Low mannitol concentrations in Arabidopsis thaliana expressing Ectocarpus genes improve salt tolerance

Mannitol is abundant in a wide range of organisms, playing important roles in biotic and abiotic stress responses. Nonetheless, mannitol is not produced by a vast majority of plants, including many important crop plants. Mannitol-producing transgenic plants displayed improved tolerance to salt stresses though mannitol production was rather low, in the µM range, compared to mM range found in plants that innately produce mannitol. Little is known about the molecular mechanisms underlying salt tolerance triggered by low concentrations of mannitol. Reported here is the production of mannitol in Arabidopsis thaliana, by expressing two mannitol biosynthesis genes from the brown alga Ectocarpus sp. strain Ec32. To date, no brown algal genes have been successfully expressed in land plants. Expression of mannitol-1-phosphate dehydrogenase and mannitol-1-phosphatase genes was associated with the production of 42.3–52.7 nmol g−1 fresh weight of mannitol, which was sufficient to impart salinity and temperature stress tolerance. Transcriptomics revealed significant differences in the expression of numerous genes, in standard and salinity stress conditions, including genes involved in K+ homeostasis, ROS signaling, plant development, photosynthesis, ABA signaling and secondary metabolism. These results suggest that the improved tolerance to salinity stress observed in transgenic plants producing mannitol in µM range is achieved by the activation of a significant number of genes, many of which are involved in priming and modulating the expression of genes involved in a variety of functions including hormone signaling, osmotic and oxidative stress, and ion homeostasis

Sinorhizobium meliloti dctA Mutants with Partial Ability To Transport Dicarboxylic Acids

Sinorhizobium meliloti dctA encodes a transport protein needed for a successful nitrogen-fixing symbiosis between the bacteria and alfalfa. Using the toxicity of the DctA substrate fluoroorotic acid as a selective agent in an iterated selection procedure, four independent S. meliloti dctA mutants were isolated that retained some ability to transport dicarboxylates. Two mutations were located in a region called motif B located in a predicted transmembrane helix of the protein that has been shown in other members of the glutamate transporter family to be involved in cation binding. A G114D mutation was located in the third transmembrane helix, which had not previously been directly implicated in transport. Multiple sequence alignment of more than 60 members of the glutamate transporter family revealed a glycine at this position in nearly all members of the family. The fourth mutant was able to transport succinate at almost wild-type levels but was impaired in malate and fumarate transport. It contains two mutations: one in a periplasmic domain and the other predicted to be in the cytoplasm. Separation of the mutations showed that each contributed to the altered substrate preference. dctA deletion mutants that contain the mutant dctA alleles on a plasmid can proceed further in symbiotic development than null mutants of dctA, but none of the plasmids could support symbiotic nitrogen fixation, although they can transport dicarboxylates, some at relatively high levels

Exploring the long-term effect of plastic on compost microbiome.

Little is known about the ecology of microbial plastic degradation. In this study, we employed next generation amplicon sequencing to assess the effect of low-density polyethylene (LDPE) films on the structure of bacterial and fungal communities in four mature compost piles with age ranging between 2 and 10 years. While, bacterial Proteobacteria, Bacteroidetes, Actinobacteria and fungi Ascomycota were most abundant across all facilities, our data indicated significant differences in compost microbiomes between compost facilities, which might be related to compost chemical parameters, age of piles and characteristics of the feedstock. In addition, a substantial shift in the interaction pattern within microbial communities from bulk and plastic-associated (PA) compost was detected. For example, cooperation between Firmicutes Bacillaceae and Thermoactinomycetaceae was detected only in PA compost. However, based on the analysis of the diversity indices and the relative abundances of microbial taxa we can conclude that the presence of plastics in compost had no significant effect on the structure of microbial community

Microbial Consortium Associated with Crustacean Shells Composting

Soil microbes play an essential role in the biodegradation of crustacean shells, which is the process of sustainable bioconversion to chitin derivatives ultimately resulting in the promotion of plant growth properties. While a number of microorganisms with chitinolytic properties have been characterized, little is known about the microbial taxa that participate in this process either by active chitin degradation or by facilitation of this activity through nutritional cooperation and composting with the chitinolytic microorganisms. In this study, we evaluated the transformation of the soil microbiome triggered by close approximation to the green crab shell surface. Our data indicate that the microbial community associated with green crab shell matter undergoes significant specialized changes, which was reflected in a decreased fungal and bacterial Shannon diversity and evenness and in a dramatic alteration in the community composition. The relative abundance of several bacterial and fungal genera including bacteria Flavobacterium, Clostridium, Pseudomonas, and Sanguibacter and fungi Mortierella, Mycochlamys, and Talaromyces were increased with approximation to the shell surface. Association with the shell triggered significant changes in microbial cooperation that incorporate microorganisms that were previously reported to be involved in chitin degradation as well as ones with no reported chitinolytic activity. Our study indicates that the biodegradation of crab shells in soil incorporates a consortium of microorganisms that might provide a more efficient way for bioconversion

Interaction between Nitrogen and Phosphate Stress Responses in Sinorhizobium meliloti

Bacteria have developed various stress response pathways to improve their assimilation and allocation of limited nutrients, such as nitrogen and phosphate. While both the Nitrogen Stress Response (NSR) and Phosphate Stress Response (PSR) have been studied individually, there are few experiments reported that characterize effects of multiple stresses on one or more pathways in Sinorhizobium meliloti, a facultatively symbiotic, nitrogen-fixing bacteria. The PII proteins, GlnB and GlnK, regulate the NSR activity, but analysis of global transcription changes in a PII deficient mutant suggest that the S. meliloti PII proteins may also regulate the PSR. PII double deletion mutants grow very slowly and pseudoreversion of the slow growth phenotype is common. To understand this phenomenon better, transposon mutants were isolated that had a faster growing phenotype. One mutation was in phoB, the response regulator for a two component regulatory system that is important in the PSR. phoB::Tn5 mutants had different phenotypes in the wild type compared to a PII deficient background. This led to the hypothesis that phosphate stress affects the NSR and conversely, that nitrogen stress affects the PSR. Our results show that phosphate availability affects glutamine synthetase activity and expression, which are often used as indicators of NSR activity, but that nitrogen availability did not affect alkaline phosphatase activity and expression, which are indicators of PSR activity. We conclude that the NSR is co-regulated by nitrogen and phosphate, whereas the PSR does not appear to be co-regulated by nitrogen in addition to its known phosphate regulation

Transcriptome Analysis of the Role of GlnD/GlnBK in Nitrogen Stress Adaptation by <em>Sinorhizobium meliloti</em> Rm1021

<div><p>Transcriptional changes in the nitrogen stress response (NSR) of wild type <i>S. meliloti</i> Rm1021, and isogenic strains missing both PII proteins, GlnB and GlnK, or carrying a Δ<i>glnD</i>-sm2 mutation were analyzed using whole-genome microarrays. This approach allowed us to identify a number of new genes involved in the NSR and showed that the response of these bacteria to nitrogen stress overlaps with other stress responses, including induction of the <i>fixK2</i> transcriptional activator and genes that are part of the phosphate stress response. Our data also show that GlnD and GlnBK proteins may regulate many genes that are not part of the NSR. Analysis of transcriptome profiles of the Rm1021 Δ<i>glnD</i>-sm2 strain allowed us to identify several genes that appear to be regulated by GlnD without the participation of the PII proteins.</p> </div

GlnB/GlnK PII Proteins and Regulation of the Sinorhizobium meliloti Rm1021 Nitrogen Stress Response and Symbiotic Function▿

The Sinorhizobium meliloti Rm1021ΔglnD-sm2 mutant, which is predicted to make a GlnD nitrogen sensor protein truncated at its amino terminus, fixes nitrogen in symbiosis with alfalfa, but the plants cannot use this nitrogen for growth (S. N. Yurgel and M. L. Kahn, Proc. Natl. Acad. Sci. U. S. A. 105:18958-18963, 2008). The mutant also has a generalized nitrogen stress response (NSR) defect. These results suggest a connection between GlnD, symbiotic metabolism, and the NSR, but the nature of this connection is unknown. In many bacteria, GlnD modifies the PII proteins, GlnB and GlnK, as it transduces a measurement of bacterial nitrogen status to a cellular response. We have now constructed and analyzed Rm1021 mutants missing GlnB, GlnK, or both proteins. Rm1021ΔglnKΔglnB was much more defective in its NSR than either single mutant, suggesting that GlnB and GlnK overlap in regulating the NSR in free-living Rm1021. The single mutants and the double mutant all formed an effective symbiosis, indicating that symbiotic nitrogen exchange could occur without the need for either GlnB or GlnK. N-terminal truncation of the GlnD protein interfered with PII protein modification in vitro, suggesting either that unmodified PII proteins were responsible for the glnD mutant's ineffective phenotype or that connecting GlnD and appropriate symbiotic behavior does not require the PII proteins