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Benjamin A Webb, K Karl Compton, Rafael Castañeda Saldaña, Timofey Arapov, W Keith Ray, Richard F Helm, Birgit E Scharf
The bacterium Sinorhizobium meliloti is attracted to seed exudates of its host plant alfalfa (Medicago sativa). Since quaternary ammonium compounds (QACs) are exuded by germinating seeds, we assayed chemotaxis of S. meliloti towards betonicine, choline, glycine betaine, stachydrine, and trigonelline. The wild type displayed a positive response to all QACs. Using LC-MS, we determined that each germinating alfalfa seed exuded QACs in the nanogram range. Compared to the closely related non-host species, spotted medic (Medicago arabica), unique profiles were released...
October 17, 2016: Molecular Microbiology
Harald Marx, Catherine E Minogue, Dhileepkumar Jayaraman, Alicia L Richards, Nicholas W Kwiecien, Alireza F Sihapirani, Shanmugam Rajasekar, Junko Maeda, Kevin Garcia, Angel R Del Valle-Echevarria, Jeremy D Volkening, Michael S Westphall, Sushmita Roy, Michael R Sussman, Jean-Michel Ané, Joshua J Coon
Legumes are essential components of agricultural systems because they enrich the soil in nitrogen and require little environmentally deleterious fertilizers. A complex symbiotic association between legumes and nitrogen-fixing soil bacteria called rhizobia culminates in the development of root nodules, where rhizobia fix atmospheric nitrogen and transfer it to their plant host. Here we describe a quantitative proteomic atlas of the model legume Medicago truncatula and its rhizobial symbiont Sinorhizobium meliloti, which includes more than 23,000 proteins, 20,000 phosphorylation sites, and 700 lysine acetylation sites...
October 17, 2016: Nature Biotechnology
Andrew G Palmer, Arijit Mukherjee, Danielle M Stacy, Stephen Lazar, Jean-Michel Ané, Helen E Blackwell
Density-dependent phenotypic switching in bacteria, the phenomenon of quorum sensing (QS), is instrumental in many pathogenic and mutualistic behaviors. In many Gram-negative bacteria, QS is regulated by N-acylated-l-homoserine lactones (AHLs). Synthetic analogues of these AHLs hold significant promise for regulating QS at the host-symbiont interface. Regulation depends on refined temporal and spatial models of quorums under native conditions. Critical to this is an understanding of how the presence of these signals may affect a prospective host...
October 14, 2016: Chembiochem: a European Journal of Chemical Biology
Maria D Molina-Sánchez, Fernando M García-Rodríguez, Nicolás Toro
The functional unit of mobile group II introns is a ribonucleoprotein particle (RNP) consisting of the intron-encoded protein (IEP) and the excised intron RNA. The IEP has reverse transcriptase activity but also promotes RNA splicing, and the RNA-protein complex triggers site-specific DNA insertion by reverse splicing, in a process called retrohoming. In vitro reconstituted ribonucleoprotein complexes from the Lactococcus lactis group II intron Ll.LtrB, which produce a double strand break, have recently been studied as a means of developing group II intron-based gene targeting methods for higher organisms...
2016: Frontiers in Molecular Biosciences
Maryam Zamani, George C diCenzo, Branislava Milunovic, Turlough M Finan
We report that the smb20752 gene of the alfalfa symbiont Sinorhizobium meliloti is a novel symbiotic gene required for full N2 -fixation. Deletion of smb20752 resulted in lower nitrogenase activity and smaller nodules without impacting overall nodule morphology. Orthologs of smb20752 were present in all alpha and beta rhizobia, including the ngr_b20860 gene of Sinorhizobium fredii NGR234. A ngr_b20860 mutant formed Fix(-) determinate nodules that developed normally to a late stage of the symbiosis on the host plants Macroptillium atropurpureum and Vigna unguiculata...
October 11, 2016: Environmental Microbiology
Mengsheng Gao, Hanh T Nguyen, Isai Salas Gonzalez, Max Teplitski
The RNA-binding chaperone Hfq plays a critical role in the establishment and functionality of the symbiosis between Sinorhizobium meliloti and its legume hosts. A mutation in hfq reduces symbiotic efficiency resulting in a Fix- phenotype, characterized by the inability of the bacterium to fix nitrogen. At least in part, this is due to the ability of Hfq to regulate the fixLJ operon, which encodes a sensor kinase-response regulator pair that controls expression of the nitrogenase genes. The ability of Hfq to bind fixLJ in vitro and in planta was demonstrated with gel shift and co-immunoprecipitation experiments...
October 7, 2016: Molecular Plant-microbe Interactions: MPMI
Min Luo, Thameesha T Gamage, Benjamin W Arentson, Katherine N Schlasner, Donald F Becker, John J Tanner
Aldehyde dehydrogenases (ALDHs) catalyze the NAD(P)(+)-dependent oxidation of aldehydes to carboxylic acids and are important for metabolism and detoxification. Although the ALDH superfamily fold is well established, some ALDHs contain an uncharacterized domain of unknown function (DUF) near the C-terminus of the polypeptide chain. Herein, we report the first structure of a protein containing the ALDH superfamily DUF. Proline utilization A from Sinorhizobium meliloti (SmPutA) is a 1233-residue bifunctional enzyme that contains the DUF in addition to proline dehydrogenase and L-glutamate-γ-semialdehyde dehydrogenase catalytic modules...
September 27, 2016: Journal of Biological Chemistry
Vikas Sharma, Ajit Kumar, G Archana, G Naresh Kumar
The Escherichia coli phytase gene appA encoding enzyme AppA was cloned in a broad host range plasmid pBBR1MCS2 (lac promoter), termed pVA1, and transformed into the Ensifer meliloti 1020. Transformation of pVA1 in Ensifer meliloti {E. m (pVA1)} increased its phosphatase and phytase activity by ∼9- and ∼50-fold, respectively, compared to the transformants containing empty plasmid as control {E. m (pBBR1MCS2)}. The western blot experiments using rabbit anti-AppA antibody showed that AppA is translocated into the periplasm of the host after its expression...
October 2016: Die Naturwissenschaften
Rafael Nisa-Martínez, María Dolores Molina-Sánchez, Nicolás Toro
Bacterial group II introns are self-splicing catalytic RNAs and mobile retroelements that have an open reading frame encoding an intron-encoded protein (IEP) with reverse transcriptase (RT) and RNA splicing or maturase activity. Some IEPs carry a DNA endonuclease (En) domain, which is required to cleave the bottom strand downstream from the intron-insertion site for target DNA-primed reverse transcription (TPRT) of the inserted intron RNA. Host factors complete the insertion of the intron. By contrast, the major retrohoming pathway of introns with IEPs naturally lacking endonuclease activity, like the Sinorhizobium meliloti intron RmInt1, is thought to involve insertion of the intron RNA into the template for lagging strand DNA synthesis ahead of the replication fork, with possible use of the nascent strand to prime reverse transcription of the intron RNA...
2016: PloS One
Mohammed Shabab, Markus F F Arnold, Jon Penterman, Andrew J Wommack, Hartmut T Bocker, Paul A Price, Joel S Griffitts, Elizabeth M Nolan, Graham C Walker
Interactions of rhizobia with legumes establish the chronic intracellular infection that underlies symbiosis. Within nodules of inverted repeat-lacking clade (IRLC) legumes, rhizobia differentiate into nitrogen-fixing bacteroids. This terminal differentiation is driven by host nodule-specific cysteine-rich (NCR) peptides that orchestrate the adaptation of free-living bacteria into intracellular residents. Medicago truncatula encodes a family of >700 NCR peptides that have conserved cysteine motifs. NCR247 is a cationic peptide with four cysteines that can form two intramolecular disulfide bonds in the oxidized forms...
September 6, 2016: Proceedings of the National Academy of Sciences of the United States of America
Elizaveta Krol, Christina Klaner, Petra Gnau, Volkhard Kaever, Lars-Oliver Essen, Anke Becker
To identify physiological processes affected by cAMP in the plant-symbiotic nitrogen-fixing proteobacterium Sinorhizobium meliloti Rm2011, cAMP levels were artificially increased by overexpression of its cognate adenylate/guanylate cyclase gene cyaJ. This resulted in high accumulation of cAMP in the culture supernatant, decreased swimming motility and increased production of succinoglycan, an exopolysaccharide involved in host invasion. Weaker, similar phenotypic changes were induced by overexpression of cyaB and cyaG1...
August 16, 2016: Microbiology
Annick Bertrand, Marie Bipfubusa, Catherine Dhont, François-P Chalifour, Pascal Drouin, Chantal J Beauchamp
Specific amino acids have protective functions in plants under stress conditions. This study assessed the effects of rhizobial strains on the amino acid composition in alfalfa under salt stress. Two alfalfa cultivars (Medicago sativa L. cv Apica and salt-tolerant cv Halo) in association with two Sinorhizobium meliloti strains with contrasting growth under salt stress (strain A2 and salt-tolerant strain Rm1521) were exposed to different levels of NaCl (0, 20, 40, 80 or 160 mM NaCl) under controlled conditions...
August 3, 2016: Plant Physiology and Biochemistry: PPB
Nicolás Toro, Francisco Martínez-Abarca, Manuel Fernández-López
BACKGROUND: Population genetic analyses based on genome-wide sequencing data have been carried out for Sinorhizobium medicae and S. meliloti, two closely related bacterial species forming nitrogen-fixing symbioses with plants of the genus Medicago. However, genome coverage was low or the isolates had a broad geographic distribution, making it difficult to interpret the estimated diversity and to unravel the early events underlying population genetic variations and ecological differentiation...
2016: BMC Genomics
Kata R Mikuláss, Krisztina Nagy, Balázs Bogos, Zsolt Szegletes, Etelka Kovács, Attila Farkas, György Váró, Éva Kondorosi, Attila Kereszt
BACKGROUND: Certain legume plants produce a plethora of AMP-like peptides in their symbiotic cells. The cationic subgroup of the nodule-specific cysteine-rich (NCR) peptides has potent antimicrobial activity against gram-negative and gram-positive bacteria as well as unicellular and filamentous fungi. FINDINGS: It was shown by scanning and atomic force microscopies that the cationic peptides NCR335, NCR247 and Polymyxin B (PMB) affect differentially on the surfaces of Sinorhizobium meliloti bacteria...
2016: Annals of Clinical Microbiology and Antimicrobials
Ye Zhang, Laura Anne Smallbone, George C diCenzo, Richard Morton, Turlough M Finan
BACKGROUND: Malic enzymes decarboxylate the tricarboxylic acid (TCA) cycle intermediate malate to the glycolytic end-product pyruvate and are well positioned to regulate metabolic flux in central carbon metabolism. Despite the wide distribution of these enzymes, their biological roles are unclear in part because the reaction catalyzed by these enzymes can be by-passed by other pathways. The N2-fixing alfalfa symbiont Sinorhizobium meliloti contains both a NAD(P)-malic enzyme (DME) and a separate NADP-malic enzyme (TME) and to help understand the role of these enzymes, we investigated growth, metabolomic, and transcriptional consequences resulting from loss of these enzymes in free-living cells...
2016: BMC Microbiology
Miguel López-Gómez, Javier Hidalgo-Castellanos, Carmen Lluch, José A Herrera-Cervera
Brassinosteroids (BRs) are steroid plant hormones that have been shown to be involved in the response to salt stress in cross-talk with other plant growth regulators such as polyamines (PAs). In addition, BRs are involved in the regulation of the nodulation in the rhizobium-legume symbiosis through the alteration of the PAs content in leaves. In this work, we have studied the effect of exogenous 24-epibrassinolide (EBL) in the response to salinity of nitrogen fixation in the symbiosis Medicago truncatula-Sinorhizobium meliloti...
July 19, 2016: Plant Physiology and Biochemistry: PPB
George C diCenzo, Alice Checcucci, Marco Bazzicalupo, Alessio Mengoni, Carlo Viti, Lukasz Dziewit, Turlough M Finan, Marco Galardini, Marco Fondi
The genome of about 10% of bacterial species is divided among two or more large chromosome-sized replicons. The contribution of each replicon to the microbial life cycle (for example, environmental adaptations and/or niche switching) remains unclear. Here we report a genome-scale metabolic model of the legume symbiont Sinorhizobium meliloti that is integrated with carbon utilization data for 1,500 genes with 192 carbon substrates. Growth of S. meliloti is modelled in three ecological niches (bulk soil, rhizosphere and nodule) with a focus on the role of each of its three replicons...
2016: Nature Communications
W O Draghi, M F Del Papa, C Hellweg, S A Watt, T F Watt, A Barsch, M J Lozano, A Lagares, M E Salas, J L López, F J Albicoro, J F Nilsson, G A Torres Tejerizo, M F Luna, M Pistorio, J L Boiardi, A Pühler, S Weidner, K Niehaus, A Lagares
Abiotic stresses in general and extracellular acidity in particular disturb and limit nitrogen-fixing symbioses between rhizobia and their host legumes. Except for valuable molecular-biological studies on different rhizobia, no consolidated models have been formulated to describe the central physiologic changes that occur in acid-stressed bacteria. We present here an integrated analysis entailing the main cultural, metabolic, and molecular responses of the model bacterium Sinorhizobium meliloti growing under controlled acid stress in a chemostat...
2016: Scientific Reports
Johannes Döhlemann, Meike Brennecke, Anke Becker
The soil-dwelling α-proteobacterium Sinorhizobium meliloti serves as model for studies of symbiotic nitrogen fixation, a highly important process in sustainable agriculture. Here, we report advancements of the genetic toolbox accelerating genome editing in S. meliloti. The hsdMSR operon encodes a type-I restriction-modification (R-M) system. Transformation of S. meliloti is counteracted by the restriction endonuclease HsdR degrading DNA which lacks the appropriate methylation pattern. We provide a stable S...
September 10, 2016: Journal of Biotechnology
Anna Pawlik, Magdalena Wójcik, Karol Rułka, Karolina Motyl-Gorzel, Monika Osińska-Jaroszuk, Jerzy Wielbo, Monika Marek-Kozaczuk, Anna Skorupska, Jerzy Rogalski, Grzegorz Janusz
The soil native bacterial strains were screened for laccase activity. Bacterial strain L3.8 with high laccase activity was identified as Sinorhizobium meliloti. The crude intracellular L3.8 enzyme extract was able to oxidize typical diagnostic substrates of plant and fungal laccases. Laccase L3.8 was purified 81-fold with a yield of 19.5%. The molecular mass of the purified bacterial laccase was found to be 70.0kDa and its pI was 4.77. UV-visible spectrum showed that L3.8 protein is a multicopper oxidase. The carbohydrate content of the purified enzyme was estimated at 3...
July 5, 2016: International Journal of Biological Macromolecules
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