of studied miRNAs in presence of target genes delivers a likely elucidation of miRNAs based abiotic stress regulation. In addition, we reported new stress induced miRNAs Ta-miR855 using computational approach. Results revealed first documentation that miR855 is regulated by salinity stress in wheat. These findings indicate that diverse miRNAs were responsive to osmotic, salt and cold stress and could function in wheat response to abiotic stresses.     

环境胁迫下HOG-MAPK途径对真菌毒素形成的调控机制北大核心CSCD

真菌为了适应在生长侵染食品、饲料等农产品的过程中所面临的各种环境胁迫的考验,包括热胁迫、氧化胁迫、渗透压胁迫、紫外胁迫等,进化出一套高渗透性甘油促分裂原活化蛋白激酶(high osmolarity glycerol mitogen-activated protein kinase,HOG-MAPK)途径。该途径对真菌的生长发育、真菌毒素的产生和致病性都具有重要影响。HOG-MAPK途径共有两个分支,其中SLN1分支相比另一分支(SHO1分支)具有较为敏感的渗透压胁迫感应能力,能在高渗压和高盐浓度下进行渗透压胁迫反应。SHO1分支参与多种信号感应传导,比如氧化胁迫、热胁迫等。本文综述了真菌HOG-MAPK途径中关键基因sln1、sho1、ste11、ssk2、pbs2和hog1在应对渗透压胁迫、氧化胁迫等不同环境胁迫时所发挥的功能,说明HOG-MAPK途径可以响应多种环境信号,并参与调控黄曲霉、赭曲霉等致病真菌的生长和黄曲霉毒素(aflatoxin)、赭曲霉毒素(ochratoxin)等真菌毒素的产生。在不同环境胁迫下,HOG-MAPK途径对真菌毒素调控机制的研究可为食品和饲料等农产品真菌毒素的防控提供理论基础和指导方向。     

Cell-type-specific calcium responses to drought, salt and cold in the Arabidopsis root

Little is known about the signalling processes involved in the response of roots to abiotic stresses. The Arabidopsis root is a model system of root anatomy with a simple architecture and is amenable to genetic manipulation. Although it is known that the root responds to cold, drought and salt stress with increases in cytoplasmic free calcium, there is currently no information about the role(s) of the functionally diverse cell types that comprise the root. Transgenic Arabidopsis with enhancer-trapped GAL4 expression in specific cell types was used to target the calcium reporting protein, aequorin, fused to a modified yellow fluorescent protein (YFP). The luminescence output of targeted aequorin enabled in vivo measurement of changes in cytosolic free calcium concentrations ([Ca2+]cyt) in specific cell types during acute cold, osmotic and salt stresses. In response to an acute cold stress, all cell types tested as well as plants constitutively expressing aequorin displayed rapid [Ca2+]cyt peaks. However, there were significant quantitative differences between different cell types in terms of their response to cold stress, osmotic stress (440 mM mannitol) and salt stress (220 mM NaCl), implying specific roles for certain cell types in the detection and/or response to these stimuli. In response to osmotic and salt stress, the endodermis and pericycle displayed prolonged oscillations in cytosolic calcium that were distinct from the responses of the other cell types tested. Targeted expression of aequorin circumvented the technical difficulties involved in fluorescent dye injection as well as the lack of cell specificity of constitutively expressed aequorin, and revealed a new level of complexity in root calcium signalling.     

Differential Responses of Nitrogen-Fixing Cyanobacteria to Salinity and Osmotic Stresses

Two nitrogen-fixing Anabaena strains were found to be differentially tolerant to salinity and osmotic stresses. Anabaena torulosa, a brackish-water, salt-tolerant strain, was relatively osmosensitive. Anabaena sp. strain L-31, a freshwater, salt-sensitive strain, on the other hand, displayed significant osmotolerance. Salinity and osmotic stresses affected nitrogenase activity differently. Nitrogen fixation in both of the strains was severely inhibited by the ionic, but not by the osmotic, component of salinity stress. Such differential sensitivity of diazotrophy to salinity-osmotic stresses was observed irrespective of the inherent tolerance of the two strains to salt-osmotic stress. Exogenously added ammonium conferred significant protection against salinity stress but was ineffective against osmotic stress. Salinity and osmotic stresses also affected stress-induced gene expression differently. Synthesis of several proteins was repressed by salinity stress but not by equivalent or higher osmotic stress. Salinity and osmotic stresses induced many common proteins. In addition, unique salt stress- or osmotic stress-specific proteins were also induced in both strains, indicating differential regulation of protein synthesis by the two stresses. These data show that cyanobacterial sensitivity and responses to salinity and osmotic stresses are distinct, independent phenomena.     

Proteomic analysis of response to long-term continuous stress in roots of germinating soybean seeds

Germination is a complex process, highly dependent on various environmental factors, including temperature and water availability. Germinating soybean seeds are especially vulnerable to unfavorable environmental conditions and exposure to long-term abiotic stresses may result in diminishing much of the yield and most importantly – restrained germination. In the present study, a proteomic approach was employed to analyze influence of cold and osmotic stress on roots of germinated soybean (Glycine max, L.) seeds. Seeds were germinating under continuous conditions of cold stress (+10 °C/H₂O), osmotic stress (+25 °C/−0.2 MPa) as well as cold and osmotic stress combined (+10 °C/−0.2 MPa). Proteome maps established for control samples and stress-treated samples displayed 1272 CBB-stained spots. A total of 59 proteins, present in both control and stress-treated samples and showing significant differences in volume, were identified with LC/nanoESI-MS. Identified proteins divided into functional categories, revealed 9 proteins involved in plant defense, 8 proteins responsible for plant destination and storage and 10 proteins involved in various tracks of carbohydrate metabolism. Furthermore, a number of proteins were assigned to electron transport, range of metabolic pathways, secondary metabolism, protein synthesis, embryogenesis and development, signal transduction, cellular transport, translocation and storage. By analyzing differences in expression patterns, it was possible to trace the soybean response to long-term abiotic stress as well as to distinguish similarities and differences between response to cold and osmotic stress.     

A novel calmodulin-binding protein functions as a negative regulator of osmotic stress tolerance in Arabidopsis thaliana seedlings

A clone for a novel Arabidopsisthaliana calmodulin (CaM)-binding protein of 25 kDa (AtCaMBP25) has been isolated by using a radiolabelled CaM probe to screen a cDNA expression library derived from A. thaliana cell suspension cultures challenged with osmotic stress. The deduced amino acid sequence of AtCaMBP25 contains putative nuclear localization sequences and shares significant degree of similarity with hypothetical plant proteins only. Fusion of the AtCaMBP25 coding sequence to reporter genes targets the hybrid protein to the nucleus. Bacterially expressed AtCaMBP25 binds, in a calcium-dependent manner, to a canonical CaM but not to a less conserved isoform of the calcium sensor. AtCaMBP25 is encoded by a single-copy gene, whose expression is induced in Arabidopsis seedlings exposed to dehydration, low temperature or high salinity. Transgenic plants overexpressing AtCaMBP25 exhibits an increased sensitivity to both ionic (NaCl) and non-ionic (mannitol) osmotic stress during seed germination and seedling growth. By contrast, transgenic lines expressing antisense AtCaMBP25 are significantly more tolerant to mannitol and NaCl stresses than the wild type. Thus, the AtCaMBP25 gene functions as a negative effector of osmotic stress tolerance and likely participates in stress signal transduction pathways.