In this work, DNA microarrays were used to investigate genome-wide transcriptional responses of Sinorhizobium meliloti to a sudden increase in external osmolarity elicited by addition of either NaCl or sucrose to exponentially growing cultures. A time course of the response within the first 4 h after the osmotic shock was established. We found that there was a general redundancy in the differentially expressed genes after NaCl or sucrose addition. Both kinds of stress resulted in induction of a large number of genes having unknown functions and in repression of many genes coding for proteins with known functions. There was a strong replicon bias in the pattern of the osmotic stress response; whereas 64% of the upregulated genes had a plasmid localization, 85% of the downregulated genes were chromosomal. Among the pSymB osmoresponsive genes, 83% were upregulated, suggesting the importance of this plasmid for S. meliloti osmoadaptation. Indeed, we identified a 200-kb region in pSymB needed for adaptation to saline shock which has a high density of osmoregulated genes.

The plant's innate immune system detects potential biotic threats through recognition of microbe-associated molecular patterns (MAMPs) or danger-associated molecular patterns (DAMPs) by pattern recognition receptors (PRR). A central regulator of pattern-triggered immunity (PTI) is the BRI1-associated kinase 1 (BAK1), which undergoes complex formation with PRR upon ligand binding. Although viral patterns inducing PTI are well known from animal systems, nothing similar has been reported for plants. Rather, antiviral defense in plants is thought to be mediated by post-transcriptional gene silencing of viral RNA or through effector-triggered immunity, i.e., recognition of virus-specific effectors by resistance proteins. Nevertheless, infection by compatible viruses can also lead to the induction of defense gene expression, indicating that plants may also recognize viruses through PTI. Here, we show that PTI, or at least the presence of the regulator BAK1, is important for antiviral defense of Arabidopsis plants. Arabidopsis bak1 mutants show increased susceptibility to three different RNA viruses during compatible interactions. Furthermore, crude viral extracts but not purified virions induce several PTI marker responses in a BAK1-dependent manner. Overall, we conclude that BAK1-dependent PTI contributes to antiviral resistance in plants.

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SummaryWe characterized the molecular function of the Pseudomonas syringae pv. tomato DC3000 (Pto) effector HopQ1.In silico studies suggest that HopQ1 might possess nucleoside hydrolase activity based on the presence of a characteristic aspartate motif. Transgenic Arabidopsis lines expressing HopQ1 or HopQ1 aspartate mutant variants were characterized with respect to flagellin triggered immunity, phenotype and changes in phytohormone content by high-performance liquid chromatography-MS (HPLC-MS).We found that HopQ1, but not its aspartate mutants, suppressed all tested immunity marker assays. Suppression of immunity was the result of a lack of the flagellin receptor FLS2, whose gene expression was abolished by HopQ1 in a promoter-dependent manner. Furthermore, HopQ1 induced cytokinin signaling in Arabidopsis and the elevation in cytokinin signaling appears to be responsible for the attenuation of FLS2 expression.We conclude that HopQ1 can activate cytokinin signaling and that moderate activation of cytokinin signaling leads to suppression of FLS2 accumulation and thus defense signaling.

BackgroundSwarming is a multicellular phenomenom characterized by the coordinated and rapid movement of bacteria across semisolid surfaces. In Sinorhizobium meliloti this type of motility has been described in a fadD mutant. To gain insights into the mechanisms underlying the process of swarming in rhizobia, we compared the transcriptome of a S. meliloti fadD mutant grown under swarming inducing conditions (semisolid medium) to those of cells grown under non-swarming conditions (broth and solid medium).ResultsMore than a thousand genes were identified as differentially expressed in response to growth on agar surfaces including genes for several metabolic activities, iron uptake, chemotaxis, motility and stress-related genes. Under swarming-specific conditions, the most remarkable response was the up-regulation of iron-related genes. We demonstrate that the pSymA plasmid and specifically genes required for the biosynthesis of the siderophore rhizobactin 1021 are essential for swarming of a S. meliloti wild-type strain but not in a fadD mutant. Moreover, high iron conditions inhibit swarming of the wild-type strain but not in mutants lacking either the iron limitation response regulator RirA or FadD.ConclusionsThe present work represents the first transcriptomic study of rhizobium growth on surfaces including swarming inducing conditions. The results have revealed major changes in the physiology of S. meliloti cells grown on a surface relative to liquid cultures. Moreover, analysis of genes responding to swarming inducing conditions led to the demonstration that iron and genes involved in rhizobactin 1021 synthesis play a role in the surface motility shown by S. meliloti which can be circumvented in a fadD mutant. This work opens a way to the identification of new traits and regulatory networks involved in swarming by rhizobia.

SummaryThe flagellin receptor of Arabidopsis, At-FLAGELLIN SENSING 2 (FLS2), has become a model for mechanistic and functional studies on plant immune receptors. Responses to flagellin or its active epitope flagellin 22 (flg22) have been extensively studied in Arabidopsis leaves. However, the perception of microbe-associated molecular patterns (MAMPs) and the immune responses in roots are poorly understood.Here, we show that isolated root tissue is able to induce pattern-triggered immunity (PTI) responses upon flg22 perception, in contrast to elf18 (the active epitope of elongation factor thermo unstable (EF-Tu)). Making use of fls2 mutant plants and tissue-specific promoters, we generated transgenic Arabidopsis lines expressing FLS2 only in certain root tissues. This allowed us to study the spatial requirements for flg22 responses in the root.Remarkably, the intensity of the immune responses did not always correlate with the expression level of the FLS2 receptor, but depended on the expressing tissue, supporting the idea that MAMP perception and sensitivity in different tissues contribute to a proper balance of defense responses according to the expected exposure to elicitors.In summary, we conclude that each investigated root tissue is able to perceive flg22 if FLS2 is present and that tissue identity is a major element of MAMP perception in roots.

The disaccharide trehalose is a well-known osmoprotectant, and trehalose accumulation through de novo biosynthesis is a common response of bacteria to abiotic stress. In this study, we have investigated the role of endogenous trehalose synthesis in the osmotolerance of Sinorhizobium meliloti. Genes coding for three possible trehalose synthesis pathways are present in the genome of S. meliloti 1021: OtsA, TreYZ, and TreS. Among these, OtsA has a major role in trehalose accumulation under all of the conditions tested and is the main system involved in osmoadaptation. Nevertheless, the other two systems are also important for growth in hyperosmotic medium. Genes for the three pathways are transcriptionally responsive to osmotic stress. The presence of at least one functional trehalose biosynthesis pathway is required for optimal competitiveness of S. meliloti to nodulate alfalfa roots.

Stimulation of potassium uptake is the most rapid response to an osmotic upshock in bacteria. This cation accumulates by a number of different transport systems whose importance has not been previously addressed for rhizobia. In silico analyses reveal the presence of genes encoding four possible potassium uptake systems in the genome of Sinorhizobium meliloti 1021: Kup1, Kup2, Trk, and Kdp. The study of the relevance of these systems under a number of different growth conditions and in symbiosis showed that the integrity of Kup1 or Trk is essential for growth under laboratory conditions even in osmotically balanced media and the absence of both systems leads to a reduced infectivity and competitiveness of the bacteria in alfalfa roots. Trk is the main system involved in the accumulation of potassium after an osmotic upshift and the most important system for growth of S. meliloti under hyperosmotic conditions. The other three systems, especially Kup1, are also relevant during the osmotic adaptation of the cell, and the relative importance of the Kdp system increases at low potassium concentrations.Rhizobia are gram-negative soil bacteria able to establish nitrogen-fixing symbiosis with leguminous plants under conditions of nitrogen deprivation. During this process, an exchange of molecular signals occurs between the two partners, leading to the formation of the root nodule, where biological nitrogen fixation takes place (12).Nearly 40% of the world's land surface can be categorized as having potential salinity problems (38). Most crops are sensitive to relatively low levels of salinity (36), and in the case of legumes, there is an additional problem since not only the plant but also the symbiotic bacteria are sensitive to salinity both at the free-living stage and during the symbiotic process (22). The Rhizobium-legume symbiosis is highly sensitive to salt or osmotic stress, since these conditions may inhibit the initial steps of the symbiotic interaction (root colonization, nodule infection, and nodule development) and also have a depressive effect on nitrogen fixation (38). It has been observed that Rhizobium mutants affected in adaptation to high salinity present deficiencies in their symbiotic capacity (25). These results emphasize the importance of studying the mechanisms of rhizobial adaptation to changes in the osmotic conditions of the soil environment.Response and adaptation to environmental stresses probably constitute a complex phenomenon involving many physiological and biochemical processes that likely reflect changes in gene expression and in the activities of enzymes and transport proteins (6,7,23). A rise in the external salinity and osmolarity triggers the outflow of water from the cell, resulting in a reduction in turgor and dehydration of the cytoplasm, causing a decrease in the cytoplasm volume and thus an increase in the ion concentration in the cytosol (33). A recent transcriptomic study of Sinorhizobium meliloti has revealed that the responses of this bacterium to ionic and nonionic compoun...