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Sample records for da alfafa medicago

  1. G-protein from Medicago sativa: functional association to photoreceptors.

    PubMed Central

    Muschietti, J P; Martinetto, H E; Coso, O A; Farber, M D; Torres, H N; Flawia, M M

    1993-01-01

    G-protein subunits were characterized from Medicago sativa (alfalfa) seedlings. Crude membranes and GTP-Sepharose-purified fractions were electrophoresed on SDS/polyacrylamide gels and analysed by Western blotting with 9193 (anti-alpha common) and AS/7 (anti-alpha t, anti-alpha i1 and anti-alpha i2) polyclonal antibodies. These procedures led to the identification of a specific polypeptide band of about 43 kDa. Another polypeptide reacting with the SW/1 (anti-beta) antibody, of about 37 kDa, was also detected. The 43 kDa polypeptide bound specifically [alpha-32P]GTP by a photoaffinity reaction and was ADP-ribosylated by activated cholera toxin, but not by pertussis toxin. Irradiation of etiolated Medicago sativa protoplast preparations at 660 nm for 1 min produced a maximal increase in the guanosine 5'-[gamma-thio]triphosphate (GTP[35S])-binding rate. After this period of irradiation, the binding rate tended to decrease. The effect of a red-light (660 nm) pulse on the binding rate was reversed when it was immediately followed by a period of far-red (> 730 nm) illumination. These results may suggest that activation of GTP[S]-binding rate was a consequence of conversion of phytochrome Pr into the Ptr form. Images Figure 1 Figure 2 Figure 3 PMID:8484719

  2. MTGD: The Medicago truncatula genome database.

    PubMed

    Krishnakumar, Vivek; Kim, Maria; Rosen, Benjamin D; Karamycheva, Svetlana; Bidwell, Shelby L; Tang, Haibao; Town, Christopher D

    2015-01-01

    Medicago truncatula, a close relative of alfalfa (Medicago sativa), is a model legume used for studying symbiotic nitrogen fixation, mycorrhizal interactions and legume genomics. J. Craig Venter Institute (JCVI; formerly TIGR) has been involved in M. truncatula genome sequencing and annotation since 2002 and has maintained a web-based resource providing data to the community for this entire period. The website (http://www.MedicagoGenome.org) has seen major updates in the past year, where it currently hosts the latest version of the genome (Mt4.0), associated data and legacy project information, presented to users via a rich set of open-source tools. A JBrowse-based genome browser interface exposes tracks for visualization. Mutant gene symbols originally assembled and curated by the Frugoli lab are now hosted at JCVI and tie into our community annotation interface, Medicago EuCAP (to be integrated soon with our implementation of WebApollo). Literature pertinent to M. truncatula is indexed and made searchable via the Textpresso search engine. The site also implements MedicMine, an instance of InterMine that offers interconnectivity with other plant 'mines' such as ThaleMine and PhytoMine, and other model organism databases (MODs). In addition to these new features, we continue to provide keyword- and locus identifier-based searches served via a Chado-backed Tripal Instance, a BLAST search interface and bulk downloads of data sets from the iPlant Data Store (iDS). Finally, we maintain an E-mail helpdesk, facilitated by a JIRA issue tracking system, where we receive and respond to questions about the website and requests for specific data sets from the community.

  3. Overexpression of Medicago SVP genes causes floral defects and delayed flowering in Arabidopsis but only affects floral development in Medicago

    PubMed Central

    Jaudal, Mauren

    2014-01-01

    The MADS-domain transcription factor SHORT VEGETATIVE PHASE plays a key role as a repressor of the transition to flowering and as a regulator of early floral development in Arabidopsis thaliana (Arabidopsis). However, no flowering-time repressors have been functionally identified in the model legume Medicago truncatula (Medicago). In this study, phylogenetic analysis of two closely-related MtSVP-like sequences, MtSVP1 and MtSVP2, showed that their predicted proteins clustered together within the eudicot SVP clade. To determine if the MtSVP-like genes have a role in flowering, they were functionally characterized in Medicago and Arabidopsis. Transcripts of both MtSVP genes were abundant and broadly expressed in vegetative tissues but were detected at much lower levels in flowers in Medicago. Over-expression of the MtSVP genes in Arabidopsis resulted in delayed flowering and flowers with many abnormal phenotypes such as leafy sepals, changes to floral organ number and longer pedicels than the wild type. By contrast, in transgenic Medicago, over-expression of MtSVP1 resulted in alterations to flower development, but did not alter flowering time, suggesting that MtSVP1 may not function to repress the transition to flowering in Medicago. PMID:24249713

  4. The Medicago truncatula gene expression atlas web server

    PubMed Central

    2009-01-01

    Background Legumes (Leguminosae or Fabaceae) play a major role in agriculture. Transcriptomics studies in the model legume species, Medicago truncatula, are instrumental in helping to formulate hypotheses about the role of legume genes. With the rapid growth of publically available Affymetrix GeneChip Medicago Genome Array GeneChip data from a great range of tissues, cell types, growth conditions, and stress treatments, the legume research community desires an effective bioinformatics system to aid efforts to interpret the Medicago genome through functional genomics. We developed the Medicago truncatula Gene Expression Atlas (MtGEA) web server for this purpose. Description The Medicago truncatula Gene Expression Atlas (MtGEA) web server is a centralized platform for analyzing the Medicago transcriptome. Currently, the web server hosts gene expression data from 156 Affymetrix GeneChip® Medicago genome arrays in 64 different experiments, covering a broad range of developmental and environmental conditions. The server enables flexible, multifaceted analyses of transcript data and provides a range of additional information about genes, including different types of annotation and links to the genome sequence, which help users formulate hypotheses about gene function. Transcript data can be accessed using Affymetrix probe identification number, DNA sequence, gene name, functional description in natural language, GO and KEGG annotation terms, and InterPro domain number. Transcripts can also be discovered through co-expression or differential expression analysis. Flexible tools to select a subset of experiments and to visualize and compare expression profiles of multiple genes have been implemented. Data can be downloaded, in part or full, in a tabular form compatible with common analytical and visualization software. The web server will be updated on a regular basis to incorporate new gene expression data and genome annotation, and is accessible at: http

  5. An Italian functional genomic resource for Medicago truncatula

    PubMed Central

    Porceddu, Andrea; Panara, Francesco; Calderini, Ornella; Molinari, Lorna; Taviani, Paola; Lanfaloni, Luisa; Scotti, Carla; Carelli, Maria; Scaramelli, Laura; Bruschi, Gianluca; Cosson, Viviane; Ratet, Pascal; de Larembergue, Henri; Duc, Gerard; Piano, Efisio; Arcioni, Sergio

    2008-01-01

    Background Medicago truncatula is a model species for legumes. Its functional genomics have been considerably boosted in recent years due to initiatives based both in Europe and US. Collections of mutants are becoming increasingly available and this will help unravel the genetic control of important traits for many species of legumes. Findings Our report is on the production of three complementary mutant collections of the model species Medicago truncatula produced in Italy in the frame of a national genomic initiative. Well established strategies were used: Tnt1 mutagenesis, TILLING and activation tagging. Both forward and reverse genetics screenings proved the efficiency of the mutagenesis approaches adopted, enabling the isolation of interesting mutants which are in course of characterization. We anticipate that the reported collections will be complementary to the recently established functional genomics tools developed for Medicago truncatula both in Europe and in the United States. PMID:19077311

  6. Medicago truncatula as a Model for Dicot Cell Wall Development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Strong interest in renewable energy has promoted an upsurge of research on plant cell wall traits that influence the availability of lignocellulosic-derived sugars for fermentation in production of biofuels. We have initiated a genome-wide transcript profiling study using the model legume Medicago t...

  7. The medicago genome provides insight into evolution of rhizobial symbiosis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Medicago truncatula is an excellent model for the study of legume-specific biology, especially endosymbiotic interactions with bacteria and fungi. This paper describes the sequence of the euchromatic portion of the M. truncatula genome based on a recently completed BAC-based assembly supplemented by...

  8. Gene silencing in Medicago truncatula roots using RNAi.

    PubMed

    Floss, Daniela S; Schmitz, Alexa M; Starker, Colby G; Gantt, J Stephen; Harrison, Maria J

    2013-01-01

    Medicago truncatula is used widely as a model system for studies of root symbioses, interactions with parasitic nematodes and fungal pathogens, as well as studies of development and secondary metabolism. In Medicago truncatula as well as other legumes, RNA interference (RNAi) coupled with Agrobacterium rhizogenes-mediated root transformation, has been used very successfully for analyses of gene function in roots. One of the major advantages of this approach is the ease and relative speed with which transgenic roots can be generated. There are several methods, both for the generation of the RNAi constructs and the root transformation. Here we provide details of an RNAi and root transformation protocol that has been used successfully in M. truncatula and which can be scaled up to enable the analysis of several hundred constructs.

  9. Adenylate cyclase activity in a higher plant, alfalfa (Medicago sativa).

    PubMed Central

    Carricarte, V C; Bianchini, G M; Muschietti, J P; Téllez-Iñón, M T; Perticari, A; Torres, N; Flawiá, M M

    1988-01-01

    An adenylate cyclase activity in Medicago sativa L. (alfalfa) roots was partially characterized. The enzyme activity remains in the supernatant fluid after centrifugation at 105,000 g and shows in crude extracts an apparent Mr of about 84,000. The enzyme is active with Mg2+ and Ca2+ as bivalent cations, and is inhibited by EGTA and by chlorpromazine. Calmodulin from bovine brain or spinach leaves activates this adenylate cyclase. PMID:3128270

  10. The Medicago Genome Initiative: a model legume database

    PubMed Central

    Bell, Callum J.; Dixon, Richard A.; Farmer, Andrew D.; Flores, Raul; Inman, Jeff; Gonzales, Robert A.; Harrison, Maria J.; Paiva, Nancy L.; Scott, Angela D.; Weller, Jennifer W.; May, Gregory D.

    2001-01-01

    The Medicago Genome Initiative (MGI) is a database of EST sequences of the model legume Medicago truncatula. The database is available to the public and has resulted from a collaborative research effort between the Samuel Roberts Noble Foundation and the National Center for Genome Resources to investigate the genome of M.truncatula. MGI is part of the greater integrated Medicago functional genomics program at the Noble Foundation (http://www.noble .org), which is taking a global approach in studying the genetic and biochemical events associated with the growth, development and environmental interactions of this model legume. Our approach will include: large-scale EST sequencing, gene expression profiling, the generation of M.truncatula activation-tagged and promoter trap insertion mutants, high-throughput metabolic profiling, and proteome studies. These multidisciplinary information pools will be interfaced with one another to provide scientists with an integrated, holistic set of tools to address fundamental questions pertaining to legume biology. The public interface to the MGI database can be accessed at http://www.ncgr.org/research/mgi. PMID:11125064

  11. Implementing a gap analysis to conserve Caucasus Medicago species for ex situ conservation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This paper discusses the results of a Medicago gap analysis of the Former Soviet Union, with a focus on the Caucasus Region. A comprehensive database of 2400 accessions of Medicago crop wild relatives that had been collected in the area of the Former Soviet Union (FSU) was developed. Using the data...

  12. Comparative Proteomic Analysis Reveals Differential Root Proteins in Medicago sativa and Medicago truncatula in Response to Salt Stress

    PubMed Central

    Long, Ruicai; Li, Mingna; Zhang, Tiejun; Kang, Junmei; Sun, Yan; Cong, Lili; Gao, Yanli; Liu, Fengqi; Yang, Qingchuan

    2016-01-01

    Salt stress is an important abiotic stress that causes decreased crop yields. Root growth and plant activities are affected by salt stress through the actions of specific genes that help roots adapt to adverse environmental conditions. For a more comprehensive understanding of proteins affected by salinity, we used two-dimensional gel electrophoresis and mass spectrometry to characterize the proteome-level changes associated with salt stress response in Medicago sativa cv. Zhongmu-1 and Medicago truncatula cv. Jemalong A17 roots. Our physiological and phenotypic observations indicated that Zhongmu-1 was more salt tolerant than Jemalong A17. We identified 93 and 30 proteins whose abundance was significantly affected by salt stress in Zhongmu-1 and Jemalong A17 roots, respectively. The tandem mass spectrometry analysis of the differentially accumulated proteins resulted in the identification of 60 and 26 proteins in Zhongmu-1 and Jemalong A17 roots, respectively. Function analyses indicated molecule binding and catalytic activity were the two primary functional categories. These proteins have known functions in various molecular processes, including defense against oxidative stress, metabolism, photosynthesis, protein synthesis and processing, and signal transduction. The transcript levels of four identified proteins were determined by quantitative reverse transcription polymerase chain reaction. Our results indicate that some of the identified proteins may play key roles in salt stress tolerance. PMID:27066057

  13. Global reprogramming of transcription and metabolism in Medicago truncatula during progressive drought and after rewatering

    PubMed Central

    Zhang, Ji-Yi; Cruz de Carvalho, Maria H; Torres-Jerez, Ivone; Kang, Yun; Allen, Stacy N; Huhman, David V; Tang, Yuhong; Murray, Jeremy; Sumner, Lloyd W; Udvardi, Michael K

    2014-01-01

    Medicago truncatula is a model legume forage crop native to the arid and semi-arid environments of the Mediterranean. Given its drought-adapted nature, it is an ideal candidate to study the molecular and biochemical mechanisms conferring drought resistance in plants. Medicago plants were subjected to a progressive drought stress over 14 d of water withholding followed by rewatering under controlled environmental conditions. Based on physiological measurements of plant water status and changes in morphology, plants experienced mild, moderate and severe water stress before rehydration. Transcriptome analysis of roots and shoots from control, mildly, moderately and severely stressed, and rewatered plants, identified many thousands of genes that were altered in expression in response to drought. Many genes with expression tightly coupled to the plant water potential (i.e. drought intensity) were identified suggesting an involvement in Medicago drought adaptation responses. Metabolite profiling of drought-stressed plants revealed the presence of 135 polar and 165 non-polar compounds in roots and shoots. Combining Medicago metabolomic data with transcriptomic data yielded insight into the regulation of metabolic pathways operating under drought stress. Among the metabolites detected in drought-stressed Medicago plants, myo-inositol and proline had striking regulatory profiles indicating involvement in Medicago drought tolerance. Global transcriptional and metabolic responses to drought and rewatering were investigated in Medicago truncatula, a naturally drought-adapted model legume species. Integration of metabolomic and transcriptomic data yielded insights into the regulation of metabolic pathways underlying drought-stress adaptation. Many genes and metabolites with expression tightly coupled to drought intensity were identified, suggesting active involvement in Medicago drought resistance. These could prove useful targets for future translational approaches to improve

  14. Crystal structure of isoflavone reductase from alfalfa (Medicago sativa L.).

    PubMed

    Wang, Xiaoqiang; He, Xianzhi; Lin, Jianqiao; Shao, Hui; Chang, Zhenzhan; Dixon, Richard A

    2006-05-19

    Isoflavonoids play important roles in plant defense and exhibit a range of mammalian health-promoting activities. Isoflavone reductase (IFR) specifically recognizes isoflavones and catalyzes a stereospecific NADPH-dependent reduction to (3R)-isoflavanone. The crystal structure of Medicago sativa IFR with deletion of residues 39-47 has been determined at 1.6A resolution. Structural analysis, molecular modeling and docking, and comparison with the structures of other NADPH-dependent enzymes, defined the putative binding sites for co-factor and substrate and potential key residues for enzyme activity and substrate specificity. Further mutagenesis has confirmed the role of Lys144 as a catalytic residue. This study provides a structural basis for understanding the enzymatic mechanism and substrate specificity of IFRs as well as the functions of IFR-like proteins.

  15. Global reprogramming of transcription and metabolism in Medicago truncatula during progressive drought and after rewatering.

    PubMed

    Zhang, Ji-Yi; Cruz DE Carvalho, Maria H; Torres-Jerez, Ivone; Kang, Yun; Allen, Stacy N; Huhman, David V; Tang, Yuhong; Murray, Jeremy; Sumner, Lloyd W; Udvardi, Michael K

    2014-11-01

    Medicago truncatula is a model legume forage crop native to the arid and semi-arid environments of the Mediterranean. Given its drought-adapted nature, it is an ideal candidate to study the molecular and biochemical mechanisms conferring drought resistance in plants. Medicago plants were subjected to a progressive drought stress over 14 d of water withholding followed by rewatering under controlled environmental conditions. Based on physiological measurements of plant water status and changes in morphology, plants experienced mild, moderate and severe water stress before rehydration. Transcriptome analysis of roots and shoots from control, mildly, moderately and severely stressed, and rewatered plants, identified many thousands of genes that were altered in expression in response to drought. Many genes with expression tightly coupled to the plant water potential (i.e. drought intensity) were identified suggesting an involvement in Medicago drought adaptation responses. Metabolite profiling of drought-stressed plants revealed the presence of 135 polar and 165 non-polar compounds in roots and shoots. Combining Medicago metabolomic data with transcriptomic data yielded insight into the regulation of metabolic pathways operating under drought stress. Among the metabolites detected in drought-stressed Medicago plants, myo-inositol and proline had striking regulatory profiles indicating involvement in Medicago drought tolerance. PMID:24661137

  16. The Medicago sativa gene index 1.2: a web-accessible gene expression atlas for investigating expression differences between Medicago sativa subspecies

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Alfalfa (Medicago sativa L.) is the primary forage legume crop species in the United States and plays essential economic and ecological roles in agricultural systems across the country. Modern alfalfa is the result of hybridization between tetraploid M. sativa ssp. sativa and M. sativa ssp. falcata....

  17. The Medicago sativa Gene Index 1.2: A web-accessible expression atlas of two Medicago sativa sub-species

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Alfalfa is an important crop, both financially and ecologically, in the agricultural system of the United States. Using Illumina RNA-seq technology we have developed a de-novo transcriptome assembly from two Medicago sativa sub-species, sativa (B47) and falcata (F56). These two genotypes have proven...

  18. Bioinformatics Analysis of MAPKKK Family Genes in Medicago truncatula.

    PubMed

    Li, Wei; Xu, Hanyun; Liu, Ying; Song, Lili; Guo, Changhong; Shu, Yongjun

    2016-04-04

    Mitogen-activated protein kinase kinase kinase (MAPKKK) is a component of the MAPK cascade pathway that plays an important role in plant growth, development, and response to abiotic stress, the functions of which have been well characterized in several plant species, such as Arabidopsis, rice, and maize. In this study, we performed genome-wide and systemic bioinformatics analysis of MAPKKK family genes in Medicago truncatula. In total, there were 73 MAPKKK family members identified by search of homologs, and they were classified into three subfamilies, MEKK, ZIK, and RAF. Based on the genomic duplication function, 72 MtMAPKKK genes were located throughout all chromosomes, but they cluster in different chromosomes. Using microarray data and high-throughput sequencing-data, we assessed their expression profiles in growth and development processes; these results provided evidence for exploring their important functions in developmental regulation, especially in the nodulation process. Furthermore, we investigated their expression in abiotic stresses by RNA-seq, which confirmed their critical roles in signal transduction and regulation processes under stress. In summary, our genome-wide, systemic characterization and expressional analysis of MtMAPKKK genes will provide insights that will be useful for characterizing the molecular functions of these genes in M. truncatula.

  19. A Snapshot of Functional Genetic Studies in Medicago truncatula.

    PubMed

    Kang, Yun; Li, Minguye; Sinharoy, Senjuti; Verdier, Jerome

    2016-01-01

    In the current context of food security, increase of plant protein production in a sustainable manner represents one of the major challenges of agronomic research, which could be partially resolved by increased cultivation of legume crops. Medicago truncatula is now a well-established model for legume genomic and genetic studies. With the establishment of genomics tools and mutant populations in M. truncatula, it has become an important resource to answer some of the basic biological questions related to plant development and stress tolerance. This review has an objective to overview a decade of genetic studies in this model plant from generation of mutant populations to nowadays. To date, the three biological fields, which have been extensively studied in M. truncatula, are the symbiotic nitrogen fixation, the seed development, and the abiotic stress tolerance, due to their significant agronomic impacts. In this review, we summarize functional genetic studies related to these three major biological fields. We integrated analyses of a nearly exhaustive list of genes into their biological contexts in order to provide an overview of the forefront research advances in this important legume model plant. PMID:27555857

  20. Transcriptional profiling of Medicago truncatula during Erysiphe pisi infection

    PubMed Central

    Curto, Miguel; Krajinski, Franziska; Schlereth, Armin; Rubiales, Diego

    2015-01-01

    Resistance to powdery mildew has been studied in a number of plant species, yet the molecular mechanisms remain largely unknown. Transcription factors (TFs) play a critical role in the plant defense response by regulating the transcriptional machinery which coordinates the expression of a large group of genes involved in plant defense. Using high-throughput quantitative real-time PCR (qPCR) technology more than 1000 Medicago truncatula TFs were screened in a pair of susceptible and resistant genotypes of M. truncatula after 4 h of Erysiphe pisi infection. Seventy nine TF genes, belonging to 33 families showed a significant transcriptional change in response to E. pisi infection. Forty eight TF genes were differentially expressed in the resistant genotypes compared to the susceptible one in response to E. pisi infection, including pathogenesis-related transcriptional factors, AP2/EREBP (APETALA2/ETHYLENE-RESPONSIVE ELEMENT BINDING FACTORS), WRKY (highly conserved WRKYGQK amino-acid sequence), MYB (Myeloblastoma), homeodomain (HD) and zinc finger C2C2 (CYS2-CYS2), C2H2, (CYS2-HIS2), LIM (Lin-11, Isl-1, Mec-3) gene families, which are involved in known defense responses. Our results suggest that these TF genes are among the E. pisi responsive genes in resistant M. truncatula that may constitute a regulatory network which controls the transcriptional changes in defense genes involved in resistance to E. pisi. PMID:26217367

  1. Bioinformatics Analysis of MAPKKK Family Genes in Medicago truncatula

    PubMed Central

    Li, Wei; Xu, Hanyun; Liu, Ying; Song, Lili; Guo, Changhong; Shu, Yongjun

    2016-01-01

    Mitogen-activated protein kinase kinase kinase (MAPKKK) is a component of the MAPK cascade pathway that plays an important role in plant growth, development, and response to abiotic stress, the functions of which have been well characterized in several plant species, such as Arabidopsis, rice, and maize. In this study, we performed genome-wide and systemic bioinformatics analysis of MAPKKK family genes in Medicago truncatula. In total, there were 73 MAPKKK family members identified by search of homologs, and they were classified into three subfamilies, MEKK, ZIK, and RAF. Based on the genomic duplication function, 72 MtMAPKKK genes were located throughout all chromosomes, but they cluster in different chromosomes. Using microarray data and high-throughput sequencing-data, we assessed their expression profiles in growth and development processes; these results provided evidence for exploring their important functions in developmental regulation, especially in the nodulation process. Furthermore, we investigated their expression in abiotic stresses by RNA-seq, which confirmed their critical roles in signal transduction and regulation processes under stress. In summary, our genome-wide, systemic characterization and expressional analysis of MtMAPKKK genes will provide insights that will be useful for characterizing the molecular functions of these genes in M. truncatula. PMID:27049397

  2. A Snapshot of Functional Genetic Studies in Medicago truncatula

    PubMed Central

    Kang, Yun; Li, Minguye; Sinharoy, Senjuti; Verdier, Jerome

    2016-01-01

    In the current context of food security, increase of plant protein production in a sustainable manner represents one of the major challenges of agronomic research, which could be partially resolved by increased cultivation of legume crops. Medicago truncatula is now a well-established model for legume genomic and genetic studies. With the establishment of genomics tools and mutant populations in M. truncatula, it has become an important resource to answer some of the basic biological questions related to plant development and stress tolerance. This review has an objective to overview a decade of genetic studies in this model plant from generation of mutant populations to nowadays. To date, the three biological fields, which have been extensively studied in M. truncatula, are the symbiotic nitrogen fixation, the seed development, and the abiotic stress tolerance, due to their significant agronomic impacts. In this review, we summarize functional genetic studies related to these three major biological fields. We integrated analyses of a nearly exhaustive list of genes into their biological contexts in order to provide an overview of the forefront research advances in this important legume model plant. PMID:27555857

  3. Beta-Amylases from Alfalfa (Medicago sativa L.) Roots 1

    PubMed Central

    Doehlert, Douglas C.; Duke, Stanley H.; Anderson, Laurens

    1982-01-01

    Amylase was found in high activity (193 international units per milligram protein) in the tap root of alfalfa (Medicago sativa L. cv. Sonora). The activity was separated by gel filtration chromatography into two fractions with molecular weights of 65,700 (heavy amylase) and 41,700 (light amylase). Activity staining of electrophoretic gels indicated the presence of one isozyme in the heavy amylase fraction and two in the light amylase fraction. Three amylase isozymes with electrophoretic mobilities identical to those in the heavy and the light amylase fractions were the only amylases identified in crude root preparations. Both heavy and light amylases hydrolyzed amylopectin, soluble starch, and amylose but did not hydrolyze pullulan or β-limit dextrin. The ratio of viscosity change to reducing power production during starch hydrolysis was identical for both alfalfa amylase fractions and sweet potato β-amylase, while that of bacterial α-amylase was considerably higher. The identification of maltose and β-limit dextrin as hydrolytic end-products confirmed that these alfalfa root amylases are all β-amylases. The pH optimum for both β-amylase fractions was 6.0. Both light and heavy β-amylases showed normal Michaelis-Menten kinetics, with soluble starch as substrate, and had respectively Km values of 5.9 and 6.8 milligrams starch per milliliter and Vmax of 640 and 130 international units per milligram protein. Arrhenius plots indicated that the energy of activation for the heavy β-amylase remained relatively unchanged (12.7 to 13.0 kilocalories per mole) from 0 to 30°C, whereas the energy of activation for the light amylase increased from 12.0 to about 28.0 kilocalories per mole at 8.7°C as temperature was lowered. The light amylase was shown to be inhibited by maltose. Images PMID:16662350

  4. Identification of Novel Defense Response Genes in Medicago truncatula for Improving Disease Resistance in Alfalfa

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Infection of plants by pathogens initiates a cascade of defense responses that halt or limit pathogen growth. However, the role of many of the genes induced by pathogens is unknown. Transcript profiling was used to identify genes associated with defense responses in the model legume Medicago truncat...

  5. Crop wild relatives of Medicago in Russia and neighboring countries: gap analysis for effective conservation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The genus Medicago includes important species used for forage, fodder and land improvement. The countries of the Former Soviet Union have a wide diversity of alfalfa crop wild relatives and annual medic species. The N.I. Vavilov Institute of Plant Industry (VIR) has a long history of collecting and ...

  6. Identification and Expression Analysis of BURP Domain-Containing Genes in Medicago truncatula.

    PubMed

    Li, Yuan; Chen, Xue; Chen, Zhu; Cai, Ronghao; Zhang, Hongmei; Xiang, Yan

    2016-01-01

    BURP domain-containing proteins belong to a newly identified protein class that is unique to plants and plays an important role in plant development and metabolism. Although systematic characterization of BURP domain-containing proteins have been carried out in many species, such as rice, poplar and maize, little is known about BURP domain-containing proteins in Medicago. In this study, multiple bioinformatics approaches were employed to identify all the members of BURP family genes in Medicago. A complete set of 39 BURP family genes were identified. These genes have diverse structures and were distributed on chromosome 1-8 except 7. According to phylogenetic analysis, these BURP family genes could be classified into eight classes. Motif and exon-intron organization, stress-related cis-elements in promoter regions and microarray analysis of MtBURPs were also performed. Furthermore, transcript level analysis of MtBURP genes in response to drought stress revealed that all of the 39 BURP genes were regulated by drought stress. The results of this study reveal a comprehensive overview of the Medicago BURP gene family and provide the first step toward the selection of MtBURP genes for cloning and functional analysis of the BURP gene family in Medicago truncatula.

  7. Sequence divergence of microsatellites for phylogeographic assessment of Moroccan Medicago species.

    PubMed

    Zitouna, N; Marghali, S; Gharbi, M; Haddioui, A; Trifi-Farah, N

    2014-01-01

    Six Medicago species were investigated to characterize and valorize plant genetic resources of pastoral interest in Morocco. Samples were obtained from the core collection of the South Australian Research and Development Institute (SARDI). The transferability of single sequence repeat markers of Medicago truncatula was successful with 97.6% efficiency across the five species. A total of 283 alleles and 243 genotypes were generated using seven SSR markers, confirming the high level of polymorphism that is characteristic of the Medicago genus, despite a heterozygosity deficit (HO = 0.378; HE = 0.705). In addition, a high level of gene flow was revealed among the species analyzed with significant intra-specific variation. The unweighted pair group method with arithmetic mean dendrogram generated by the dissimilarity matrix revealed that M. polymorpha and M. orbicularis are closely related, and that M. truncatula is likely the ancestral species. The Pearson correlation index revealed no significant correlations between the geographic distribution of the Moroccan species and genetic similarities, indicating local adaptation of these species to different ecological environments independent of their topographical proximities. The substantial genetic variation observed was likely due to the predominance of selfing species, the relative proximity of prospected sites, human impacts, and the nature of the SARDI core collections, which are selected for their high genetic diversity. The results of this first report on Moroccan Medicago species will be of great interest for establishing strategies aiming at reasonable management and selection programs for local and Mediterranean germplasm in the face of increasing environmental change.

  8. Induction of peroxidases and superoxide dismutases in transformed embryogenic calli of alfalfa (Medicago sativa L.)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Activities of peroxidase (POD) and superoxide dismutase (SOD) enzymes were analyzed in non-regenerative transformed embryogenic lines of alfalfa (Medicago sativa L.) carrying wound-inducible oryzacystatin I (OC-I), wound-inducible oryzacystatin I antisense (OC-Ias) or hygromycin phosphotransferase (...

  9. Identification and Expression Analysis of BURP Domain-Containing Genes in Medicago truncatula

    PubMed Central

    Li, Yuan; Chen, Xue; Chen, Zhu; Cai, Ronghao; Zhang, Hongmei; Xiang, Yan

    2016-01-01

    BURP domain-containing proteins belong to a newly identified protein class that is unique to plants and plays an important role in plant development and metabolism. Although systematic characterization of BURP domain-containing proteins have been carried out in many species, such as rice, poplar and maize, little is known about BURP domain-containing proteins in Medicago. In this study, multiple bioinformatics approaches were employed to identify all the members of BURP family genes in Medicago. A complete set of 39 BURP family genes were identified. These genes have diverse structures and were distributed on chromosome 1–8 except 7. According to phylogenetic analysis, these BURP family genes could be classified into eight classes. Motif and exon-intron organization, stress-related cis-elements in promoter regions and microarray analysis of MtBURPs were also performed. Furthermore, transcript level analysis of MtBURP genes in response to drought stress revealed that all of the 39 BURP genes were regulated by drought stress. The results of this study reveal a comprehensive overview of the Medicago BURP gene family and provide the first step toward the selection of MtBURP genes for cloning and functional analysis of the BURP gene family in Medicago truncatula. PMID:27148311

  10. A set of GFP organelle marker lines for intracellular localization studies in Medicago truncatula

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genomics advances in the model legume Medicago truncatula have led to an increase in the number of identified genes encoding proteins with unknown biological function. Determining the intracellular location of uncharacterized proteins often aids in the elucidation of biological function. To expedite...

  11. Physical characteristics of Medicago truncatula calcium oxalate crystals determine their effectiveness in insect defense

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Plant structural traits can act as defense against herbivorous insects, causing them to avoid feeding on a given plant or tissue. Mineral crystals of calcium oxalate in leaves of Medicago truncatula Gaertn. have previously been shown to be effective deterrents of lepidopteran insect feeding. They ar...

  12. Structural and chemical insect defenses in calcium oxalate defective mutants of Medicago truncatula

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Plant structures can act as defense against herbivorous insects, causing them to avoid feeding on a given plant or tissue. Mineral crystals of calcium oxalate in leaves of Medicago truncatula Gaertn. are effective deterrents of lepidopteran feeding, and they inhibit conversion of leaves into insect ...

  13. Transcriptome profiling identified novel genes associated with aluminum toxicity, resistance and tolerance in Medicago truncatula

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Oligonucleotide microarrays corresponding to more than 16,000 genes were used to analyze changes in transcript accumulation in root tips of the Al-sensitive Medicago truncatula cultivar Jemalong genotype A17 in response to Al stress treatment. Out of 2,782 genes with significant changes in transcrip...

  14. Chromosomal and symbiotic relationships of rhizobia nodulating Medicago truncatula and M. laciniata.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    MultiLocus Sequence Typing (MLST) is a sequence-based method used to characterize bacterial genomes. This method was used to examine the genetic structure of Medicago-nodulating rhizobia at the Amra site, which is located in an arid region of Tunisia. Here the annual medics M. laciniata and M. tru...

  15. Medicago ciliaris growing in Tunisian soils is preferentially nodulated by Sinorhizobium medicae

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Variation in growth of Medicago ciliaris was recorded across soils from five different regions in Tunisia that represented different soil types and climatic zones. In four of these soils (Mateur, Enfidha, Rhayet and Soliman) this variation appeared to be related to the nodule number on the roots of ...

  16. Physiological and molecular characterization of aluminum resistance in Medicago truncatula

    PubMed Central

    Chandran, Divya; Sharopova, Natasha; VandenBosch, Kathryn A; Garvin, David F; Samac, Deborah A

    2008-01-01

    Background Aluminum (Al) toxicity is an important factor limiting crop production on acid soils. However, little is known about the mechanisms by which legumes respond to and resist Al stress. To explore the mechanisms of Al toxicity and resistance in legumes, we compared the impact of Al stress in Al-resistant and Al-sensitive lines of the model legume, Medicago truncatula Gaertn. Results A screen for Al resistance in 54 M. truncatula accessions identified eight Al-resistant and eight Al-sensitive lines. Comparisons of hydroponic root growth and root tip hematoxylin staining in an Al-resistant line, T32, and an Al-sensitive line, S70, provided evidence that an inducible Al exclusion mechanism occurs in T32. Transcriptional events associated with the Al resistance response were analyzed in T32 and S70 after 12 and 48 h Al treatment using oligonucleotide microarrays. Fewer genes were differentially regulated in response to Al in T32 compared to S70. Expression patterns of oxidative stress-related genes, stress-response genes and microscopic examination of Al-treated root tips suggested a lower degree of Al-induced oxidative damage to T32 root tips compared to S70. Furthermore, genes associated with cell death, senescence, and cell wall degradation were induced in both lines after 12 h of Al treatment but preferentially in S70 after 48 h of Al treatment. A multidrug and toxin efflux (MATE) transporter, previously shown to exude citrate in Arabidopsis, showed differential expression patterns in T32 and S70. Conclusion Our results identified novel genes induced by Al in Al-resistant and sensitive M. truncatula lines. In T32, transcription levels of genes related to oxidative stress were consistent with reactive oxygen species production, which would be sufficient to initiate cell death of Al-accumulating cells thereby contributing to Al exclusion and root growth recovery. In contrast, transcriptional levels of oxidative stress-related genes were consistent with

  17. Expression of coordinately regulated defense response genes and analysis of their role in disease resistance in Medicago truncatula

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Microarray technology was used to identify genes associated with disease defense responses in the model legume Medicago truncatula. Transcript profiles from leaves inoculated with Colletotrichum trifolii and Erysiphe pisi and roots infected with Phytophthora medicaginis were compared to identify gen...

  18. Databases and information integration for the Medicago truncatula genome and transcriptome.

    PubMed

    Cannon, Steven B; Crow, John A; Heuer, Michael L; Wang, Xiaohong; Cannon, Ethalinda K S; Dwan, Christopher; Lamblin, Anne-Francoise; Vasdewani, Jayprakash; Mudge, Joann; Cook, Andrew; Gish, John; Cheung, Foo; Kenton, Steve; Kunau, Timothy M; Brown, Douglas; May, Gregory D; Kim, Dongjin; Cook, Douglas R; Roe, Bruce A; Town, Chris D; Young, Nevin D; Retzel, Ernest F

    2005-05-01

    An international consortium is sequencing the euchromatic genespace of Medicago truncatula. Extensive bioinformatic and database resources support the marker-anchored bacterial artificial chromosome (BAC) sequencing strategy. Existing physical and genetic maps and deep BAC-end sequencing help to guide the sequencing effort, while EST databases provide essential resources for genome annotation as well as transcriptome characterization and microarray design. Finished BAC sequences are joined into overlapping sequence assemblies and undergo an automated annotation process that integrates ab initio predictions with EST, protein, and other recognizable features. Because of the sequencing project's international and collaborative nature, data production, storage, and visualization tools are broadly distributed. This paper describes databases and Web resources for the project, which provide support for physical and genetic maps, genome sequence assembly, gene prediction, and integration of EST data. A central project Web site at medicago.org/genome provides access to genome viewers and other resources project-wide, including an Ensembl implementation at medicago.org, physical map and marker resources at mtgenome.ucdavis.edu, and genome viewers at the University of Oklahoma (www.genome.ou.edu), the Institute for Genomic Research (www.tigr.org), and Munich Information for Protein Sequences Center (mips.gsf.de). PMID:15888676

  19. Legume genome evolution viewed through the Medicago truncatula and Lotus japonicus genomes

    PubMed Central

    Cannon, Steven B.; Sterck, Lieven; Rombauts, Stephane; Sato, Shusei; Cheung, Foo; Gouzy, Jérôme; Wang, Xiaohong; Mudge, Joann; Vasdewani, Jayprakash; Schiex, Thomas; Spannagl, Manuel; Monaghan, Erin; Nicholson, Christine; Humphray, Sean J.; Schoof, Heiko; Mayer, Klaus F. X.; Rogers, Jane; Quétier, Francis; Oldroyd, Giles E.; Debellé, Frédéric; Cook, Douglas R.; Retzel, Ernest F.; Roe, Bruce A.; Town, Christopher D.; Tabata, Satoshi; Van de Peer, Yves; Young, Nevin D.

    2006-01-01

    Genome sequencing of the model legumes, Medicago truncatula and Lotus japonicus, provides an opportunity for large-scale sequence-based comparison of two genomes in the same plant family. Here we report synteny comparisons between these species, including details about chromosome relationships, large-scale synteny blocks, microsynteny within blocks, and genome regions lacking clear correspondence. The Lotus and Medicago genomes share a minimum of 10 large-scale synteny blocks, each with substantial collinearity and frequently extending the length of whole chromosome arms. The proportion of genes syntenic and collinear within each synteny block is relatively homogeneous. Medicago–Lotus comparisons also indicate similar and largely homogeneous gene densities, although gene-containing regions in Mt occupy 20–30% more space than Lj counterparts, primarily because of larger numbers of Mt retrotransposons. Because the interpretation of genome comparisons is complicated by large-scale genome duplications, we describe synteny, synonymous substitutions and phylogenetic analyses to identify and date a probable whole-genome duplication event. There is no direct evidence for any recent large-scale genome duplication in either Medicago or Lotus but instead a duplication predating speciation. Phylogenetic comparisons place this duplication within the Rosid I clade, clearly after the split between legumes and Salicaceae (poplar). PMID:17003129

  20. Isolation and characterization of a gene from Medicago sativa L., encoding a bZIP transcription factor.

    PubMed

    Li, Yan; Sun, Yan; Yang, Qingchuan; Fang, Feng; Kang, Junmei; Zhang, Tiejun

    2013-02-01

    A full-length cDNA of 1,537 nucleotides was cloned from Medicago sativa L. cv. "Zhongmu No. 1" by rapid amplification of cDNA ends. It was designated as MsZIP, encoding a protein of 340 amino acids. The protein molecular weight was 36.43 kDa, and the theoretical isoelectric point was 5.72. The MsZIP preferentially localized in nucleus and have signal peptide. Blast analysis revealed that MsZIP shared the highest homology with some bZIP proteins of M. truncatula. The transcript of MsZIP was strongly enriched in leaf compared with root and stem of mature alfalfa plants. MsZIP was strongly induced by 15 % PEG6000 (polyethylene glycol), 50 μM abscisic acid, 200 mM NaCl, 70 μM gibberellic acid, 5 mM salicylic acid and 200 μM methyl jasmonate. Physiological resistance parameters were measured in the transgenic tobacco. Malondialdehyde content, relative water content, soluble sugar content, soluble protein content and proline content in transgenic tobacco increased compared with non-transgenic tobacco under salt stress or drought stress. The results showed that accumulation of the MsZIP protein in the vegetative tissues of transgenic plants enhanced their tolerance to osmotic pressure stress. These results demonstrate a role for the MsZIP protein in stress protection and suggest the potential of the MsZIP gene for genetic engineering of salt tolerance and drought tolerance.

  1. Molecular docking of Glycine max and Medicago truncatula ureases with urea; bioinformatics approaches.

    PubMed

    Filiz, Ertugrul; Vatansever, Recep; Ozyigit, Ibrahim Ilker

    2016-03-01

    Urease (EC 3.5.1.5) is a nickel-dependent metalloenzyme catalyzing the hydrolysis of urea into ammonia and carbon dioxide. It is present in many bacteria, fungi, yeasts and plants. Most species, with few exceptions, use nickel metalloenzyme urease to hydrolyze urea, which is one of the commonly used nitrogen fertilizer in plant growth thus its enzymatic hydrolysis possesses vital importance in agricultural practices. Considering the essentiality and importance of urea and urease activity in most plants, this study aimed to comparatively investigate the ureases of two important legume species such as Glycine max (soybean) and Medicago truncatula (barrel medic) from Fabaceae family. With additional plant species, primary and secondary structures of 37 plant ureases were comparatively analyzed using various bioinformatics tools. A structure based phylogeny was constructed using predicted 3D models of G. max and M. truncatula, whose crystallographic structures are not available, along with three additional solved urease structures from Canavalia ensiformis (PDB: 4GY7), Bacillus pasteurii (PDB: 4UBP) and Klebsiella aerogenes (PDB: 1FWJ). In addition, urease structures of these species were docked with urea to analyze the binding affinities, interacting amino acids and atom distances in urease-urea complexes. Furthermore, mutable amino acids which could potentially affect the protein active site, stability and flexibility as well as overall protein stability were analyzed in urease structures of G. max and M. truncatula. Plant ureases demonstrated similar physico-chemical properties with 833-878 amino acid residues and 89.39-90.91 kDa molecular weight with mainly acidic (5.15-6.10 pI) nature. Four protein domain structures such as urease gamma, urease beta, urease alpha and amidohydro 1 characterized the plant ureases. Secondary structure of plant ureases also demonstrated conserved protein architecture, with predominantly α-helix and random coil structures. In

  2. Border sequences of Medicago truncatula CLE36 are specifically cleaved by endoproteases common to the extracellular fluids of Medicago and soybean.

    PubMed

    Djordjevic, Michael A; Oakes, Marie; Wong, Chui E; Singh, Mohan; Bhalla, Prem; Kusumawati, Lucia; Imin, Nijat

    2011-08-01

    CLE (CLAVATA3/ESR-related) peptides are developmental regulators that are secreted into the apoplast. Little is known about the role of the sequences that flank CLE peptides in terms of their biological activity or how they are targeted by proteases that are known to liberate the final active CLE peptides from their precursor sequences. The biological activity of Medicago truncatula CLE36, which possesses broadly conserved border sequences flanking the putative final active CLE36 peptide product, was assessed. Using in vitro root growth assays and an in vitro root and callus formation assay it is shown that CLE36 peptides of different lengths possess differential biological activities. Using mass spectrometry, Glycine max and Medicago extracellular fluids were each shown to possess an endoproteolytic activity that recognizes and cleaves at border sequences in a synthetic 31 amino acid CLE36 'propeptide bait' to liberate biologically active peptide products. Inhibitor studies suggest that a subtilisin, in combination with a carboxypeptidase, liberated and trimmed CLE36, respectively, to form biologically relevant 11-15 amino acid cleavage products. The 15 amino acid cleavage product is more biologically potent on Arabidopsis than shorter or longer CLE peptides. In situ hybridization shows that the soybean orthologue of CLE36 (GmCLE34) is expressed in the provascular tissue. The results suggest that secreted subtilisins can specifically recognize the border sequences of CLE36 propeptides and liberate biologically active cleavage products. These secreted proteases may affect the stability and biological activity of CLE peptides in the apoplast or be involved in CLE36 processing.

  3. Mycorrhiza Symbiosis Increases the Surface for Sunlight Capture in Medicago truncatula for Better Photosynthetic Production

    PubMed Central

    Adolfsson, Lisa; Keresztes, Áron; Uddling, Johan; Schoefs, Benoît; Spetea, Cornelia

    2015-01-01

    Arbuscular mycorrhizal (AM) fungi play a prominent role in plant nutrition by supplying mineral nutrients, particularly inorganic phosphate (Pi), and also constitute an important carbon sink. AM stimulates plant growth and development, but the underlying mechanisms are not well understood. In this study, Medicago truncatula plants were grown with Rhizophagus irregularis BEG141 inoculum (AM), mock inoculum (control) or with Pi fertilization. We hypothesized that AM stimulates plant growth through either modifications of leaf anatomy or photosynthetic activity per leaf area. We investigated whether these effects are shared with Pi fertilization, and also assessed the relationship between levels of AM colonization and these effects. We found that increased Pi supply by either mycorrhization or fertilization led to improved shoot growth associated with increased nitrogen uptake and carbon assimilation. Both mycorrhized and Pi-fertilized plants had more and longer branches with larger and thicker leaves than the control plants, resulting in an increased photosynthetically active area. AM-specific effects were earlier appearance of the first growth axes and increased number of chloroplasts per cell section, since they were not induced by Pi fertilization. Photosynthetic activity per leaf area remained the same regardless of type of treatment. In conclusion, the increase in growth of mycorrhized and Pi-fertilized Medicago truncatula plants is linked to an increase in the surface for sunlight capture, hence increasing their photosynthetic production, rather than to an increase in the photosynthetic activity per leaf area. PMID:25615871

  4. Mycorrhiza symbiosis increases the surface for sunlight capture in Medicago truncatula for better photosynthetic production.

    PubMed

    Adolfsson, Lisa; Solymosi, Katalin; Andersson, Mats X; Keresztes, Áron; Uddling, Johan; Schoefs, Benoît; Spetea, Cornelia

    2015-01-01

    Arbuscular mycorrhizal (AM) fungi play a prominent role in plant nutrition by supplying mineral nutrients, particularly inorganic phosphate (Pi), and also constitute an important carbon sink. AM stimulates plant growth and development, but the underlying mechanisms are not well understood. In this study, Medicago truncatula plants were grown with Rhizophagus irregularis BEG141 inoculum (AM), mock inoculum (control) or with P(i) fertilization. We hypothesized that AM stimulates plant growth through either modifications of leaf anatomy or photosynthetic activity per leaf area. We investigated whether these effects are shared with P(i) fertilization, and also assessed the relationship between levels of AM colonization and these effects. We found that increased P(i) supply by either mycorrhization or fertilization led to improved shoot growth associated with increased nitrogen uptake and carbon assimilation. Both mycorrhized and P(i)-fertilized plants had more and longer branches with larger and thicker leaves than the control plants, resulting in an increased photosynthetically active area. AM-specific effects were earlier appearance of the first growth axes and increased number of chloroplasts per cell section, since they were not induced by P(i) fertilization. Photosynthetic activity per leaf area remained the same regardless of type of treatment. In conclusion, the increase in growth of mycorrhized and P(i)-fertilized Medicago truncatula plants is linked to an increase in the surface for sunlight capture, hence increasing their photosynthetic production, rather than to an increase in the photosynthetic activity per leaf area.

  5. Identification of a molecular dialogue between developing seeds of Medicago truncatula and seedborne xanthomonads.

    PubMed

    Terrasson, Emmanuel; Darrasse, Armelle; Righetti, Karima; Buitink, Julia; Lalanne, David; Ly Vu, Benoit; Pelletier, Sandra; Bolingue, William; Jacques, Marie-Agnès; Leprince, Olivier

    2015-07-01

    Plant pathogenic bacteria disseminate and survive mainly in association with seeds. This study addresses whether seeds are passive carriers or engage a molecular dialogue with pathogens during their development. We developed two pathosystems using Medicago truncatula with Xanthomonas alfalfae subsp. alfalfae (Xaa), the natural Medicago sp. pathogen and Xanthomonas campestris pv. campestris (Xcc), a Brassicaceae pathogen. Three days after flower inoculation, the transcriptome of Xcc-infected pods showed activation of an innate immune response that was strongly limited in Xcc mutated in the type three secretion system, demonstrating an incompatible interaction of Xcc with the reproductive structures. In contrast, the presence of Xaa did not result in an activation of defence genes. Transcriptome profiling during development of infected seeds exhibited time-dependent and differential responses to Xcc and Xaa. Gene network analysis revealed that the transcriptome of Xcc-infected seeds was mainly affected during seed filling whereas that of Xaa-infected seeds responded during late maturation. The Xcc-infected seed transcriptome exhibited an activation of defence response and a repression of targeted seed maturation pathways. Fifty-one percent of putative ABSCISIC ACID INSENSITIVE3 targets were deregulated by Xcc, including oleosin, cupin, legumin and chlorophyll degradation genes. At maturity, these seeds displayed decreased weight and increased chlorophyll content. In contrast, these traits were not affected by Xaa infection. These findings demonstrate the existence of a complex molecular dialogue between xanthomonads and developing seeds and provides insights into a previously unexplored trade-off between seed development and pathogen defence.

  6. Expression analysis of Medicago stem internodes identifies genes associated with regulatory control and biogenesis of plant cell walls

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Legumes have many traits that make them attractive bioenergy crops, especially as components of mixed grass stands or in crop rotations with maize. Alfalfa (Medicago sativa) is a legume with an advantage as a biomass crop because of its ability to fix atmospheric nitrogen and the easy separation of ...

  7. Effective conservation of Medicago Crop Wild Relatives in Russia and neighbouring countries: a gap analysis points the way forward

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Gap analysis is an effective way to review and refine conservation strategies for crop wild relatives. We developed a comprehensive database containing over 2400 accessions of Medicago crop wild relatives that had been collected in the area of the Former Soviet Union (FSU). Using the data we develop...

  8. Effect of wheel traffic and green manure treatments on forage yield and crown rot in alfalfa (Medicago sativa)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Harvesting alfalfa (Medicago sativa) results in mechanical wheel traffic on plants, which may damage crowns and increase the opportunity for entry of pathogens causing crown rot. Developing resistance to crown rot is problematic due to the large number of pathogens involved. Incorporation of plant b...

  9. Thiaminase activities and thiamine content of Pteridium aquilinum, Equisetum ramosissimum, Malva parviflora, Pennisetum clandestinum and Medicago sativa.

    PubMed

    Meyer, P

    1989-06-01

    Thiaminase type 1 and 2 activities and thiamine content of five plants were determined. Of these Pteridium aquilinum and Equisetum ramosissimum were found to have considerably more thiaminase activity and lower thiamine content than Malva parviflora, Pennisetum clandestinum and Medicago sativa.

  10. Application of Multilocus Sequence Typing To Study the Genetic Structure of Megaplasmids in Medicago-Nodulating Rhizobia

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A multilocus sequence typing (MLST) analysis was used to examine the relatedness and distribution of genotypic variants of the two large extrachromosomal replicons in Medicago-nodulating rhizobia (Sinorhizobium meliloti and S. medicae). One goal was to develop a strategy for the characterization of...

  11. Detection of QTL for forage yield, lodging resistance and spring vigor traits in alfalfa (Medicago sativa L.)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Alfalfa (Medicago sativa L.) is an internationally significant forage crop. Forage yield, lodging resistance and spring vigor are important agronomic traits conditioned by quantitative genetic and environmental effects. The objective of this study was to identify quantitative trait loci (QTL) and mo...

  12. Growth habit and photo-synthetic activity of shoot cultures of Medicago sativa L. transformed with the oryzacystatin II gene

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In vitro maintained shoot cultures of alfalfa (Medicago sativa L. cv. Zajeÿarska 83) that were transformed with the oryzacystatin II (OCII) gene and propagated on growth regulator-free medium were subjected to analysis of morphological characteristics and photosynthetic activity. The most striking f...

  13. ECOLOGICAL RISK ASSESSMENT OF ALFALFA (MEDICAGO VARIA L.) GENETICLALY ENGINEERED TO EXPRESS A HUMAN METALLOTHIONEIN (HMT) GENE

    EPA Science Inventory

    The objectives of these studies were two-fold: (1) to determine efficacy of low and high expression hMT gene constructs by assessing accumulation of Cu in shoots of parental and transgenic plants of alfalfa (Medicago varia L.) exposed to different concentrations of CuSO4 by addit...

  14. Identification of molecular markers associated with verticillium wilt resistance in alfalfa (medicago sativa l.) using high-resolution melting

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Verticillium wilt (VW), caused by the soilborne fungus, Verticillium alfalfae, is one of the most serious diseases of alfalfa (Medicago sativa L.) worldwide. To identify loci associated with resistance to VW, an association study was conducted using autotetraploid alfalfa populations composed of 352...

  15. Occurrence of transgenic feral alfalfa (Medicago sativa subsp. sativa L.) in alfalfa seed production areas in the United States

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genetically-engineered glyphosate-resistant alfalfa (Medicago sativa subsp. sativa) was commercialized in 2011. The potential risk of transgene dispersal into the environment is not clearly understood for alfalfa, a perennial crop that is cross-pollinated by insects. We gathered data on feral and tr...

  16. Contrasting calcium localization and speciation in leaves of Medicago trunculata mutant COD5 analyzed via synchrotron X-ray techniques

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Oxalate-producing plants accumulate calcium oxalate crystals (CaOx(C)) in the range of 3-80%(w/w) of their dry weight, reducing calcium (Ca) bioavailability. The calcium oxalate deficient 5 (cod5) mutant of Medicago truncatula has been previously shown to contain similar Ca, but lower oxalate and Ca...

  17. Medicago truncatula-derived calcium oxalate crystals have a negative impact on chewing insect performance via their physical properties

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Plant structural traits often act as defenses against herbivorous insects, causing them to avoid feeding on a given plant or tissue. Mineral crystals of calcium oxalate in Medicago truncatula Gaertn. (Fabaceae) leaves have previously been shown to be effective deterrents of lepidopteran insect feedi...

  18. Genetically modified Medicago truncatula lacking calcium oxalate has increased calcium bioavailability and partially rescues vitamin D receptor knockout mice phenotypes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    How the distribution and sequestered form of plant macro/micro-nutrients influence their bioavailability, and ultimately impact human health, is poorly understood. The legume Medicago truncatula has a portion of its tissue calcium sequestered in the form of the calcium oxalate crystal, which reduces...

  19. Leaf calcium oxalate crystal structure and its role in defense against a chewing insect in Medicago truncatula

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Crystals of calcium oxalate are common in plants and widely distributed among many plant families. These hard and largely insoluble crystals take on many shapes and sizes depending on the tissue and species. In Medicago truncatula, calcium oxalate crystals are abundant in leaves and accumulate in sh...

  20. Genetic evidence for differences in the pathways of druse and prismatic calcium oxalate crystal formation in Medicago truncatula

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Current evidence supports a single pathway of oxalate biosynthesis utilising ascorbic acid as the precursor. In this study, we begin to address the possibility that more than one pathway of oxalate biosynthesis and calcium oxalate formation occurs in Medicago truncatula Gaertn. (cv. Jemalong genotyp...

  1. Differential expression of TLP, ERF1, and R2R3MYB in annual Medicago species under salinity conditions.

    PubMed

    Gharaghani, F; Rafiei, F; Mirakhorli, N; Ebrahimie, E

    2015-08-21

    The present study was conducted to evaluate the responses of three annual Medicago species (M. truncatula, M. laciniata, and M. polymorpha) to salinity. We analyzed publicly available microarray data in NCBI pertaining to salinity-response genes in M. truncatula. Our data search identified Tubby C2 (TLP) and ethylene responsive transcription factor 1 (ERF1) as genes that potentially respond to salinity. We evaluated morpho-physiological traits and the expression of the genes in three Medicago species that had been maintained under control and saline conditions. The analysis of morpho-physiological traits showed that M. polymorpha and M. laciniata were more tolerant to salinity than M. truncatula, as they had lower reductions in biomass and dry root weight and lower increases in anthocyanin concentration. The saline conditions caused a significant increase (P < 0.01) in the expression of TLP in all Medicago species, but caused a significant decrease in the expression of ERF1. Considerable variation in anthocyanin concentrations was found among the three Medicago species. To investigate the cause of this variation, we examined the expression of R2R3MYB, a gene involved in the biosynthesis of anthocyanins. Our analysis showed that saline conditions induced high over-expression of R2R3MYB in all three Medicago spp. The high efficiency of the primer pairs used in qRT-PCR enabled us to compare the expression levels of each gene in the three species. We concluded that the more salt tolerant species showed higher expression of TLP and R2R3MYB under both control and salinity conditions.

  2. Relationships of the Woody Medicago Species (Section Dendrotelis) Assessed by Molecular Cytogenetic Analyses

    PubMed Central

    Rosato, Marcela; Castro, Mercedes; Rosselló, Josep A.

    2008-01-01

    Background and Aims The organization of rDNA genes in the woody medic species from the agronomically important Medicago section Dendrotelis was analysed to gain insight into their taxonomic relationships, to assess the levels of infraspecific variation concerning ribosomal loci in a restricted and fragmented insular species (M. citrina) and to assess the nature of its polyploidy. Methods Fluorescence in situ hybridization (FISH) was used for physical mapping of 5S and 45S ribosomal DNA genes in the three species of section Dendrotelis (M. arborea, M. citrina, M. strasseri) and the related M. marina from section Medicago. Genomic in situ hybridization (GISH) was used to assess the genomic relationships of the polyploid M. citrina with the putatively related species from section Dendrotelis. Key Results The diploid (2n = 16) M. marina has a single 45S and two 5S rDNA loci, a pattern usually detected in previous studies of Medicago diploid species. However, polyploid species from section Dendrotelis depart from expectations. The tetraploid species (2n = 32) M. arborea and M. strasseri have one 45S rDNA locus and two 5S rDNA loci, whereas in the hexaploid (2n = 48) M. citrina four 45S rDNA and five 5S rDNA loci have been detected. No single chromosome of M. citrina was uniformly labelled after using genomic probes from M. arborea and M. strasseri. Instead, cross-hybridization signals in M. citrina were restricted to terminal chromosome arms and NOR regions. Conclusions FISH results support the close taxonomic interrelationship between M. arborea and M. strasseri. In these tetraploid species, NOR loci have experienced a diploidization event through physical loss of sequences, a cytogenetic feature so far not reported in other species of the genus. The high number of rDNA loci and GISH results support the specific status for the hexaploid M. citrina, and it is suggested that this species is not an autopolyploid derivative of M. arborea or M. strasseri. Further, molecular

  3. 3D imaging and mechanical modeling of helical buckling in Medicago truncatula plant roots.

    PubMed

    Silverberg, Jesse L; Noar, Roslyn D; Packer, Michael S; Harrison, Maria J; Henley, Christopher L; Cohen, Itai; Gerbode, Sharon J

    2012-10-16

    We study the primary root growth of wild-type Medicago truncatula plants in heterogeneous environments using 3D time-lapse imaging. The growth medium is a transparent hydrogel consisting of a stiff lower layer and a compliant upper layer. We find that the roots deform into a helical shape just above the gel layer interface before penetrating into the lower layer. This geometry is interpreted as a combination of growth-induced mechanical buckling modulated by the growth medium and a simultaneous twisting near the root tip. We study the helical morphology as the modulus of the upper gel layer is varied and demonstrate that the size of the deformation varies with gel stiffness as expected by a mathematical model based on the theory of buckled rods. Moreover, we show that plant-to-plant variations can be accounted for by biomechanically plausible values of the model parameters.

  4. Medicaol, a strigolactone identified as a putative didehydro-orobanchol isomer, from Medicago truncatula.

    PubMed

    Tokunaga, Tamami; Hayashi, Hideo; Akiyama, Kohki

    2015-03-01

    A major strigolactone produced by the model legume Medicago truncatula (barrel medic) has been tentatively identified as a didehydro-orobanchol isomer. In this study, a putative didehydro-orobanchol isomer was isolated from root exudates collected from barrel medic grown hydroponically under phosphate-starved conditions. The structure and absolute configurations of this strigolactone, named medicaol, were determined by a combination of spectroscopic analysis and spectral comparison with 4-deoxymedicaol which was synthesized using solvolysis and rearrangement of hydroxymethylhexahydroindenone to tetrahydroazulenone as a key step. Medicaol has a seven-membered cycloheptadiene in the A ring instead of a typical six-membered cyclohexene. Medicaol and 4-deoxymedicaol showed activity comparable to their corresponding six-membered A ring relatives, orobanchol and 4-deoxyorobanchol (ent-2'-epi-5-deoxystrigol), in inducing hyphal branching of germinating spores of an arbuscular mycorrhizal fungus Gigaspora margarita. Plausible biosynthetic pathways from 4-deoxyorobanchol to medicaol are also proposed.

  5. Regulation of symbiotic nitrogen fixation in root nodules of alfalfa (Medicago sativa) infected with Rhizobium meliloti.

    PubMed

    Kamberger, W

    1977-10-24

    Symbiotic nitrogen fixation of Rhizobium meliloti bacteroids in Medicago sativa root nodules was suppressed by several inorganic nitrogen sources. Amino acids like glutamine, glutamic acid and aspartic acid, which can serve as sole nitrogen sources for the unnodulated plant did not influence nitrogenase activity of effective nodules, even at high concetrations. Ammonia and nitrate suppressed symbiotic nitrogen fixation in vivo only at concentrations much higher than those needed for suppression of nitrogenase activity in free living nitrogen fixing bacteria. The kinetics of suppression were slow compared with that of free living nitrogen fixing bacteria. On the other hand, nitrite, which acts as a direct inhibitor of nitrogenase, suppressed very quickly and at low concentrations. Glutamic acid and glutamine enhanced the effect of ammonia dramatically, while the suppression by nitrate was enhanced only slightly.

  6. Genomic Inventory and Transcriptional Analysis of Medicago truncatula Transporters1[W][OA

    PubMed Central

    Benedito, Vagner A.; Li, Haiquan; Dai, Xinbin; Wandrey, Maren; He, Ji; Kaundal, Rakesh; Torres-Jerez, Ivone; Gomez, S. Karen; Harrison, Maria J.; Tang, Yuhong; Zhao, Patrick X.; Udvardi, Michael K.

    2010-01-01

    Transporters move hydrophilic substrates across hydrophobic biological membranes and play key roles in plant nutrition, metabolism, and signaling and, consequently, in plant growth, development, and responses to the environment. To initiate and support systematic characterization of transporters in the model legume Medicago truncatula, we identified 3,830 transporters and classified 2,673 of these into 113 families and 146 subfamilies. Analysis of gene expression data for 2,611 of these transporters identified 129 that are expressed in an organ-specific manner, including 50 that are nodule specific and 36 specific to mycorrhizal roots. Further analysis uncovered 196 transporters that are induced at least 5-fold during nodule development and 44 in roots during arbuscular mycorrhizal symbiosis. Among the nodule- and mycorrhiza-induced transporter genes are many candidates for known transport activities in these beneficial symbioses. The data presented here are a unique resource for the selection and functional characterization of legume transporters. PMID:20023147

  7. Transcriptomic Analysis of Sinorhizobium meliloti and Medicago truncatula Symbiosis Using Nitrogen Fixation-Deficient Nodules.

    PubMed

    Lang, Claus; Long, Sharon R

    2015-08-01

    The bacterium Sinorhizobium meliloti interacts symbiotically with legume plant hosts such as Medicago truncatula to form nitrogen-fixing root nodules. During symbiosis, plant and bacterial cells differentiate in a coordinated manner, resulting in specialized plant cells that contain nitrogen-fixing bacteroids. Both plant and bacterial genes are required at each developmental stage of symbiosis. We analyzed gene expression in nodules formed by wild-type bacteria on six plant mutants with defects in nitrogen fixation. We observed differential expression of 482 S. meliloti genes with functions in cell envelope homeostasis, cell division, stress response, energy metabolism, and nitrogen fixation. We simultaneously analyzed gene expression in M. truncatula and observed differential regulation of host processes that may trigger bacteroid differentiation and control bacterial infection. Our analyses of developmentally arrested plant mutants indicate that plants use distinct means to control bacterial infection during early and late symbiotic stages.

  8. Molecular mobility in Medicago truncatula seed during early stage of germination: Neutron scattering and NMR investigations

    NASA Astrophysics Data System (ADS)

    Falourd, Xavier; Natali, Francesca; Peters, Judith; Foucat, Loïc

    2014-01-01

    First hours of Medicago truncatula (MT) seeds germination were investigated using elastic incoherent neutron scattering (EINS) and nuclear magnetic resonance (NMR), to follow respectively how macromolecular motions and water mobility evolve when water permeates into the seed. From EINS results, it was shown that there is an increase in macromolecular mobility with the water uptake. Changes in NMR relaxation parameters reflected microstructural changes associated with the recovery of the metabolic processes. The EINS investigation of the effect of temperature on macromolecular motions showed that there is a relationship between the amount of water in the seeds and the effect of freezing-thawing cycle. The NMR relaxometry results obtained at 253 K allowed establishing possible link between the freezing of water molecules tightly bound to macromolecules and their drastic motion restriction around 250 K, as observed with EINS at the highest water content.

  9. Alteration of enod40 expression modifies medicago truncatula root nodule development induced by sinorhizobium meliloti

    PubMed Central

    Charon, C; Sousa, C; Crespi, M; Kondorosi, A

    1999-01-01

    Molecular mechanisms involved in the control of root nodule organogenesis in the plant host are poorly understood. One of the nodulin genes associated with the earliest phases of this developmental program is enod40. We show here that transgenic Medicago truncatula plants overexpressing enod40 exhibit accelerated nodulation induced by Sinorhizobium meliloti. This resulted from increased initiation of primordia, which was accompanied by a proliferation response of the region close to the root tip and enhanced root length. The root cortex of the enod40-transformed plants showed increased sensitivity to nodulation signals. T(1) and T(2) descendants of two transgenic lines with reduced amounts of enod40 transcripts (probably from cosuppression) formed only a few and modified nodulelike structures. Our results suggest that induction of enod40 is a limiting step in primordium formation, and its function is required for appropriate nodule development. PMID:10521525

  10. Triterpenoid glycosides from Medicago sativa as antifungal agents against Pyricularia oryzae.

    PubMed

    Abbruscato, Pamela; Tosi, Solveig; Crispino, Laura; Biazzi, Elisa; Menin, Barbara; Picco, Anna M; Pecetti, Luciano; Avato, Pinarosa; Tava, Aldo

    2014-11-19

    The antifungal properties of saponin mixtures from alfalfa (Medicago sativa L.) tops and roots, the corresponding mixtures of prosapogenins from tops, and purified saponins and sapogenins against the causal agent of rice blast Pyricularia oryzae isolates are presented. In vitro experiments highlighted a range of activities, depending upon the assayed metabolite. The antifungal effects of the most promising prosapogenin mixture from alfalfa tops were confirmed by means of in planta tests using three different Italian cultivars of rice (Oryza sativa L. ssp. japonica), known to possess high, medium, and low blast resistance. The evidenced antifungal properties of the tested metabolites allowed some considerations on their structure-activity relationship. Results indicate that prosapogenins are active compounds to prevent the fungal attack of P. oryzae on different rice cultivars. Therefore, if properly formulated, these substances could represent a promising and environmentally friendly treatment to control rice blast. PMID:25361378

  11. Identification of a molecular dialogue between developing seeds of Medicago truncatula and seedborne xanthomonads.

    PubMed

    Terrasson, Emmanuel; Darrasse, Armelle; Righetti, Karima; Buitink, Julia; Lalanne, David; Ly Vu, Benoit; Pelletier, Sandra; Bolingue, William; Jacques, Marie-Agnès; Leprince, Olivier

    2015-07-01

    Plant pathogenic bacteria disseminate and survive mainly in association with seeds. This study addresses whether seeds are passive carriers or engage a molecular dialogue with pathogens during their development. We developed two pathosystems using Medicago truncatula with Xanthomonas alfalfae subsp. alfalfae (Xaa), the natural Medicago sp. pathogen and Xanthomonas campestris pv. campestris (Xcc), a Brassicaceae pathogen. Three days after flower inoculation, the transcriptome of Xcc-infected pods showed activation of an innate immune response that was strongly limited in Xcc mutated in the type three secretion system, demonstrating an incompatible interaction of Xcc with the reproductive structures. In contrast, the presence of Xaa did not result in an activation of defence genes. Transcriptome profiling during development of infected seeds exhibited time-dependent and differential responses to Xcc and Xaa. Gene network analysis revealed that the transcriptome of Xcc-infected seeds was mainly affected during seed filling whereas that of Xaa-infected seeds responded during late maturation. The Xcc-infected seed transcriptome exhibited an activation of defence response and a repression of targeted seed maturation pathways. Fifty-one percent of putative ABSCISIC ACID INSENSITIVE3 targets were deregulated by Xcc, including oleosin, cupin, legumin and chlorophyll degradation genes. At maturity, these seeds displayed decreased weight and increased chlorophyll content. In contrast, these traits were not affected by Xaa infection. These findings demonstrate the existence of a complex molecular dialogue between xanthomonads and developing seeds and provides insights into a previously unexplored trade-off between seed development and pathogen defence. PMID:25922487

  12. Definition and evolution of a new symbiovar, sv. rigiduloides, among Ensifer meliloti efficiently nodulating Medicago species.

    PubMed

    Gubry-Rangin, Cécile; Béna, Gilles; Cleyet-Marel, Jean-Claude; Brunel, Brigitte

    2013-10-01

    Understanding functional diversity is one of the main goals of microbial ecology, and definition of new bacterial ecotypes contributes significantly to this objective. Nitrogen-fixing bacteria provide a good system for investigation of ecotypes/biovars/symbiovars, as they present different specific associations with several host plants. This specific symbiosis is reflected both in the nodulation and fixation efficiency and in genetic characters of the bacteria, and several biovars have already been described in the bacterial species Ensifer meliloti. In the present study, the species affiliation of E. meliloti strains trapped from nodules sampled from Medicago rigiduloïdes roots was analyzed using housekeeping recA genes and DNA-DNA hybridization. The genetic diversity of these isolates was also investigated using several symbiotic markers: nodulation (nodA, nodB, nodC) and nitrogen fixation (nifH) genes, as well as the performance of phenotypic tests of nodulation capacity and nitrogen fixation efficiency. These analyses led to the proposal of a new bacterial symbiovar, E. meliloti sv. rigiduloides, that fixed nitrogen efficiently on M. rigiduloïdes, but not on Medicago truncatula. Using phylogenetic reconstructions, including the different described symbiovars, several hypotheses of lateral gene transfer and gene loss are proposed to explain the emergence of symbiovars within this species. The widespread geographical distribution of this symbiovar around the Mediterranean Basin, in contrast to restriction of M. rigiduloïdes to Eastern European countries, suggests that these isolates might also be associated with other plant species. The description of a new symbiovar within E. meliloti confirms the need for accurate bacterial ecological classification, especially for analysis of bacterial populations. PMID:23871297

  13. Evolutionary history of mitogen-activated protein kinase (MAPK) genes in Lotus, Medicago, and Phaseolus

    PubMed Central

    Neupane, Achal; Nepal, Madhav P; Benson, Benjamin V; MacArthur, Kenton J; Piya, Sarbottam

    2013-01-01

    Mitogen-Activated Protein Kinase (MAPK) genes encode proteins that mediate various signaling pathways associated with biotic and abiotic stress responses in eukaryotes. The MAPK genes form a 3-tier signal transduction cascade between cellular stimuli and physiological responses. Recent identification of soybean MAPKs and availability of genome sequences from other legume species allowed us to identify their MAPK genes. The main objectives of this study were to identify MAPKs in 3 legume species, Lotus japonicus, Medicago truncatula, and Phaseolus vulgaris, and to assess their phylogenetic relationships. We used approaches in comparative genomics for MAPK gene identification and named the newly identified genes following Arabidopsis MAPK nomenclature model. We identified 19, 18, and 15 MAPKs and 7, 4, and 9 MAPKKs in the genome of Lotus japonicus, Medicago truncatula, and Phaseolus vulgaris, respectively. Within clade placement of MAPKs and MAPKKs in the 3 legume species were consistent with those in soybean and Arabidopsis. Among 5 clades of MAPKs, 4 founder clades were consistent to MAPKs of other plant species and orthologs of MAPK genes in the fifth clade-"Clade E" were consistent with those in soybean. Our results also indicated that some gene duplication events might have occurred prior to eudicot-monocot divergence. Highly diversified MAPKs in soybean relative to those in 3 other legume species are attributable to the polyploidization events in soybean. The identification of the MAPK genes in the legume species is important for the legume crop improvement; and evolutionary relationships and functional divergence of these gene members provide insights into plant genome evolution. PMID:24317362

  14. Overall picture of expressed Heat Shock Factors in Glycine max, Lotus japonicus and Medicago truncatula.

    PubMed

    Soares-Cavalcanti, Nina M; Belarmino, Luís C; Kido, Ederson A; Pandolfi, Valesca; Marcelino-Guimarães, Francismar C; Rodrigues, Fabiana A; Pereira, Gonçalo A G; Benko-Iseppon, Ana M

    2012-06-01

    Heat shock (HS) leads to the activation of molecular mechanisms, known as HS-response, that prevent damage and enhance survival under stress. Plants have a flexible and specialized network of Heat Shock Factors (HSFs), which are transcription factors that induce the expression of heat shock proteins. The present work aimed to identify and characterize the Glycine max HSF repertory in the Soybean Genome Project (GENOSOJA platform), comparing them with other legumes (Medicago truncatula and Lotus japonicus) in view of current knowledge of Arabidopsis thaliana. The HSF characterization in leguminous plants led to the identification of 25, 19 and 21 candidate ESTs in soybean, Lotus and Medicago, respectively. A search in the SuperSAGE libraries revealed 68 tags distributed in seven HSF gene types. From the total number of obtained tags, more than 70% were related to root tissues (water deficit stress libraries vs. controls), indicating their role in abiotic stress responses, since the root is the first tissue to sense and respond to abiotic stress. Moreover, as heat stress is related to the pressure of dryness, a higher HSF expression was expected at the water deficit libraries. On the other hand, expressive HSF candidates were obtained from the library inoculated with Asian Soybean Rust, inferring crosstalk among genes associated with abiotic and biotic stresses. Evolutionary relationships among sequences were consistent with different HSF classes and subclasses. Expression profiling indicated that regulation of specific genes is associated with the stage of plant development and also with stimuli from other abiotic stresses pointing to the maintenance of HSF expression at a basal level in soybean, favoring its activation under heat-stress conditions.

  15. A CDPK isoform participates in the regulation of nodule number in Medicago truncatula.

    PubMed

    Gargantini, Pablo R; Gonzalez-Rizzo, Silvina; Chinchilla, Delphine; Raices, Marcela; Giammaria, Verónica; Ulloa, Rita M; Frugier, Florian; Crespi, Martin D

    2006-12-01

    Medicago spp. are able to develop root nodules via symbiotic interaction with Sinorhizobium meliloti. Calcium-dependent protein kinases (CDPKs) are involved in various signalling pathways in plants, and we found that expression of MtCPK3, a CDPK isoform present in roots of the model legume Medicago truncatula, is regulated during the nodulation process. Early inductions were detected 15 min and 3-4 days post-inoculation (dpi). The very early induction of CPK3 messengers was also present in inoculated M. truncatula dmi mutants and in wild-type roots subjected to salt stress, indicating that this rapid response is probably stress-related. In contrast, the later response was concomitant with cortical cell division and the formation of nodule primordia, and was not observed in wild-type roots inoculated with nod (-) strains. This late induction correlated with a change in the subcellular distribution of CDPK activities. Accordingly, an anti-MtCPK3 antibody detected two bands in soluble root extracts and one in the particulate fraction. CPK3::GFP fusions are targeted to the plasma membrane in epidermal onion cells, a localization that depends on myristoylation and palmitoylation sites of the protein, suggesting a dual subcellular localization. MtCPK3 mRNA and protein were also up-regulated by cytokinin treatment, a hormone linked to the regulation of cortical cell division and other nodulation-related responses. An RNAi-CDPK construction was used to silence CPK3 in Agrobacterium rhizogenes-transformed roots. Although no major phenotype was detected in these roots, when infected with rhizobia, the total number of nodules was, on average, twofold higher than in controls. This correlates with the lack of MtCPK3 induction in the inoculated super-nodulator sunn mutant. Our results suggest that CPK3 participates in the regulation of the symbiotic interaction. PMID:17132148

  16. Microgravity Effects on the Early Events of Biological Nitrogen Fixation in Medicago Truncatula: Results from the SyNRGE Experiment

    NASA Technical Reports Server (NTRS)

    Stutte, Gary W.; Roberts, Michael

    2012-01-01

    SyNRGE (Symbiotic Nodulation in a Reduced Gravity Environment) was a sortie mission on STS-135 in the Biological Research in Canisters (BRIC) hardware to study the effect of microgravity on a plant-microbe symbiosis resulting in biological nitrogen fixation. Medicago truncatula, a model species for th legume family, was inoculated with its bacterial symbiont, Sinorhizobium meliloti, to observe early biomolecular events associated with infection and nodulation in Petri Dish Fixation Units (PDFU's).

  17. Effects of simulated acidic rain on yields of Raphanus sativus, Lactuca sativa, Triticum aestivum and Medicago sativa

    SciTech Connect

    Evans, L.S.; Gmur, N.F.; Mancini, D.

    1982-01-01

    Experiments were performed to determine effects of simulated acidic rain on radishes (Raphanus sativus), wheat(Triticum aestivum) and alfalfa (Medicago sativa) grown under greenhouse conditions. Experimental designs allowed the detection of statistically significant differences among means that differed by less than 10%. These results suggest that the efficiency of radish foliage in increasing; root mass decreases with increased rainfall acidity since only foliage was exposed to the treatments.

  18. The impact of CdSe/ZnS Quantum Dots in cells of Medicago sativa in suspension culture

    PubMed Central

    2010-01-01

    Background Nanotechnology has the potential to provide agriculture with new tools that may be used in the rapid detection and molecular treatment of diseases and enhancement of plant ability to absorb nutrients, among others. Data on nanoparticle toxicity in plants is largely heterogeneous with a diversity of physicochemical parameters reported, which difficult generalizations. Here a cell biology approach was used to evaluate the impact of Quantum Dots (QDs) nanocrystals on plant cells, including their effect on cell growth, cell viability, oxidative stress and ROS accumulation, besides their cytomobility. Results A plant cell suspension culture of Medicago sativa was settled for the assessment of the impact of the addition of mercaptopropanoic acid coated CdSe/ZnS QDs. Cell growth was significantly reduced when 100 mM of mercaptopropanoic acid -QDs was added during the exponential growth phase, with less than 50% of the cells viable 72 hours after mercaptopropanoic acid -QDs addition. They were up taken by Medicago sativa cells and accumulated in the cytoplasm and nucleus as revealed by optical thin confocal imaging. As part of the cellular response to internalization, Medicago sativa cells were found to increase the production of Reactive Oxygen Species (ROS) in a dose and time dependent manner. Using the fluorescent dye H2DCFDA it was observable that mercaptopropanoic acid-QDs concentrations between 5-180 nM led to a progressive and linear increase of ROS accumulation. Conclusions Our results showed that the extent of mercaptopropanoic acid coated CdSe/ZnS QDs cytotoxicity in plant cells is dependent upon a number of factors including QDs properties, dose and the environmental conditions of administration and that, for Medicago sativa cells, a safe range of 1-5 nM should not be exceeded for biological applications. PMID:20929583

  19. Transcriptome analysis of Glomus mosseae/Medicago sativa mycorrhiza on atrazine stress.

    PubMed

    Song, Fuqiang; Li, Jize; Fan, Xiaoxu; Zhang, Quan; Chang, Wei; Yang, Fengshan; Geng, Gui

    2016-01-01

    Arbuscular mycorrhizal fungi (AMF) protect host plants against diverse biotic and abiotic stresses, and promote biodegradation of various contaminants. In this study effect of Glomus mosseae/Medicago sativa mycorrhiza on atrazine degradation was investigated. It was observed that the atrazine degradation rates with any addition level in mycorrhizal treatments were all significantly higher than those in non-mycorrhizal treatments. When atrazine was applied at 20 mg kg(-1), the removal efficiency was up to 74.65%. Therefore, G. mosseae can be considered as ideal inhabitants of technical installations to facilitate phytoremediation. Furthermore, a total of 10.4 Gb was used for de novo transcriptome assembly, resulting in a comprehensive data set for the identification of genes corresponding to atrazine stress in the AM association. After comparative analysis with edgeR, a total of 2,060 differential expressed genes were identified, including 570 up-regulated genes and 1490 down-regulated genes. After excluding 'function unknown' and 'general function predictions only' genes, 172 up-regulated genes were obtained. The differentially expressed genes in AM association with and without atrazine stress were associated with molecular processes/other proteins, zinc finger protein, intracellular/extracellular enzymes, structural proteins, anti-stress/anti-disease protein, electron transport-related protein, and plant growth associated protein. Our results not only prove AMF has important ecological significance on atrazine degradation but also provide evidence for the molecular mechanisms of atrazine degradation by AMF. PMID:26833403

  20. Role of motility and chemotaxis in efficiency of nodulation by Rhizobium meliloti. [Medicago sation

    SciTech Connect

    Caetano-Anolles, G.; Wall, L.G.; De Micheli, A.T.; Macchi, E.M.; Bauer, W.D.; Favelukes, G. )

    1988-04-01

    Spontaneous mutants of Rhizobium meliloti L5-30 defective in motility or chemotaxis were isolated and compared against the parent with respect to symbiotic competence. Each of the mutants were able to generate normal nodules on the host plant alfalfa (Medicago sativa), but had slightly delayed nodule formation, diminished nodulation int he initially susceptible region of the host root, and relatively low representation in nodules following co-inoculation with equal numbers of the parent. When inoculated in growth pouches with increasing dosages of the parental strain, the number of nodules formed in the initially susceptible region of the root increased sigmoidally, with an optimum concentration of about 10{sup 5} to 10{sup 6} bacteria/plant. The dose-response behavior of the nonmotile and nonchemotactic mutants was similar, but they required 10- to 30-fold higher concentrations of bacteria to generate the same number of nodules. The distribution frequencies of nodules at different positions along the primary root were very similar for the mutants and parent, indicating that reduced nodulation by the mutants in dose-response experiments probably reflects reduced efficiency of nodule initiation rather than developmentally delayed nodule initiation. The number of bacteria that firmly adsorbed to the host root surface during several hours of incubation was 5- to 20-fold greater for the parent than the mutants.

  1. The Effects of Clinorotation on the Host Plant, Medicago truncatula, and Its Microbial Symbionts

    NASA Astrophysics Data System (ADS)

    Dauzart, Ariel; Vandenbrink, Joshua; Kiss, John

    2016-02-01

    Understanding the outcome of the plant-microbe symbiosis in altered gravity is vital to developing life support systems for long-distance space travel and colonization of other planets. Thus, the aim of this research was to understand mutualistic relationships between plants and endophytic microbes under the influence of altered gravity. This project utilized the model tripartite relationship among Medicago truncatula ¬- Sinorhizobium meliloti - Rhizophagus irregularis. Plants were inoculated with rhizobial bacteria (S. meliloti), arbuscular mycorrhizal fungi (R. irregularis), or both microbes, and placed on a rotating clinostat. Vertical and horizontal static controls were also performed. Clinorotation significantly reduced M. truncatula dry mass and fresh mass compared to the static controls. The addition of rhizobia treatments under clinorotation also altered total root length and root-to-shoot fresh mass ratio. Nodule size decreased under rhizobia + clinorotation treatment, and nodule density was significantly decreased compared to the vertical treatment. However, inoculation with arbuscular mycorrhizal fungi was shown to increase biomass accumulation and nodule size. Thus, clinorotation significantly affected M. truncatula and its symbiotic relationships with S. meliloti and R. irregularis. In the long term, the results observed in this clinostat study on the changes of plant-microbe mutualism need to be investigated in spaceflight experiments. Thus, careful consideration of the symbiotic microbes of plants should be included in the design of bioregenerative life support systems needed for space travel.

  2. MtSWEET11, a Nodule-Specific Sucrose Transporter of Medicago truncatula1[OPEN

    PubMed Central

    Torres-Jerez, Ivone; Sosso, Davide; Guan, Dian; Frommer, Wolf B.

    2016-01-01

    Optimization of nitrogen fixation by rhizobia in legumes is a key area of research for sustainable agriculture. Symbiotic nitrogen fixation (SNF) occurs in specialized organs called nodules and depends on a steady supply of carbon to both plant and bacterial cells. Here we report the functional characterization of a nodule-specific Suc transporter, MtSWEET11 from Medicago truncatula. MtSWEET11 belongs to a clade of plant SWEET proteins that are capable of transporting Suc and play critical roles in pathogen susceptibility. When expressed in mammalian cells, MtSWEET11 transported sucrose (Suc) but not glucose (Glc). The MtSWEET11 gene was found to be expressed in infected root hair cells, and in the meristem, invasion zone, and vasculature of nodules. Expression of an MtSWEET11-GFP fusion protein in nodules resulted in green fluorescence associated with the plasma membrane of uninfected cells and infection thread and symbiosome membranes of infected cells. Two independent Tnt1-insertion sweet11 mutants were uncompromised in SNF. Therefore, although MtSWEET11 appears to be involved in Suc distribution within nodules, it is not crucial for SNF, probably because other Suc transporters can fulfill its role(s). PMID:27021190

  3. Glutamine synthetase in Medicago truncatula, unveiling new secrets of a very old enzyme.

    PubMed

    Seabra, Ana R; Carvalho, Helena G

    2015-01-01

    Glutamine synthetase (GS) catalyzes the first step at which nitrogen is brought into cellular metabolism and is also involved in the reassimilation of ammonium released by a number of metabolic pathways. Due to its unique position in plant nitrogen metabolism, GS plays essential roles in all aspects of plant development, from germination to senescence, and is a key component of nitrogen use efficiency (NUE) and plant yield. Understanding the mechanisms regulating GS activity is therefore of utmost importance and a great effort has been dedicated to understand how GS is regulated in different plant species. The present review summarizes exciting recent developments concerning the structure and regulation of GS isoenzymes, using the model legume Medicago truncatula. These include the understanding of the structural determinants of both the cytosolic and plastid located isoenzymes, the existence of a seed-specific GS gene unique to M. truncatula and closely related species and the discovery that GS isoenzymes are regulated by nitric oxide at the post-translational level. The data is discussed and integrated with the potential roles of the distinct GS isoenzymes within the whole plant context. PMID:26284094

  4. Transcriptional changes in response to arbuscular mycorrhiza development in the model plant Medicago truncatula.

    PubMed

    Wulf, Anne; Manthey, Katja; Doll, Jasmin; Perlick, Andreas M; Linke, Burkhard; Bekel, Thomas; Meyer, Folker; Franken, Philipp; Küster, Helge; Krajinski, Franziska

    2003-04-01

    Significant changes in root morphology and physiology during arbuscular mycorrhiza (AM) development are likely to be controlled by specific gene expression pattern in the host plant. Until now, little was known about transcriptional changes which occur AM-exclusively; that is, they do not occur during other root-microbe associations, nor are they induced by improved phosphate nutrition. In order to identify such AM-exclusive gene inductions of Medicago truncatula, we used a pool of different RNA samples as subtractor population in a suppressive subtractive hybridization (SSH) experiment. This approach resulted in the identification of a number of new AM-regulated genes. None of these genes were expressed in nonmycorrhiza roots or leaves. Electronic data obtained by comparison of the cDNA sequences to expressed sequence tag (EST) sequences from a wide range of cDNA libraries in the M. truncatula EST database (Gene Index, MtGI) support the mycorrhiza specificity of the corresponding genes, because sequences in the MtGI that were found to match the identified SSH-cDNA sequences originated exclusively from AM cDNA libraries. The promoter of one of those genes, MtGst1, showing similarities to plant glutathione-S-transferase (GST) encoding genes, was cloned and used in reporter gene studies. In contrast to studies with the potato GST gene PRP, MtGst 1 promoter activity was detected in all zones of the root cortex colonized by Glomus intraradices, but nowhere else.

  5. Transcriptional responses of Medicago truncatula upon sulfur deficiency stress and arbuscular mycorrhizal symbiosis

    PubMed Central

    Wipf, Daniel; Mongelard, Gaëlle; van Tuinen, Diederik; Gutierrez, Laurent; Casieri, Leonardo

    2014-01-01

    Sulfur plays an essential role in plants' growth and development and in their response to various abiotic and biotic stresses despite its leachability and its very low abundance in the only form that plant roots can uptake (sulfate). It is part of amino acids, glutathione (GSH), thiols of proteins and peptides, membrane sulfolipids, cell walls and secondary products, so reduced availability can drastically alter plant growth and development. The nutritional benefits of symbiotic interactions can help the plant in case of S deficiency. In particular the arbuscular mycorrhizal (AM) interaction improves N, P, and S plant nutrition, but the mechanisms behind these exchanges are not fully known yet. Although the transcriptional changes in the leguminous model plant Medicago truncatula have been already assessed in several biotic and/or abiotic conditions, S deficiency has not been considered so far. The aim of this work is to get a first overview on S-deficiency responses in the leaf and root tissues of plants interacting with the AM fungus Rhizophagus irregularis. Several hundred genes displayed significantly different transcript accumulation levels. Annotation and GO ID association were used to identify biological processes and molecular functions affected by sulfur starvation. Beside the beneficial effects of AM interaction, plants were greatly affected by the nutritional status, showing various differences in their transcriptomic footprints. Several pathways in which S plays an important role appeared to be differentially affected according to mycorrhizal status, with a generally reduced responsiveness to S deficiency in mycorrhized plants. PMID:25520732

  6. Ractopamine up take by alfalfa (Medicago sativa) and wheat (Triticum aestivum) from soil.

    PubMed

    Shelver, Weilin L; DeSutter, Thomas M

    2015-08-01

    Ractopamine is a beta adrenergic agonist used as a growth promoter in swine, cattle and turkeys. To test whether ractopamine has the potential to accumulate in plants grown in contaminated soil, a greenhouse study was conducted with alfalfa (Medicago sativa) and wheat (Triticum aestivum) grown in two soils having different concentrations of organic matter (1.3% and 2.1%), amended with 0, 0.5, and 10 μg/g of ractopamine. Plant growth ranged from 2.7 to 8.8 g dry weight (dw) for alfalfa, and 8.7 to 40 g dw for wheat and was generally greater in the higher organic matter content soil. The uptake of ractopamine in plant tissues ranged from non-detectable to 897 ng/g and was strongly dependent on soil ractopamine concentration across soil and plant tissue. When adjusted to the total fortified quantities, the amount of ractopamine taken up by the plant tissue was low, <0.01% for either soil.

  7. Transcriptome analysis of Glomus mosseae/Medicago sativa mycorrhiza on atrazine stress.

    PubMed

    Song, Fuqiang; Li, Jize; Fan, Xiaoxu; Zhang, Quan; Chang, Wei; Yang, Fengshan; Geng, Gui

    2016-02-02

    Arbuscular mycorrhizal fungi (AMF) protect host plants against diverse biotic and abiotic stresses, and promote biodegradation of various contaminants. In this study effect of Glomus mosseae/Medicago sativa mycorrhiza on atrazine degradation was investigated. It was observed that the atrazine degradation rates with any addition level in mycorrhizal treatments were all significantly higher than those in non-mycorrhizal treatments. When atrazine was applied at 20 mg kg(-1), the removal efficiency was up to 74.65%. Therefore, G. mosseae can be considered as ideal inhabitants of technical installations to facilitate phytoremediation. Furthermore, a total of 10.4 Gb was used for de novo transcriptome assembly, resulting in a comprehensive data set for the identification of genes corresponding to atrazine stress in the AM association. After comparative analysis with edgeR, a total of 2,060 differential expressed genes were identified, including 570 up-regulated genes and 1490 down-regulated genes. After excluding 'function unknown' and 'general function predictions only' genes, 172 up-regulated genes were obtained. The differentially expressed genes in AM association with and without atrazine stress were associated with molecular processes/other proteins, zinc finger protein, intracellular/extracellular enzymes, structural proteins, anti-stress/anti-disease protein, electron transport-related protein, and plant growth associated protein. Our results not only prove AMF has important ecological significance on atrazine degradation but also provide evidence for the molecular mechanisms of atrazine degradation by AMF.

  8. Does mycorrhization influence herbivore-induced volatile emission in Medicago truncatula?

    PubMed

    Leitner, Margit; Kaiser, Roland; Hause, Bettina; Boland, Wilhelm; Mithöfer, Axel

    2010-02-01

    Symbiosis with mycorrhizal fungi substantially impacts secondary metabolism and defensive traits of colonised plants. In the present study, we investigated the influence of mycorrhization (Glomus intraradices) on inducible indirect defences against herbivores using the model legume Medicago truncatula. Volatile emission by mycorrhizal and non-mycorrhizal plants was measured in reaction to damage inflicted by Spodoptera spp. and compared to the basal levels of volatile emission by plants of two different cultivars. Emitted volatiles were recorded using closed-loop stripping and gas chromatography/mass spectrometry. The documented volatile patterns were evaluated using multidimensional scaling to visualise patterns and stepwise linear discriminant analysis to distinguish volatile blends of plants with distinct physiological status and genetic background. Volatile blends emitted by different cultivars of M. truncatula prove to be clearly distinct, whereas mycorrhization only slightly influenced herbivore-induced volatile emissions. Still, the observed differences were sufficient to create classification rules to distinguish mycorrhizal and non-mycorrhizal plants by the volatiles emitted. Moreover, the effect of mycorrhization turned out to be opposed in the two cultivars examined. Root symbionts thus seem to alter indirect inducible defences of M. truncatula against insect herbivores. The impact of this effect strongly depends on the genetic background of the plant and, hence, in part explains the highly contradictory results on tripartite interactions gathered to date.

  9. The arbuscular mycorrhizal symbiosis influences sulfur starvation responses of Medicago truncatula.

    PubMed

    Sieh, Daniela; Watanabe, Mutsumi; Devers, Emanuel A; Brueckner, Franziska; Hoefgen, Rainer; Krajinski, Franziska

    2013-01-01

    Arbuscular mycorrhizal (AM) symbiosis is a mutualistic interaction that occurs between the large majority of vascular plants and fungi of the phylum Glomeromycota. In addition to other nutrients, sulfur compounds are symbiotically transferred from AM fungus to host plants; however, the physiological importance of mycorrhizal-mediated sulfur for plant metabolism has not yet been determined. We applied different sulfur and phosphate fertilization treatments to Medicago truncatula and investigated whether mycorrhizal colonization influences leaf metabolite composition and the expression of sulfur starvation-related genes. The expression pattern of sulfur starvation-related genes indicated reduced sulfur starvation responses in mycorrhizal plants grown at 1 mM phosphate nutrition. Leaf metabolite concentrations clearly showed that phosphate stress has a greater impact than sulfur stress on plant metabolism, with no demand for sulfur at strong phosphate starvation. However, when phosphate nutrition is high enough, mycorrhizal colonization reduces sulfur stress responses, probably as a result of symbiotic sulfur uptake. Mycorrhizal colonization is able to reduce sulfur starvation responses in M. truncatula when the plant's phosphate status is high enough that sulfur starvation is of physiological importance. This clearly shows the impact of mycorrhizal sulfur transfer on plant metabolism.

  10. A Medicago truncatula mutant hyper-responsive to mycorrhiza and defective for nodulation.

    PubMed

    Morandi, Dominique; le Signor, Christine; Gianinazzi-Pearson, Vivienne; Duc, Gérard

    2009-08-01

    One key strategy for the identification of plant genes required for mycorrhizal development is the use of plant mutants affected in mycorrhizal colonisation. In this paper, we report a new Medicago truncatula mutant defective for nodulation but hypermycorrhizal for symbiosis development and response. This mutant, called B9, presents a poor shoot and, especially, root development with short laterals. Inoculation with Glomus intraradices results in significantly higher root colonisation of the mutant than the wild-type genotype A17 (+20% for total root length, +16% for arbuscule frequency in the colonised part of the root, +39% for arbuscule frequency in the total root system). Mycorrhizal effects on shoot and root biomass of B9 plants are about twofold greater than in the wild-type genotype. The B9 mutant of M. truncatula is characterised by considerably higher root concentrations of the phytoestrogen coumestrol and by the novel synthesis of the coumestrol conjugate malonyl glycoside, absent from roots of wild-type plants. In conclusion, this is the first time that a hypermycorrhizal plant mutant affected negatively for nodulation (Myc(++), Nod (-/+) phenotype) is reported. This mutant represents a new tool for the study of plant genes differentially regulating mycorrhiza and nodulation symbioses, in particular, those related to autoregulation mechanisms.

  11. Medicago truncatula gene responses specific to arbuscular mycorrhiza interactions with different species and genera of Glomeromycota.

    PubMed

    Massoumou, M; van Tuinen, D; Chatagnier, O; Arnould, C; Brechenmacher, L; Sanchez, L; Selim, S; Gianinazzi, S; Gianinazzi-Pearson, V

    2007-05-01

    Plant genes exhibiting common responses to different arbuscular mycorrhizal (AM) fungi and not induced under other biological conditions have been sought for to identify specific markers for monitoring the AM symbiosis. A subset of 14 candidate Medicago truncatula genes was identified as being potentially mycorrhiza responsive in previous cDNA microarray analyses and exclusive to cDNA libraries derived from mycorrhizal root tissues. Transcriptional activity of the selected plant genes was compared during root interactions with seven AM fungi belonging to different species of Glomus, Acaulospora, Gigaspora, or Scutellospora, and under widely different biological conditions (mycorrhiza, phosphate fertilization, pathogenic/beneficial microbe interactions, incompatible plant genotype). Ten of the M. truncatula genes were commonly induced by all the tested AM fungal species, and all were activated by at least two fungi. Most of the plant genes were transcribed uniquely in mycorrhizal roots, and several were already active at the appressorium stage of fungal development. Novel data provide evidence that common recognition responses to phylogenetically different Glomeromycota exist in plants during events that are unique to mycorrhiza interactions. They indicate that plants should possess a mycorrhiza-specific genetic program which is comodulated by a broad spectrum of AM fungi.

  12. Cotyledon damage affects seed number through final plant size in the annual grassland species Medicago lupulina

    PubMed Central

    Zhang, Shiting; Zhao, Chuan; Lamb, Eric G.

    2011-01-01

    Background and Aims The effects of cotyledon damage on seedling growth and survival are relatively well established, but little is known about the effects on aspects of plant fitness such as seed number and size. Here the direct and indirect mechanisms linking cotyledon damage and plant fitness in the annual species Medicago lupulina are examined. Methods Growth and reproductive traits, including mature plant size, time to first flowering, flower number, seed number and individual seed mass were monitored in M. lupulina plants when zero, one or two cotyledons were removed at 7 d old. Structural equation modelling (SEM) was used to examine the mechanisms linking cotyledon damage to seed number and seed mass. Key Results Cotyledon damage reduced seed number but not individual seed mass. The primary mechanism was a reduction in plant biomass with cotyledon damage that in turn reduced seed number primarily through a reduction in flower numbers. Although cotyledon damage delayed flower initiation, it had little effect on seed number. Individual seed mass was not affected by cotyledon removal, but there was a trade-off between seed number and seed mass. Conclusions It is shown how a network of indirect mechanisms link damage to cotyledons and fitness in M. lupulina. Cotyledon damage had strong direct effects on both plant size and flowering phenology, but an analysis of the causal relationships among plant traits and fitness components showed that a reduction in plant size associated with cotyledon damage was an important mechanism influencing fitness. PMID:21196450

  13. Conserved genetic determinant of motor organ identity in Medicago truncatula and related legumes

    PubMed Central

    Chen, Jianghua; Moreau, Carol; Liu, Yu; Kawaguchi, Masayoshi; Hofer, Julie; Ellis, Noel; Chen, Rujin

    2012-01-01

    Plants exhibit various kinds of movements that have fascinated scientists and the public for centuries. Physiological studies in plants with the so-called motor organ or pulvinus suggest that cells at opposite sides of the pulvinus mediate leaf or leaflet movements by swelling and shrinking. How motor organ identity is determined is unknown. Using a genetic approach, we isolated a mutant designated elongated petiolule1 (elp1) from Medicago truncatula that fails to fold its leaflets in the dark due to loss of motor organs. Map-based cloning indicated that ELP1 encodes a putative plant-specific LOB domain transcription factor. RNA in situ analysis revealed that ELP1 is expressed in primordial cells that give rise to the motor organ. Ectopic expression of ELP1 resulted in dwarf plants with petioles and rachises reduced in length, and the epidermal cells gained characteristics of motor organ epidermal cells. By identifying ELP1 orthologs from other legume species, namely pea (Pisum sativum) and Lotus japonicus, we show that this motor organ identity is regulated by a conserved molecular mechanism. PMID:22689967

  14. Unveiling common responses of Medicago truncatula to appropriate and inappropriate rust species.

    PubMed

    Vaz Patto, Maria Carlota; Rubiales, Diego

    2014-01-01

    Little is known about the nature of effective defense mechanisms in legumes to pathogens of remotely related plant species. Some rust species are among pathogens with broad host range causing dramatic losses in various crop plants. To understand and compare the different host and nonhost resistance (NHR) responses of legume species against rusts, we characterized the reaction of the model legume Medicago truncatula to one appropriate (Uromyces striatus) and two inappropriate (U. viciae-fabae and U. lupinicolus) rusts. We found that similar pre and post-haustorial mechanisms of resistance appear to be operative in M. truncatula against appropriate and inappropriate rust fungus. The appropriate U. striatus germinated better on M. truncatula accessions then the inappropriate U. viciae-fabae and U. lupinicolus, but once germinated, germ tubes of the three rusts had a similar level of success in finding stomata and forming an appressoria over a stoma. However, responses to different inappropriate rust species also showed some specificity, suggesting a combination of non-specific and specific responses underlying this legume NHR to rust fungi. Further genetic and expression analysis studies will contribute to the development of the necessary molecular tools to use the present information on host and NHR mechanisms to breed for broad-spectrum resistance to rust in legume species.

  15. [Symbiosis between the nodule bacterium Sinorhizobium meliloti and alfalfa (Medicago sativa) under salinization conditions].

    PubMed

    Ibragimova, M V; Rumiantseva, M L; Onishchuk, O P; Belova, V S; Kurchak, O N; Andronov, E E; Dziubenko, N I; Simarov, B V

    2006-01-01

    Two hundred forty-three isolates of alfalfa nodule bacteria (Sinorhizobium meliloti) were obtained from legume nodules and soils sampled in the northern Aral region, experiencing secondary salinization. Isolates obtained from nodules (N isolates) were significantly more salt-tolerant than those from soils (S isolates) when grown in a liquid medium with 3.5% NaCl. It was found that wild species of alfalfa, melilot, and trigonella preferably formed symbioses with salt-tolerant nodule bacteria in both salinized and nonsalinized soils. Only two alfalfa species, Medicago falcata and M. trautvetteri, formed efficient symbioses in soils contrasting in salinity. The formation of efficient symbiosis with alfalfa in the presence of 0.6% NaCl was studied in 36 isolates (N and S) differing in salt tolerance and symbiotic efficiency. Fifteen isolates formed efficient symbioses in the presence of salt. The increase in the dry weight of the plants was 25-68% higher than in the control group. The efficiency of symbiotic interaction under salinization conditions depended on the efficiency of the isolates under standard conditions but did not correlate with the source of nodule bacteria (soil or nodule) or their salt tolerance. The results indicate that nodule bacterium strains forming efficient symbioses under salinization conditions can be found.

  16. Transcriptome analysis of Glomus mosseae/Medicago sativa mycorrhiza on atrazine stress

    PubMed Central

    Song, Fuqiang; Li, Jize; Fan, Xiaoxu; Zhang, Quan; Chang, Wei; Yang, Fengshan; Geng, Gui

    2016-01-01

    Arbuscular mycorrhizal fungi (AMF) protect host plants against diverse biotic and abiotic stresses, and promote biodegradation of various contaminants. In this study effect of Glomus mosseae/Medicago sativa mycorrhiza on atrazine degradation was investigated. It was observed that the atrazine degradation rates with any addition level in mycorrhizal treatments were all significantly higher than those in non- mycorrhizal treatments. When atrazine was applied at 20 mg kg−1, the removal efficiency was up to 74.65%. Therefore, G. mosseae can be considered as ideal inhabitants of technical installations to facilitate phytoremediation. Furthermore, a total of 10.4 Gb was used for de novo transcriptome assembly, resulting in a comprehensive data set for the identification of genes corresponding to atrazine stress in the AM association. After comparative analysis with edgeR, a total of 2,060 differential expressed genes were identified, including 570 up-regulated genes and 1490 down-regulated genes. After excluding ‘function unknown’ and ‘general function predictions only’ genes, 172 up-regulated genes were obtained. The differentially expressed genes in AM association with and without atrazine stress were associated with molecular processes/other proteins, zinc finger protein, intracellular/extracellular enzymes, structural proteins, anti-stress/anti-disease protein, electron transport-related protein, and plant growth associated protein. Our results not only prove AMF has important ecological significance on atrazine degradation but also provide evidence for the molecular mechanisms of atrazine degradation by AMF. PMID:26833403

  17. Mixed Nodule Infection in Sinorhizobium meliloti–Medicago sativa Symbiosis Suggest the Presence of Cheating Behavior

    PubMed Central

    Checcucci, Alice; Azzarello, Elisa; Bazzicalupo, Marco; Galardini, Marco; Lagomarsino, Alessandra; Mancuso, Stefano; Marti, Lucia; Marzano, Maria C.; Mocali, Stefano; Squartini, Andrea; Zanardo, Marina; Mengoni, Alessio

    2016-01-01

    In the symbiosis between rhizobia and legumes, host plants can form symbiotic root nodules with multiple rhizobial strains, potentially showing different symbiotic performances in nitrogen fixation. Here, we investigated the presence of mixed nodules, containing rhizobia with different degrees of mutualisms, and evaluate their relative fitness in the Sinorhizobium meliloti–Medicago sativa model symbiosis. We used three S. meliloti strains, the mutualist strains Rm1021 and BL225C and the non-mutualist AK83. We performed competition experiments involving both in vitro and in vivo symbiotic assays with M. sativa host plants. We show the occurrence of a high number (from 27 to 100%) of mixed nodules with no negative effect on both nitrogen fixation and plant growth. The estimation of the relative fitness as non-mutualist/mutualist ratios in single nodules shows that in some nodules the non-mutualist strain efficiently colonized root nodules along with the mutualist ones. In conclusion, we can support the hypothesis that in S. meliloti–M. sativa symbiosis mixed nodules are formed and allow non-mutualist or less-mutualist bacterial partners to be less or not sanctioned by the host plant, hence allowing a potential form of cheating behavior to be present in the nitrogen fixing symbiosis. PMID:27379128

  18. Adaptation to climate through flowering phenology: a case study in Medicago truncatula.

    PubMed

    Burgarella, Concetta; Chantret, Nathalie; Gay, Laurène; Prosperi, Jean-Marie; Bonhomme, Maxime; Tiffin, Peter; Young, Nevin D; Ronfort, Joelle

    2016-07-01

    Local climatic conditions likely constitute an important selective pressure on genes underlying important fitness-related traits such as flowering time, and in many species, flowering phenology and climatic gradients strongly covary. To test whether climate shapes the genetic variation on flowering time genes and to identify candidate flowering genes involved in the adaptation to environmental heterogeneity, we used a large Medicago truncatula core collection to examine the association between nucleotide polymorphisms at 224 candidate genes and both climate variables and flowering phenotypes. Unlike genome-wide studies, candidate gene approaches are expected to enrich for the number of meaningful trait associations because they specifically target genes that are known to affect the trait of interest. We found that flowering time mediates adaptation to climatic conditions mainly by variation at genes located upstream in the flowering pathways, close to the environmental stimuli. Variables related to the annual precipitation regime reflected selective constraints on flowering time genes better than the other variables tested (temperature, altitude, latitude or longitude). By comparing phenotype and climate associations, we identified 12 flowering genes as the most promising candidates responsible for phenological adaptation to climate. Four of these genes were located in the known flowering time QTL region on chromosome 7. However, climate and flowering associations also highlighted largely distinct gene sets, suggesting different genetic architectures for adaptation to climate and flowering onset. PMID:27144929

  19. NODULES WITH ACTIVATED DEFENSE 1 is required for maintenance of rhizobial endosymbiosis in Medicago truncatula.

    PubMed

    Wang, Chao; Yu, Haixiang; Luo, Li; Duan, Liujian; Cai, Liuyang; He, Xinxing; Wen, Jiangqi; Mysore, Kirankumar S; Li, Guoliang; Xiao, Aifang; Duanmu, Deqiang; Cao, Yangrong; Hong, Zonglie; Zhang, Zhongming

    2016-10-01

    The symbiotic interaction between legume plants and rhizobia results in the formation of root nodules, in which symbiotic plant cells host and harbor thousands of nitrogen-fixing rhizobia. Here, a Medicago truncatula nodules with activated defense 1 (nad1) mutant was identified using reverse genetics methods. The mutant phenotype was characterized using cell and molecular biology approaches. An RNA-sequencing technique was used to analyze the transcriptomic reprogramming of nad1 mutant nodules. In the nad1 mutant plants, rhizobial infection and propagation in infection threads are normal, whereas rhizobia and their symbiotic plant cells become necrotic immediately after rhizobia are released from infection threads into symbiotic cells of nodules. Defense-associated responses were detected in nad1 nodules. NAD1 is specifically present in root nodule symbiosis plants with the exception of Morus notabilis, and the transcript is highly induced in nodules. NAD1 encodes a small uncharacterized protein with two predicted transmembrane helices and is localized at the endoplasmic reticulum. Our data demonstrate a positive role for NAD1 in the maintenance of rhizobial endosymbiosis during nodulation. PMID:27245091

  20. Influence of arbuscular mycorrhizal colonisation on cadmium induced Medicago truncatula root isoflavonoid accumulation.

    PubMed

    Aloui, Achref; Dumas-Gaudot, Eliane; Daher, Zeina; van Tuinen, Diederik; Aschi-Smit, Samira; Morandi, Dominique

    2012-11-01

    Cadmium is a serious environmental pollution threats to the planet. Its accumulation in plants affects many cellular functions, resulting in growth and development inhibition, whose mechanisms are not fully understood. However, some fungi forming arbuscular mycorrhizal symbiosis with the majority of plant species have the capacity to buffer the deleterious effect of this heavy metal. In the present work we investigated the capacity of Rhizophagus irregularis (syn. Glomus irregularis) to alleviate cadmium stress in Medicago truncatula. In spite of a reduction in all mycorrhizal parameters, plants colonized for 21 days by R. irregularis and treated by 2 mg kg⁻¹ cadmium displayed less growth inhibition in comparison to plants grown without cadmium. Cadmium strongly increased the accumulation of some isoflavonoids and their derivates: formononetin, malonylononin, medicarpin 3-O-β-(6'-malonylglucoside), medicarpin and coumestrol. Interestingly, in plants colonized by R. irregularis we noticed a strong reduction of the cadmium-induced accumulation of root isoflavonoids, a part for medicarpin and coumestrol. Moreover, transcripts of chalcone reductase, a protein that we reported previously as being down-regulated in R. irregularis-colonized M. truncatula roots, revealed a similar expression pattern with a strong increase in response to cadmium and a reduced expression in cadmium-treated mycorrhizal roots.

  1. Caesium inhibits the colonization of Medicago truncatula by arbuscular mycorrhizal fungi.

    PubMed

    Wiesel, Lea; Dubchak, Sergiy; Turnau, Katarzyna; Broadley, Martin R; White, Philip J

    2015-03-01

    Contamination of soils with radioisotopes of caesium (Cs) is of concern because of their emissions of harmful β and γ radiation. Radiocaesium enters the food chain through vegetation and the intake of Cs can affect the health of organisms. Arbuscular mycorrhizal (AM) fungi form mutualistic symbioses with plants through colonization of the roots and previous studies on the influence of AM on Cs concentrations in plants have given inconsistent results. These studies did not investigate the influence of Cs on AM fungi and it is therefore not known if Cs has a direct effect on AM colonization. Here, we investigated whether Cs influences AM colonization and if this effect impacts on the influence of Rhizophagus intraradices on Cs accumulation by Medicago truncatula. M. truncatula was grown with or without R. intraradices in pots containing different concentrations of Cs. Here, we present the first evidence that colonization of plants by AM fungi can be negatively affected by increasing Cs concentrations in the soil. Mycorrhizal colonization had little effect on root or shoot Cs concentrations. In conclusion, the colonization by AM fungi is impaired by high Cs concentrations and this direct effect of soil Cs on AM colonization might explain the inconsistent results reported in literature that have shown increased, decreased or unaffected Cs concentrations in AM plants.

  2. Biogenesis of protein bodies during vicilin accumulation in Medicago truncatula immature seeds

    PubMed Central

    2012-01-01

    Background Grain legumes play a worldwide role as a source of plant proteins for feed and food. In the model legume Medicago truncatula, the organisation of protein storage vacuoles (PSV) in maturing seeds remains unknown. Findings The sub-cellular events accompanying the accumulation of vicilin (globulin7S) were analysed during seed mid-maturation. Immuno-detection of vicilin in light microscopy, allowed a semi-quantitative assessment of the protein body complement. The identified populations of vicilin-containing protein bodies are distinguished by their number and size which allowed to propose a model of their biogenesis. Two distributions were detected, enabling a separation of their processing at early and mid maturation stages. The largest protein bodies, at 16 and 20 days after pollination (DAP), were formed by the fusion of small bodies. They have probably attained their final size and correspond to mature vicilin aggregations. Electron microscopic observations revealed the association of the dense protein bodies with rough endoplasmic reticulum. The presence of a ribosome layer surrounding protein bodies, would support an endoplasmic reticulum–vacuole trafficking pathway. Conclusions The stastistic analysis may be useful for screening mutations of candidate genes governing protein content. The definitive evidence for an ER-storage vacuole pathway corresponds to a challenge, for the storage of post-translationally unstable proteins. It was proposed for the accumulation of one class of storage protein, the vicilins. This alternative pathway is a matter of controversy in dicotyledonous seeds. PMID:22862819

  3. [Determination of Hard Rate of Alfalfa (Medicago sativa L.) Seeds with Near Infrared Spectroscopy].

    PubMed

    Wang, Xin-xun; Chen, Ling-ling; Zhang, Yun-wei; Mao, Pei-sheng

    2016-03-01

    Alfalfa (Medicago sativa L.) is the most commonly grown forage crop due to its better quality characteristics and high adaptability in China. However, there was 20%-80% hard seeds in alfalfa which could not be identified easily from non hard seeds which would cause the loss of seed utilization value and plant production. This experiment was designed for 121 samples of alfalfa. Seeds were collected according to different regions, harvested year and varieties. 31 samples were artificial matched as hard rates ranging from 20% to 80% to establish a model for hard seed rate by near infrared spectroscopy (NIRS) with Partial Least Square (PLS). The objective of this study was to establish a model and to estimate the efficiency of NIRS for determining hard rate of alfalfa seeds. The results showed that the correlation coefficient (R2(cal)) of calibration model was 0.981 6, root mean square error of cross validation (RMSECV) was 5.32, and the ratio of prediction to deviation (RPD) was 3.58. The forecast model in this experiment presented the satisfied precision. The proposed method using NIRS technology is feasible for identification and classification of hard seed in alfalfa. A new method, as nondestructive testing of hard seed rate, was provided to theoretical basis for fast nondestructive detection of hard seed rates in alfalfa. PMID:27400509

  4. Stem nematode counteracts plant resistance of aphids in alfalfa, Medicago sativa.

    PubMed

    Ramirez, Ricardo A; Spears, Lori R

    2014-10-01

    Plants are exploited by a diverse community of insect herbivores and phytopathogens that interact indirectly through plant-mediated interactions. Generally, plants are thought to respond to insects and pathogens through different defensive signaling pathways. As plants are selected for resistance to one phytophagous organism type (insect vs. pathogen) in managed systems, it is not clear how this selection may affect community interactions. This study examined the effect of nematode-resistant varieties on aphid (Acyrthosiphon pisum) suppression, and then determined how infection by the stem nematode, Ditylenchus dipsaci, mediated ecological effects on aphids and on plant defense proteins. Four alfalfa (Medicago sativa) varieties were selected with resistance to nematodes only (+,-), aphids only (-,+), nematodes and aphids (+,+), and susceptibility to nematodes and aphids (-,-). Field and greenhouse experiments were conducted to isolate the effect of nematode infection and aphid abundance on each variety. We found that varieties resistant to nematode, regardless of aphid resistance, had the lowest aphid counts, suggesting possible cross-resistance. Aphid abundance, however, increased when plants were exposed to nematodes. Resistant varieties were associated with elevated saponins but these compounds were not affected by insect or pathogen feeding. Concentrations of peroxidases and trypsin inhibitors, however, were increased in nematode resistant varieties when exposed to nematodes and aphids, respectively. The patterns of plant defense were variable, and a combination of resistance traits and changes in nutrient availability may drive positive interactions between nematodes and aphids aboveground.

  5. Complete genome sequence of the Medicago microsymbiont Ensifer (Sinorhizobium) medicae strain WSM419

    SciTech Connect

    Reeve, Wayne; Chain, Patrick S. G.; O'Hara, Graham; Ardley, Julie; Nandesena, Kemanthi; Brau, Lambert; Tiwari, Ravi; Malfatti, Stephanie; Kiss, Hajnalka; Lapidus, Alla L.; Copeland, A; Nolan, Matt; Land, Miriam L; Hauser, Loren John; Chang, Yun-Juan; Ivanova, N; Mavromatis, K; Markowitz, Victor; Kyrpides, Nikos C; Gollagher, Margaret; Yates, Ron; Dilworth, Michael; Howieson, John

    2010-01-01

    Ensifer (Sinorhizobium) medicae is an effective nitrogen fixing microsymbiont of a diverse range of annual Medicago (medic) species. Strain WSM419 is an aerobic, motile, non-spore forming, Gram-negative rod isolated from a M. murex root nodule collected in Sardinia, Italy in 1981. WSM419 was manufactured commercially in Australia as an inoculant for annual medics during 1985 to 1993 due to its nitrogen fixation, saprophytic competence and acid tolerance properties. Here we describe the basic features of this organism, together with the complete genome sequence, and annotation. This is the first report of a complete genome sequence for a microsymbiont of the group of annual medic species adapted to acid soils. We reveal that its genome size is 6,817,576 bp encoding 6,518 protein-coding genes and 81 RNA only encoding genes. The genome contains a chromosome of size 3,781,904 bp and 3 plasmids of size 1,570,951 bp, 1,245,408 bp and 219,313 bp. The smallest plasmid is a feature unique to this medic microsymbiont.

  6. Mixed Nodule Infection in Sinorhizobium meliloti-Medicago sativa Symbiosis Suggest the Presence of Cheating Behavior.

    PubMed

    Checcucci, Alice; Azzarello, Elisa; Bazzicalupo, Marco; Galardini, Marco; Lagomarsino, Alessandra; Mancuso, Stefano; Marti, Lucia; Marzano, Maria C; Mocali, Stefano; Squartini, Andrea; Zanardo, Marina; Mengoni, Alessio

    2016-01-01

    In the symbiosis between rhizobia and legumes, host plants can form symbiotic root nodules with multiple rhizobial strains, potentially showing different symbiotic performances in nitrogen fixation. Here, we investigated the presence of mixed nodules, containing rhizobia with different degrees of mutualisms, and evaluate their relative fitness in the Sinorhizobium meliloti-Medicago sativa model symbiosis. We used three S. meliloti strains, the mutualist strains Rm1021 and BL225C and the non-mutualist AK83. We performed competition experiments involving both in vitro and in vivo symbiotic assays with M. sativa host plants. We show the occurrence of a high number (from 27 to 100%) of mixed nodules with no negative effect on both nitrogen fixation and plant growth. The estimation of the relative fitness as non-mutualist/mutualist ratios in single nodules shows that in some nodules the non-mutualist strain efficiently colonized root nodules along with the mutualist ones. In conclusion, we can support the hypothesis that in S. meliloti-M. sativa symbiosis mixed nodules are formed and allow non-mutualist or less-mutualist bacterial partners to be less or not sanctioned by the host plant, hence allowing a potential form of cheating behavior to be present in the nitrogen fixing symbiosis. PMID:27379128

  7. Comparative sequence and expression analysis of tapetum specific male sterility related genes in Medicago truncatula.

    PubMed

    Shao, L H; Zheng, X W; Yi, D X; Li, C

    2016-01-01

    Heterosis, or enhancement through outbreeding, is one of the most promising approaches for increasing crop yield. Male sterility (MS), which promotes heterosis, has been widely applied in hybrid crop production. Medicago truncatula is a model legume species and is closely related to M. sativa, an important legume forage plant. Although the molecular mechanisms of MS in M. truncatula and M. sativa remain unclear, several studies of MS have been conducted in Arabidopsis thaliana. Previous research has shown that MS is associated with the destruction of tapetal cell layers. Disruption of tapetum developmental processes may result in pollen abortion. In an effort to identify genes useful for breeding in M. sativa, we identified MS related genes in M. truncatula using BLAST and homology to A. thaliana genes. In this study, we identified 63 tapetum specific male sterility (TSMS) related genes. The length of TSMS genes varied from 225 to 3747 bp. We identified 15 conserved domains and 8 cis-elements associated with TSMS related genes. Analysis of the phylogenetic relationships among these genes allowed them to be classified into three groups, MtTsms A, MtTsms B, and MtTsms C. Expression analyses revealed that these genes may be involved in developmental processes and response to abiotic stress. PMID:27421020

  8. Evolution by gene duplication of Medicago truncatula PISTILLATA-like transcription factors

    PubMed Central

    Roque, Edelín; Fares, Mario A.; Yenush, Lynne; Rochina, Mari Cruz; Wen, Jiangqi; Mysore, Kirankumar S.; Gómez-Mena, Concepción; Beltrán, José Pío; Cañas, Luis A.

    2016-01-01

    PISTILLATA (PI) is a member of the B-function MADS-box gene family, which controls the identity of both petals and stamens in Arabidopsis thaliana. In Medicago truncatula (Mt), there are two PI-like paralogs, known as MtPI and MtNGL9. These genes differ in their expression patterns, but it is not known whether their functions have also diverged. Describing the evolution of certain duplicated genes, such as transcription factors, remains a challenge owing to the complex expression patterns and functional divergence between the gene copies. Here, we report a number of functional studies, including analyses of gene expression, protein–protein interactions, and reverse genetic approaches designed to demonstrate the respective contributions of each M. truncatula PI-like paralog to the B-function in this species. Also, we have integrated molecular evolution approaches to determine the mode of evolution of Mt PI-like genes after duplication. Our results demonstrate that MtPI functions as a master regulator of B-function in M. truncatula, maintaining the overall ancestral function, while MtNGL9 does not seem to have a role in this regard, suggesting that the pseudogenization could be the functional evolutionary fate for this gene. However, we provide evidence that purifying selection is the primary evolutionary force acting on this paralog, pinpointing the conservation of its biochemical function and, alternatively, the acquisition of a new role for this gene. PMID:26773809

  9. Medicago truncatula Mtha1-2 mutants loose metabolic responses to mycorrhizal colonization.

    PubMed

    Hubberten, Hans-Michael; Sieh, Daniela; Zöller, Daniela; Hoefgen, Rainer; Krajinski, Franziska

    2015-01-01

    Bidirectional nutrient transfer is one of the key features of the arbuscular mycorrhizal symbiosis. Recently we were able to identify a Medicago truncatula mutant (mtha1-2) that is defective in the uptake of phosphate from the periarbuscular space due to a lack of the energy providing proton gradient provided by the symbiosis specific proton ATPase MtHA1 In order to further characterize the impact of fungal colonization on the plant metabolic status, without the beneficial aspect of improved mineral nutrition, we performed leaf ion analyses in mutant and wildtype plants with and without fungal colonization. Although frequency of fungal colonization was unaltered, the mutant did not show a positive growth response to mycorrhizal colonization. This indicates that nutrient transfer into the plant cell fails in the truncated arbuscules due to lacking expression of a functional MtHA1 protein. The leaves of wildtype plants showed clear metabolic responses to root mycorrhizal colonization, whereas no changes of leaf metabolite levels of mycorrhizal mtha1-2 plants were detected, even though they were colonized. These results show that MtHa1 is indispensable for a functional mycorrhizal symbiosis and, moreover, suggest that fungal root colonization per se does not depend on nutrient transfer to the plant host. PMID:25751449

  10. [Influence of salt stress on the genetically polymorphic system of Sinorhizobium meliloti-Medicago truncatula].

    PubMed

    Kurchak, O N; Provorov, N A; Onishchuk, O P; Vorobyov, N I; Roumiantseva, M L; Simarov, B V

    2014-07-01

    The impacts of salt stress (75 mM NaC1) on the ecological efficiency of the genetically polymorphic Sinorhizobium meliloti-Medicago truncatula system were studied. Its impact on a symbiotic system results in an increase of the partners' variability for symbiotic traits and of the symbiosis integrity as indicated by: a) the specificity of the partners' interactions--the nonadditive inputs of their genotypes into the variation of symbiotic parameters; and b) the correlative links between these parameters. The structure of the nodDI locus and the content correlates to the efficiency of the symbiosis between S. meliloti and M. truncatula genotypes under stress conditions more sufficiently than in the absence of stress. Correlations between the symbiotic efficiency of rhizobia strains and their growth rate outside symbiosis are expressed under stress conditions, not in the absence of stress. Under salt stress symbiotic effectiveness was decreased for M. truncatula line F83005.5, which was salt sensitive for mineral nutrition. The inhibition of symbiotic activity for this line is linked with decreased nodule formation, whereas for Jemalong 6 and DZA315.16 lines it is associated with repressed N2-fixation. It was demonstrated for the first time that salt stress impairs the M. truncatula habitus (the mass : height ratio increased 2- to 6-fold), which in the salt-resistant cultivar Jemalong 6 is normalized as the result of rhizobia inoculation.

  11. Enhanced salt tolerance of alfalfa (Medicago sativa) by rstB gene transformation.

    PubMed

    Zhang, Wan-Jun; Wang, Tao

    2015-05-01

    Generating salt tolerance forage plant is essential for use of the land affected by high salinity. A salt tolerance gene rstB was used as a selectable marker gene in Agrobacterium-mediated transformation of tobacco under a selective regime of 170mM NaCl. The transgenic plants showed clear improvement in salt tolerance. To improve salt tolerance of alfalfa (Medicago sativa L.), rstB gene was introduced into alfalfa genome by Agrobacterium-mediated transformation. No abnormal phenotype was observed among the transgenic plants when compared with wild type (wt) plants. Significant enhancement of resistance to salt-shock treatment was noted on the rstB transgenic (T0) plants. Transgenic second-generation (T1) seeds showed improved germination rate and seedling growth under salt-stress condition. Hindered Na(+) accumulation, but enhanced Ca(2+) accumulation was observed on the rstB T1 plants when subjected to salt-stresses. Enhanced calcium accumulation in transgenic plants was also verified by cytohistochemical localization of calcium. Under salt-stress of 50mM NaCl, about 15% of the transgenic plants finished their life-cycle but the wt plants had no flower formation. The results demonstrated that the expression of rstB gene improved salt tolerance in transgenic alfalfa.

  12. Concerted changes in N and C primary metabolism in alfalfa (Medicago sativa) under water restriction.

    PubMed

    Aranjuelo, Iker; Tcherkez, Guillaume; Molero, Gemma; Gilard, Françoise; Avice, Jean-Christophe; Nogués, Salvador

    2013-02-01

    Although the mechanisms of nodule N(2) fixation in legumes are now well documented, some uncertainty remains on the metabolic consequences of water deficit. In most cases, little consideration is given to other organs and, therefore, the coordinated changes in metabolism in leaves, roots, and nodules are not well known. Here, the effect of water restriction on exclusively N(2)-fixing alfalfa (Medicago sativa L.) plants was investigated, and proteomic, metabolomic, and physiological analyses were carried out. It is shown that the inhibition of nitrogenase activity caused by water restriction was accompanied by concerted alterations in metabolic pathways in nodules, leaves, and roots. The data suggest that nodule metabolism and metabolic exchange between plant organs nearly reached homeostasis in asparagine synthesis and partitioning, as well as the N demand from leaves. Typically, there was (i) a stimulation of the anaplerotic pathway to sustain the provision of C skeletons for amino acid (e.g. glutamate and proline) synthesis; (ii) re-allocation of glycolytic products to alanine and serine/glycine; and (iii) subtle changes in redox metabolites suggesting the implication of a slight oxidative stress. Furthermore, water restriction caused little change in both photosynthetic efficiency and respiratory cost of N(2) fixation by nodules. In other words, the results suggest that under water stress, nodule metabolism follows a compromise between physiological imperatives (N demand, oxidative stress) and the lower input to sustain catabolism.

  13. Accumulation and residue of napropamide in alfalfa (Medicago sativa) and soil involved in toxic response.

    PubMed

    Cui, Li E; Yang, Hong

    2011-06-15

    Napropamide belongs to the amide herbicide family and widely used to control weeds in farmland. Intensive use of the herbicide has resulted in widespread contamination to ecosystems. The present study demonstrated an analysis on accumulation of the toxic pesticide napropamide in six genotypes of alfalfa (Medicago sativa), along with biological parameters and its residues in soils. Soil was treated with napropamide at 3 mg kg(-1) dry soil and alfalfa plants were cultured for 10 or 30 d, respectively. The maximum value for napropamide accumulation is 0.426 mg kg(-1) in shoots and 2.444 mg kg(-1) in roots. The napropamide-contaminated soil with alfalfa cultivation had much lower napropamide concentrations than the control (soil without alfalfa cultivation). Also, the content of napropamide residue in the rhizosphere was significantly lower than that in the non-rhizosphere soil. M. sativa exposed to 3 mg kg(-1) napropamide showed inhibited growth. Further analysis revealed that plants treated with napropamide accumulated more reactive oxygen species (O(2)(-) and H(2)O(2)) and less amounts of chlorophyll. However, not all cultivars showed oxidative injury, suggesting that the alfalfa cultivars display different tolerance to napropamide. PMID:21439724

  14. The Effects of Heavy Metals and Root Immersion on Isoflavonoid Metabolism in Alfalfa (Medicago sativa L.).

    PubMed Central

    Parry, A. D.; Tiller, S. A.; Edwards, R.

    1994-01-01

    Modest increases in the concentration of medicarpin, 6-fold in leaves and 4-fold in roots, were observed in alfalfa (Medicago sativa L.) seedlings treated with 1 mM metal salts for 72 h. However, medicarpin-3-O-glucoside-6"-O-malonate (MGM) and formononetin-7-O-glucoside-6"-O-malonate (FGM) levels were up to 50-fold lower in metal-treated compared to control roots. Approximately 10% of the "missing" conjugates could be accounted for in the root treatment solution, where FGM and MGM transiently accumulated prior to their hydrolysis. Time-course studies revealed that total isoflavonoid content (roots plus solution) increased slightly after CuCl2 treatment, whereas the levels of FGM and MGM increased rapidly in alfalfa roots immersed in water. This increase was reduced by aeration. The phenylalanine ammonia-lyase inhibitor L-[alpha]-aminooxy-[beta]-phenylpropionic acid was used to show that immersion of the roots reduced conjugate rates of degradation, which explains their accumulation. In contrast, conjugate rates of degradation were elevated in CuCl2-treated roots, with 50% of the increase being due to hydrolysis. Up to 90% of formononetin and medicarpin produced in response to CuCl2 treatment arose via conjugate hydrolysis. Our results demonstrate that both immersion/anaerobiosis and abiotic elicitation modify isoflavonoid metabolism in alfalfa, and that metal-stimulated accumulation of phytoalexins may arise through the release from preformed stores rather than de novo synthesis. PMID:12232319

  15. Association Mapping for Fiber-Related Traits and Digestibility in Alfalfa (Medicago sativa)

    PubMed Central

    Wang, Zan; Qiang, Haiping; Zhao, Haiming; Xu, Ruixuan; Zhang, Zhengli; Gao, Hongwen; Wang, Xuemin; Liu, Guibo; Zhang, Yingjun

    2016-01-01

    Association mapping is a powerful approach for exploring the molecular genetic basis of complex quantitative traits. An alfalfa (Medicago sativa) association panel comprised of 336 genotypes from 75 alfalfa accessions represented by four to eight genotypes for each accession. Each genotype was genotyped using 85 simple sequence repeat (SSR) markers and phenotyped for five fiber-related traits in four different environments. A model-based structure analysis was used to group all genotypes into two groups. Most of the genotypes have a low relative kinship (<0.3), suggesting population stratification not be an issue for association analysis. Generally, the Q + K model exhibited the best performance to eliminate the false associated positives. In total, 124 marker-trait associations were predicted (p < 0.005). Among these, eight associations were predicted in two environments repeatedly and 20 markers were predicted to be associated with multiple traits. These trait-associated markers will greatly help marker-assisted breeding programs to improve fiber-related quality traits in alfalfa. PMID:27047512

  16. Functional specialization of duplicated AP3-like genes in Medicago truncatula.

    PubMed

    Roque, Edelín; Serwatowska, Joanna; Cruz Rochina, M; Wen, Jiangqi; Mysore, Kirankumar S; Yenush, Lynne; Beltrán, José Pío; Cañas, Luis A

    2013-02-01

    The B-class of MADS box genes has been studied in a wide range of plant species, but has remained largely uncharacterized in legumes. Here we investigate the evolutionary fate of the duplicated AP3-like genes of a legume species. To obtain insight into the extent to which B-class MADS box gene functions are conserved or have diversified in legumes, we isolated and characterized the two members of the AP3 lineage in Medicago truncatula: MtNMH7 and MtTM6 (euAP3 and paleoAP3 genes, respectively). A non-overlapping and complementary expression pattern of both genes was observed in petals and stamens. MtTM6 was expressed predominantly in the outer cell layers of both floral organs, and MtNMH7 in the inner cell layers of petals and stamens. Functional analyses by reverse genetics approaches (RNAi and Tnt1 mutagenesis) showed that the contribution of MtNMH7 to petal identity is more important than that of MtTM6, whereas MtTM6 plays a more important role in stamen identity than its paralog MtNMH7. Our results suggest that the M. truncatula AP3-like genes have undergone a functional specialization process associated with complete partitioning of gene expression patterns of the ancestral gene lineage. We provide information regarding the similarities and differences in petal and stamen development among core eudicots.

  17. Evolution by gene duplication of Medicago truncatula PISTILLATA-like transcription factors.

    PubMed

    Roque, Edelín; Fares, Mario A; Yenush, Lynne; Rochina, Mari Cruz; Wen, Jiangqi; Mysore, Kirankumar S; Gómez-Mena, Concepción; Beltrán, José Pío; Cañas, Luis A

    2016-03-01

    PISTILLATA (PI) is a member of the B-function MADS-box gene family, which controls the identity of both petals and stamens in Arabidopsis thaliana. In Medicago truncatula (Mt), there are two PI-like paralogs, known as MtPI and MtNGL9. These genes differ in their expression patterns, but it is not known whether their functions have also diverged. Describing the evolution of certain duplicated genes, such as transcription factors, remains a challenge owing to the complex expression patterns and functional divergence between the gene copies. Here, we report a number of functional studies, including analyses of gene expression, protein-protein interactions, and reverse genetic approaches designed to demonstrate the respective contributions of each M. truncatula PI-like paralog to the B-function in this species. Also, we have integrated molecular evolution approaches to determine the mode of evolution of Mt PI-like genes after duplication. Our results demonstrate that MtPI functions as a master regulator of B-function in M. truncatula, maintaining the overall ancestral function, while MtNGL9 does not seem to have a role in this regard, suggesting that the pseudogenization could be the functional evolutionary fate for this gene. However, we provide evidence that purifying selection is the primary evolutionary force acting on this paralog, pinpointing the conservation of its biochemical function and, alternatively, the acquisition of a new role for this gene.

  18. Effect of salt stress on glycine betaine biosynthesis and catabolism by Medicago sativa bacteroids

    SciTech Connect

    Fougere, F.; Poggi, M.-C.; Le Rudulier, D. )

    1990-05-01

    Previous works have shown that glycine betaine (GB) and choline (Cho) are actively taken up by Medicago sativa bacteroids isolated from 4-week-old nodules. Here, we have investigated the effects of NaCl on the fte of Cho and GB. Bacteroids were incubated in low- or high-salt-medium (0.4 M NaCl) and supplemented with {sup 14}C 1,2-Cho or {sup 14}C 1,2-GB. After 3 hours, radioactivity was measured in CO{sub 2} released, in ethanol-soluble and insoluble fractions. In absence of salt, a low proportion of the labelling was found in soluble fraction: 47 and 19% after Cho or GB supply, respectively. On the contrary, in high-salt-medium, the soluble fraction still contained 85% of the radioactivity with GB corresponding to 92-98%. Both enzymes involved in GB biosynthesis from Cho were studied. Choline oxidase activity was enhanced by 59%, while betainal dehydrogenase activity remained unchanged after bacteroid incubation in high-salt-medium. Thus, GB accumulation in salt-stressed bacteroids would be likely a consequence of a decrease of its catabolism rather than an increase of its biosynthesis.

  19. (Homo)glutathione Deficiency Impairs Root-knot Nematode Development in Medicago truncatula

    PubMed Central

    Baldacci-Cresp, Fabien; Chang, Christine; Maucourt, Mickaël; Deborde, Catherine; Hopkins, Julie; Lecomte, Philippe; Bernillon, Stéphane; Brouquisse, Renaud; Moing, Annick; Abad, Pierre; Hérouart, Didier; Puppo, Alain; Favery, Bruno; Frendo, Pierre

    2012-01-01

    Root-knot nematodes (RKN) are obligatory plant parasitic worms that establish and maintain an intimate relationship with their host plants. During a compatible interaction, RKN induce the redifferentiation of root cells into multinucleate and hypertrophied giant cells essential for nematode growth and reproduction. These metabolically active feeding cells constitute the exclusive source of nutrients for the nematode. Detailed analysis of glutathione (GSH) and homoglutathione (hGSH) metabolism demonstrated the importance of these compounds for the success of nematode infection in Medicago truncatula. We reported quantification of GSH and hGSH and gene expression analysis showing that (h)GSH metabolism in neoformed gall organs differs from that in uninfected roots. Depletion of (h)GSH content impaired nematode egg mass formation and modified the sex ratio. In addition, gene expression and metabolomic analyses showed a substantial modification of starch and γ-aminobutyrate metabolism and of malate and glucose content in (h)GSH-depleted galls. Interestingly, these modifications did not occur in (h)GSH-depleted roots. These various results suggest that (h)GSH have a key role in the regulation of giant cell metabolism. The discovery of these specific plant regulatory elements could lead to the development of new pest management strategies against nematodes. PMID:22241996

  20. Molecular Signals Controlling the Inhibition of Nodulation by Nitrate in Medicago truncatula

    PubMed Central

    van Noorden, Giel E.; Verbeek, Rob; Dinh, Quy Dung; Jin, Jian; Green, Alexandra; Ng, Jason Liang Pin; Mathesius, Ulrike

    2016-01-01

    The presence of nitrogen inhibits legume nodule formation, but the mechanism of this inhibition is poorly understood. We found that 2.5 mM nitrate and above significantly inhibited nodule initiation but not root hair curling in Medicago trunatula. We analyzed protein abundance in M. truncatula roots after treatment with either 0 or 2.5 mM nitrate in the presence or absence of its symbiont Sinorhizobium meliloti after 1, 2 and 5 days following inoculation. Two-dimensional gel electrophoresis combined with mass spectrometry was used to identify 106 differentially accumulated proteins responding to nitrate addition, inoculation or time point. While flavonoid-related proteins were less abundant in the presence of nitrate, addition of Nod gene-inducing flavonoids to the Sinorhizobium culture did not rescue nodulation. Accumulation of auxin in response to rhizobia, which is also controlled by flavonoids, still occurred in the presence of nitrate, but did not localize to a nodule initiation site. Several of the changes included defense- and redox-related proteins, and visualization of reactive oxygen species indicated that their induction in root hairs following Sinorhizobium inoculation was inhibited by nitrate. In summary, the presence of nitrate appears to inhibit nodulation via multiple pathways, including changes to flavonoid metabolism, defense responses and redox changes. PMID:27384556

  1. Concerted changes in N and C primary metabolism in alfalfa (Medicago sativa) under water restriction

    PubMed Central

    Aranjuelo, Iker

    2013-01-01

    Although the mechanisms of nodule N2 fixation in legumes are now well documented, some uncertainty remains on the metabolic consequences of water deficit. In most cases, little consideration is given to other organs and, therefore, the coordinated changes in metabolism in leaves, roots, and nodules are not well known. Here, the effect of water restriction on exclusively N2-fixing alfalfa (Medicago sativa L.) plants was investigated, and proteomic, metabolomic, and physiological analyses were carried out. It is shown that the inhibition of nitrogenase activity caused by water restriction was accompanied by concerted alterations in metabolic pathways in nodules, leaves, and roots. The data suggest that nodule metabolism and metabolic exchange between plant organs nearly reached homeostasis in asparagine synthesis and partitioning, as well as the N demand from leaves. Typically, there was (i) a stimulation of the anaplerotic pathway to sustain the provision of C skeletons for amino acid (e.g. glutamate and proline) synthesis; (ii) re-allocation of glycolytic products to alanine and serine/glycine; and (iii) subtle changes in redox metabolites suggesting the implication of a slight oxidative stress. Furthermore, water restriction caused little change in both photosynthetic efficiency and respiratory cost of N2 fixation by nodules. In other words, the results suggest that under water stress, nodule metabolism follows a compromise between physiological imperatives (N demand, oxidative stress) and the lower input to sustain catabolism. PMID:23440170

  2. [Determination of Hard Rate of Alfalfa (Medicago sativa L.) Seeds with Near Infrared Spectroscopy].

    PubMed

    Wang, Xin-xun; Chen, Ling-ling; Zhang, Yun-wei; Mao, Pei-sheng

    2016-03-01

    Alfalfa (Medicago sativa L.) is the most commonly grown forage crop due to its better quality characteristics and high adaptability in China. However, there was 20%-80% hard seeds in alfalfa which could not be identified easily from non hard seeds which would cause the loss of seed utilization value and plant production. This experiment was designed for 121 samples of alfalfa. Seeds were collected according to different regions, harvested year and varieties. 31 samples were artificial matched as hard rates ranging from 20% to 80% to establish a model for hard seed rate by near infrared spectroscopy (NIRS) with Partial Least Square (PLS). The objective of this study was to establish a model and to estimate the efficiency of NIRS for determining hard rate of alfalfa seeds. The results showed that the correlation coefficient (R2(cal)) of calibration model was 0.981 6, root mean square error of cross validation (RMSECV) was 5.32, and the ratio of prediction to deviation (RPD) was 3.58. The forecast model in this experiment presented the satisfied precision. The proposed method using NIRS technology is feasible for identification and classification of hard seed in alfalfa. A new method, as nondestructive testing of hard seed rate, was provided to theoretical basis for fast nondestructive detection of hard seed rates in alfalfa.

  3. Correlations between environmental factors and wild bee behavior on alfalfa (Medicago sativa) in northwestern China.

    PubMed

    Wang, Xiaojuan; Liu, Hongping; Li, Xiaoxia; Song, Yu; Chen, Li; Jin, Liang

    2009-10-01

    To discover the effect of environmental factors on pollinator visitation to flowering Medicago sativa, several field experiments were designed to examine the diurnal movement patterns of wild bee species in the Hexi Corridor of northwestern China. Our study results showed that Megachile abluta, M. spissula, and Xylocopa valga showed unimodal diurnal foraging behavior, whereas Andrena parvula and Anthophora melanognatha showed bimodal diurnal foraging behavior. Correlation analysis indicated that diurnal foraging activities of pollinators were significantly correlated with environmental factors. Correlations of foraging activities versus environmental factors for M. abluta, M. spissula, and X. valga best fit a linear model, whereas those of A. parvula and A. melanognatha best fit a parallel quadratic model. Results of this study indicated that solitary wild bees such as M. abluta, M. spissula, X. valga, A. parvula, and A. melanognatha are potential alfalfa pollinators in the Hexi Corridor. An understanding of the environmental factors that affect the behaviors of different wild bees foraging in alfalfa are basic to the utilization of solitary wild bees in a practical way for increased, or more consistent, pollination of alfalfa for seed production. PMID:19825303

  4. Stoichiometric Characteristics of Carbon, Nitrogen, and Phosphorus in Leaves of Differently Aged Lucerne (Medicago sativa) Stands

    PubMed Central

    Wang, Zhennan; Lu, Jiaoyun; Yang, Mei; Yang, Huimin; Zhang, Qingping

    2015-01-01

    Element concentration within a plant which is vital to function maintenance and adaptation to environment, may change with plant growth. However, how carbon (C), nitrogen (N), and phosphorus (P) vary stoichiometrically with stand growth, i.e., ages or cuts, was still untouched in perennial species. This study tested the hypothesis that lucerne (Medicago sativa) C:N, C:P, and N:P should change with stand age and cut. Leaf C:N, C:P, and N:P changed with stand age, showing various trends in different cuts of lucerne. Generally the greatest stoichiometric ratios were measured in 8 year stand and in the second cut. They were affected significantly and negatively by total N and P concentrations of leaf, but not by organic C concentration. There were significantly positive correlations among leaf C:N, C:P, and N:P. However, leaf C:N, C:P, and N:P were hardly affected by soil features. Conclusively, lucerne C, N, and P stoichiometry are age- and cut-specific, and regulated mainly by leaf N, P concentrations and stoichiometry. There are few correlations with soil fertility. To our knowledge, it is the first try to elucidate the stoichiometry in the viewpoint of age and cut with a perennial herbaceous legume. PMID:26697029

  5. Micromonospora from nitrogen fixing nodules of alfalfa (Medicago sativa L.). A new promising Plant Probiotic Bacteria.

    PubMed Central

    Martínez-Hidalgo, Pilar; Galindo-Villardón, Purificación; Igual, José M.; Martínez-Molina, Eustoquio

    2014-01-01

    Biotic interactions can improve agricultural productivity without costly and environmentally challenging inputs. Micromonospora strains have recently been reported as natural endophytes of legume nodules but their significance for plant development and productivity has not yet been established. The aim of this study was to determine the diversity and function of Micromonospora isolated from Medicago sativa root nodules. Micromonospora-like strains from field alfalfa nodules were characterized by BOX-PCR fingerprinting and 16S rRNA gene sequencing. The ecological role of the interaction of the 15 selected representative Micromonospora strains was tested in M. sativa. Nodulation, plant growth and nutrition parameters were analyzed. Alfalfa nodules naturally contain abundant and highly diverse populations of Micromonospora, both at the intra- and at interspecific level. Selected Micromonospora isolates significantly increase the nodulation of alfalfa by Ensifer meliloti 1021 and also the efficiency of the plant for nitrogen nutrition. Moreover, they promote aerial growth, the shoot-to-root ratio, and raise the level of essential nutrients. Our results indicate that Micromonospora acts as a Rhizobia Helper Bacteria (RHB) agent and has probiotic effects, promoting plant growth and increasing nutrition efficiency. Its ecological role, biotechnological potential and advantages as a plant probiotic bacterium (PPB) are also discussed. PMID:25227415

  6. Evaluation of some weed extracts against field dodder on alfalfa (Medicago sativa).

    PubMed

    Habib, S A; Rahman, A A

    1988-02-01

    Experiments were conducted to evaluate the effectiveness of some weed extracts against field dodderCuscuta campestris Yunck. on alfalfa (Medicago stiva L.) and to identify and quantify the phytotoxic agents of these extracts. All concentrations of aqueous extract of every weed showed significant effectiveness on dodder when compared to the untreated plant under lath house and field conditions. Control percentages of the (0.5 g) of Bermuda grass (Cynodon dactylon) and wall goosefoot (Chenopodium murale) ranged between 83 and 96, and the same concentration caused injury to alfalfa foliage up to 43% when applied in the field. Phytotoxic agents were identified as phenolic compounds such as chlorogenic, isochlorogenic,p-coumaric acids, and scopoletin. Their quantities varied with species; Bermuda grass had the highest content (32.2 μg/g dry weight) followed by Johnson grass (Sorghum halepense), well goosefoot, and tumble pigweed (Amaranthus albus). These results might aid in screening for effective alternate approaches for controlling dodder on alfalfa planted for seeds.

  7. Chemical modification and degradation of atrazine in Medicago sativa through multiple pathways.

    PubMed

    Zhang, Jing Jing; Lu, Yi Chen; Yang, Hong

    2014-10-01

    Atrazine is a member of the triazine herbicide family intensively used to control weeds for crop production. In this study, atrazine residues and its degraded products in alfalfa (Medicago sativa) were characterized using UPLC-TOF-MS/MS. Most of atrazine absorbed in plants was found as chemically modified derivatives like deisopropylated atrazine (DIA), dehydrogenated atrazine (DHA), or methylated atrazine (MEA), and some atrazine derivatives were conjugated through different functional groups such as sugar, glutathione, and amino acids. Interestingly, the specific conjugates DHA+hGSH (homoglutathione) and MEA-HCl+hGSH in alfalfa were detected. These results suggest that atrazine in alfalfa can be degraded through different pathways. The increased activities of glycosyltransferase and glutathione S-transferase were determined to support the atrazine degradation models. The outcome of the work uncovered the detailed mechanism for the residual atrazine accumulation and degradation in alfalfa and will help to evaluate whether the crop is suitable to be cultivated in the atrazine-polluted soil.

  8. Transcriptional and post-transcriptional regulation of a NAC1 transcription factor in Medicago truncatula roots.

    PubMed

    D'haeseleer, Katrien; Den Herder, Griet; Laffont, Carole; Plet, Julie; Mortier, Virginie; Lelandais-Brière, Christine; De Bodt, Stefanie; De Keyser, Annick; Crespi, Martin; Holsters, Marcelle; Frugier, Florian; Goormachtig, Sofie

    2011-08-01

    • Legume roots develop two types of lateral organs, lateral roots and nodules. Nodules develop as a result of a symbiotic interaction with rhizobia and provide a niche for the bacteria to fix atmospheric nitrogen for the plant. • The Arabidopsis NAC1 transcription factor is involved in lateral root formation, and is regulated post-transcriptionally by miRNA164 and by SINAT5-dependent ubiquitination. We analyzed in Medicago truncatula the role of the closest NAC1 homolog in lateral root formation and in nodulation. • MtNAC1 shows a different expression pattern in response to auxin than its Arabidopsis homolog and no changes in lateral root number or nodulation were observed in plants affected in MtNAC1 expression. In addition, no interaction was found with SINA E3 ligases, suggesting that post-translational regulation of MtNAC1 does not occur in M. truncatula. Similar to what was found in Arabidopsis, a conserved miR164 target site was retrieved in MtNAC1, which reduced protein accumulation of a GFP-miR164 sensor. Furthermore, miR164 and MtNAC1 show an overlapping expression pattern in symbiotic nodules, and overexpression of this miRNA led to a reduction in nodule number. • This work suggests that regulatory pathways controlling a conserved transcription factor are complex and divergent between M. truncatula and Arabidopsis.

  9. Evaluation of Medicago sativa L. sprouts as antihyperlipidemic and antihyperglycemic agent.

    PubMed

    Seida, Ahmed; El-Hefnawy, Hala; Abou-Hussein, Dina; Mokhtar, Fatma Alzahraa; Abdel-Naim, Ashraf

    2015-11-01

    Medicago sativa L. (Alfalfa) is traditionally used to treat diabetes. This study was designed to investigate the potential antihyperlipidemic and antihyperglycemic activity of M. sativa sprouts in streptozotocin (STZ) induced diabetes via i.p. injection of 55 mg/kg of STZ. Experimental animals were divided into the following groups: GP1 (normal), GP2 (STZ-hyperlipidemic), GP3 (rouvastatin), GP4 (metformin), GP 5-9 (diabetic treated with methanolic, petroleum ether, chloroform, ethyl acetate and butanol extracts). The administration of the total methanolic extract (500 mg/kg), the petroleum ether (32.5mg) and butanol fractions (60 mg) for 4 weeks significantly decreased (p<0.05) triglycerides (TG), total cholesterol (TC), low-density lipoproteins (LDL) and very low density lipoproteins (VLDL) in comparison to rouvastatin. Petroleum ether fraction proved to exhibit the best activity as antihyperlipidemic agent (12.23%). On the other hand, ethyl acetate fraction retained the best activity (vs. metformin) as antihyperglycemic agent. Histopathological evidences on liver, pancreas and spleen were in agreement with the above mentioned results. Purification, characterization, and identification of isolated compounds from the active fractions afforded 9 compounds: β-sitosterol and stigmasterol from the petroleum ether fraction; 10-hydroxy-coumestrol, apigenin, genistein, p-hydroxy-benzoic-acid, 7, 4'- dihydroxyflavone, quercetin-3-glucoside and sissotrin from the ethyl acetate fraction. PMID:26639479

  10. Selective lignin downregulation leads to constitutive defense response expression in alfalfa (Medicago sativa L.).

    PubMed

    Gallego-Giraldo, Lina; Jikumaru, Yusuke; Kamiya, Yuji; Tang, Yuhong; Dixon, Richard A

    2011-05-01

    • Downregulation of hydroxycinnamoyl CoA: shikimate hydroxycinnamoyl transferase (HCT) in alfalfa (Medicago sativa) reduces lignin levels and improves forage quality and saccharification efficiency for bioethanol production. However, the plants have reduced stature. It was previously reported that HCT-down-regulated Arabidopsis have impaired auxin transport, but this has recently been disproved. • To address the basis for the phenotypes of lignin-modified alfalfa, we measured auxin transport, profiled a range of metabolites including flavonoids and hormones, and performed in depth transcriptome analyses. • Auxin transport is unaffected in HCT antisense alfalfa despite increased flavonoid biosynthesis. The plants show increased cytokinin and reduced auxin levels, and gibberellin levels and sensitivity are both reduced. Levels of salicylic, jasmonic and abscisic acids are elevated, associated with massive upregulation of pathogenesis and abiotic stress-related genes and enhanced tolerance to fungal infection and drought. • We suggest that HCT downregulated alfalfa plants exhibit constitutive activation of defense responses, triggered by release of bioactive cell wall fragments and production of hydrogen peroxide as a result of impaired secondary cell wall integrity.

  11. Expression Analysis of PIN Genes in Root Tips and Nodules of Medicago truncatula.

    PubMed

    Sańko-Sawczenko, Izabela; Łotocka, Barbara; Czarnocka, Weronika

    2016-01-01

    Polar auxin transport is dependent on the family of PIN-formed proteins (PINs), which are membrane transporters of anionic indole-3-acetic acid (IAA(-)). It is assumed that polar auxin transport may be essential in the development and meristematic activity maintenance of Medicago truncatula (M. truncatula) root nodules. However, little is known about the involvement of specific PIN proteins in M. truncatula nodulation. Using real-time quantitative PCR, we analyzed the expression patterns of all previously identified MtPIN genes and compared them between root nodules and root tips of M. truncatula. Our results demonstrated significant differences in the expression level of all 11 genes (MtPIN1-MtPIN11) between examined organs. Interestingly, MtPIN9 was the only PIN gene with higher expression level in root nodules compared to root tips. This result is the first indication of PIN9 transporter potential involvement in M. truncatula nodulation. Moreover, relatively high expression level in root nodules was attributed to MtPINs encoding orthologs of Arabidopsis thaliana PIN5 subclade. PIN proteins from this subclade have been found to localize in the endoplasmic reticulum, which may indicate that the development and meristematic activity maintenance of M. truncatula root nodules is associated with intracellular homeostasis of auxins level and their metabolism in the endoplasmic reticulum. PMID:27463709

  12. TE7, An Inefficient Symbiotic Mutant of Medicago truncatula Gaertn. cv Jemalong.

    PubMed Central

    Benaben, V.; Duc, G.; Lefebvre, V.; Huguet, T.

    1995-01-01

    A mutagenesis program using ethylmethane sulfonate on Medicago truncatula Gaertn cv Jemalong, an annual, autogamous and diploid lucerne, permitted the isolation of a mutant (TE7) unable to establish an effective nitrogen-fixing symbiosis, [Nod+Fix-], with Rhizobium meliloti wild-type strains. The mutant phenotype is characterized by an altered infection process that leads to the formation of two kinds of inefficient nodules on the same root system. A certain proportion of the nodules are small, round, and uninfected, with infection threads limited to the outer root cortical cells. Others develop to a normal elongated shape and are infected; bacterial release occurs but the bacteria do not differentiate into bacteroids. The ratio of invaded to uninvaded nodules depends on the bacterial strain used. Throughout the infection process, certain events correlated with the plant defense response against pathogens can be observed: (a) the presence of polyphenolic compounds associated with the walls of infected cells and also with some parts of infection threads in the root cortex; (b) appositions on infection thread walls during the early stage of infection and also within the central tissue of infected nodules; and (c) autophagy of the plant cells that contain released bacteria. Genetic data suggest that the phenotype of TE7 is under monogenic and recessive control; this gene has been designated Mtsym1. PMID:12228341

  13. Medicago sativa--Sinorhizobium meliloti Symbiosis Promotes the Bioaccumulation of Zinc in Nodulated Roots.

    PubMed

    Zribi, Kais; Nouairi, Issam; Slama, Ines; Talbi-Zribi, Ons; Mhadhbi, Haythem

    2015-01-01

    In this study we investigated effects of Zn supply on germination, growth, inorganic solutes (Zn, Ca, Fe, and Mg) partitioning and nodulation of Medicago sativa This plant was cultivated with and without Zn (2 mM). Treatments were plants without (control) and with Zn tolerant strain (S532), Zn intolerant strain (S112) and 2 mM urea nitrogen fertilisation. Results showed that M. sativa germinates at rates of 50% at 2 mM Zn. For plants given nitrogen fertilisation, Zn increased plant biomass production. When grown with symbionts, Zn supply had no effect on nodulation. Moreover, plants with S112 showed a decrease of shoot and roots biomasses. However, in symbiosis with S532, an increase of roots biomass was observed. Plants in symbiosis with S. meliloti accumulated more Zn in their roots than nitrogen fertilised plants. Zn supply results in an increase of Ca concentration in roots of fertilised nitrogen plants. However, under Zn supply, Fe concentration decreased in roots and increased in nodules of plants with S112. Zn supply showed contrasting effects on Mg concentrations for plants with nitrogen fertilisation (increase) and plants with S112 (decrease). The capacity of M. sativa to accumulate Zn in their nodulated roots encouraged its use in phytostabilisation processes.

  14. MtSWEET11, a Nodule-Specific Sucrose Transporter of Medicago truncatula.

    PubMed

    Kryvoruchko, Igor S; Sinharoy, Senjuti; Torres-Jerez, Ivone; Sosso, Davide; Pislariu, Catalina I; Guan, Dian; Murray, Jeremy; Benedito, Vagner A; Frommer, Wolf B; Udvardi, Michael K

    2016-05-01

    Optimization of nitrogen fixation by rhizobia in legumes is a key area of research for sustainable agriculture. Symbiotic nitrogen fixation (SNF) occurs in specialized organs called nodules and depends on a steady supply of carbon to both plant and bacterial cells. Here we report the functional characterization of a nodule-specific Suc transporter, MtSWEET11 from Medicago truncatula MtSWEET11 belongs to a clade of plant SWEET proteins that are capable of transporting Suc and play critical roles in pathogen susceptibility. When expressed in mammalian cells, MtSWEET11 transported sucrose (Suc) but not glucose (Glc). The MtSWEET11 gene was found to be expressed in infected root hair cells, and in the meristem, invasion zone, and vasculature of nodules. Expression of an MtSWEET11-GFP fusion protein in nodules resulted in green fluorescence associated with the plasma membrane of uninfected cells and infection thread and symbiosome membranes of infected cells. Two independent Tnt1-insertion sweet11 mutants were uncompromised in SNF Therefore, although MtSWEET11 appears to be involved in Suc distribution within nodules, it is not crucial for SNF, probably because other Suc transporters can fulfill its role(s). PMID:27021190

  15. Mixed Nodule Infection in Sinorhizobium meliloti-Medicago sativa Symbiosis Suggest the Presence of Cheating Behavior.

    PubMed

    Checcucci, Alice; Azzarello, Elisa; Bazzicalupo, Marco; Galardini, Marco; Lagomarsino, Alessandra; Mancuso, Stefano; Marti, Lucia; Marzano, Maria C; Mocali, Stefano; Squartini, Andrea; Zanardo, Marina; Mengoni, Alessio

    2016-01-01

    In the symbiosis between rhizobia and legumes, host plants can form symbiotic root nodules with multiple rhizobial strains, potentially showing different symbiotic performances in nitrogen fixation. Here, we investigated the presence of mixed nodules, containing rhizobia with different degrees of mutualisms, and evaluate their relative fitness in the Sinorhizobium meliloti-Medicago sativa model symbiosis. We used three S. meliloti strains, the mutualist strains Rm1021 and BL225C and the non-mutualist AK83. We performed competition experiments involving both in vitro and in vivo symbiotic assays with M. sativa host plants. We show the occurrence of a high number (from 27 to 100%) of mixed nodules with no negative effect on both nitrogen fixation and plant growth. The estimation of the relative fitness as non-mutualist/mutualist ratios in single nodules shows that in some nodules the non-mutualist strain efficiently colonized root nodules along with the mutualist ones. In conclusion, we can support the hypothesis that in S. meliloti-M. sativa symbiosis mixed nodules are formed and allow non-mutualist or less-mutualist bacterial partners to be less or not sanctioned by the host plant, hence allowing a potential form of cheating behavior to be present in the nitrogen fixing symbiosis.

  16. Nodule carbohydrate catabolism is enhanced in the Medicago truncatula A17-Sinorhizobium medicae WSM419 symbiosis

    PubMed Central

    Larrainzar, Estíbaliz; Gil-Quintana, Erena; Seminario, Amaia; Arrese-Igor, Cesar; González, Esther M.

    2014-01-01

    The symbiotic association between Medicago truncatula and Sinorhizobium meliloti is a well-established model system in the legume–Rhizobium community. Despite its wide use, the symbiotic efficiency of this model has been recently questioned and an alternative microsymbiont, S. medicae, has been proposed. However, little is known about the physiological mechanisms behind the higher symbiotic efficiency of S. medicae WSM419. In the present study, we inoculated M. truncatula Jemalong A17 with either S. medicae WSM419 or S. meliloti 2011 and compared plant growth, photosynthesis, N2-fixation rates, and plant nodule carbon and nitrogen metabolic activities in the two systems. M. truncatula plants in symbiosis with S. medicae showed increased biomass and photosynthesis rates per plant. Plants grown in symbiosis with S. medicae WSM419 also showed higher N2-fixation rates, which were correlated with a larger nodule biomass, while nodule number was similar in both systems. In terms of plant nodule metabolism, M. truncatula–S. medicae WSM419 nodules showed increased sucrose-catabolic activity, mostly associated with sucrose synthase, accompanied by a reduced starch content, whereas nitrogen-assimilation activities were comparable to those measured in nodules infected with S. meliloti 2011. Taken together, these results suggest that S. medicae WSM419 is able to enhance plant carbon catabolism in M. truncatula nodules, which allows for the maintaining of high symbiotic N2-fixation rates, better growth and improved general plant performance. PMID:25221545

  17. Exploring root symbiotic programs in the model legume Medicago truncatula using EST analysis

    PubMed Central

    Journet, Etienne-Pascal; van Tuinen, Diederik; Gouzy, Jérome; Crespeau, Hervé; Carreau, Véronique; Farmer, Mary-Jo; Niebel, Andreas; Schiex, Thomas; Jaillon, Olivier; Chatagnier, Odile; Godiard, Laurence; Micheli, Fabienne; Kahn, Daniel; Gianinazzi-Pearson, Vivienne; Gamas, Pascal

    2002-01-01

    We report on a large-scale expressed sequence tag (EST) sequencing and analysis program aimed at characterizing the sets of genes expressed in roots of the model legume Medicago truncatula during interactions with either of two microsymbionts, the nitrogen-fixing bacterium Sinorhizobium meliloti or the arbuscular mycorrhizal fungus Glomus intraradices. We have designed specific tools for in silico analysis of EST data, in relation to chimeric cDNA detection, EST clustering, encoded protein prediction, and detection of differential expression. Our 21 473 5′- and 3′-ESTs could be grouped into 6359 EST clusters, corresponding to distinct virtual genes, along with 52 498 other M.truncatula ESTs available in the dbEST (NCBI) database that were recruited in the process. These clusters were manually annotated, using a specifically developed annotation interface. Analysis of EST cluster distribution in various M.truncatula cDNA libraries, supported by a refined R test to evaluate statistical significance and by ‘electronic northern’ representation, enabled us to identify a large number of novel genes predicted to be up- or down-regulated during either symbiotic root interaction. These in silico analyses provide a first global view of the genetic programs for root symbioses in M.truncatula. A searchable database has been built and can be accessed through a public interface. PMID:12490726

  18. Exploring root symbiotic programs in the model legume Medicago truncatula using EST analysis.

    PubMed

    Journet, Etienne-Pascal; van Tuinen, Diederik; Gouzy, Jérome; Crespeau, Hervé; Carreau, Véronique; Farmer, Mary-Jo; Niebel, Andreas; Schiex, Thomas; Jaillon, Olivier; Chatagnier, Odile; Godiard, Laurence; Micheli, Fabienne; Kahn, Daniel; Gianinazzi-Pearson, Vivienne; Gamas, Pascal

    2002-12-15

    We report on a large-scale expressed sequence tag (EST) sequencing and analysis program aimed at characterizing the sets of genes expressed in roots of the model legume Medicago truncatula during interactions with either of two microsymbionts, the nitrogen-fixing bacterium Sinorhizobium meliloti or the arbuscular mycorrhizal fungus Glomus intraradices. We have designed specific tools for in silico analysis of EST data, in relation to chimeric cDNA detection, EST clustering, encoded protein prediction, and detection of differential expression. Our 21 473 5'- and 3'-ESTs could be grouped into 6359 EST clusters, corresponding to distinct virtual genes, along with 52 498 other M.truncatula ESTs available in the dbEST (NCBI) database that were recruited in the process. These clusters were manually annotated, using a specifically developed annotation interface. Analysis of EST cluster distribution in various M.truncatula cDNA libraries, supported by a refined R test to evaluate statistical significance and by 'electronic northern' representation, enabled us to identify a large number of novel genes predicted to be up- or down-regulated during either symbiotic root interaction. These in silico analyses provide a first global view of the genetic programs for root symbioses in M.truncatula. A searchable database has been built and can be accessed through a public interface. PMID:12490726

  19. Adaptation to climate through flowering phenology: a case study in Medicago truncatula.

    PubMed

    Burgarella, Concetta; Chantret, Nathalie; Gay, Laurène; Prosperi, Jean-Marie; Bonhomme, Maxime; Tiffin, Peter; Young, Nevin D; Ronfort, Joelle

    2016-07-01

    Local climatic conditions likely constitute an important selective pressure on genes underlying important fitness-related traits such as flowering time, and in many species, flowering phenology and climatic gradients strongly covary. To test whether climate shapes the genetic variation on flowering time genes and to identify candidate flowering genes involved in the adaptation to environmental heterogeneity, we used a large Medicago truncatula core collection to examine the association between nucleotide polymorphisms at 224 candidate genes and both climate variables and flowering phenotypes. Unlike genome-wide studies, candidate gene approaches are expected to enrich for the number of meaningful trait associations because they specifically target genes that are known to affect the trait of interest. We found that flowering time mediates adaptation to climatic conditions mainly by variation at genes located upstream in the flowering pathways, close to the environmental stimuli. Variables related to the annual precipitation regime reflected selective constraints on flowering time genes better than the other variables tested (temperature, altitude, latitude or longitude). By comparing phenotype and climate associations, we identified 12 flowering genes as the most promising candidates responsible for phenological adaptation to climate. Four of these genes were located in the known flowering time QTL region on chromosome 7. However, climate and flowering associations also highlighted largely distinct gene sets, suggesting different genetic architectures for adaptation to climate and flowering onset.

  20. Expression Analysis of PIN Genes in Root Tips and Nodules of Medicago truncatula

    PubMed Central

    Sańko-Sawczenko, Izabela; Łotocka, Barbara; Czarnocka, Weronika

    2016-01-01

    Polar auxin transport is dependent on the family of PIN-formed proteins (PINs), which are membrane transporters of anionic indole-3-acetic acid (IAA−). It is assumed that polar auxin transport may be essential in the development and meristematic activity maintenance of Medicago truncatula (M. truncatula) root nodules. However, little is known about the involvement of specific PIN proteins in M. truncatula nodulation. Using real-time quantitative PCR, we analyzed the expression patterns of all previously identified MtPIN genes and compared them between root nodules and root tips of M. truncatula. Our results demonstrated significant differences in the expression level of all 11 genes (MtPIN1–MtPIN11) between examined organs. Interestingly, MtPIN9 was the only PIN gene with higher expression level in root nodules compared to root tips. This result is the first indication of PIN9 transporter potential involvement in M. truncatula nodulation. Moreover, relatively high expression level in root nodules was attributed to MtPINs encoding orthologs of Arabidopsis thaliana PIN5 subclade. PIN proteins from this subclade have been found to localize in the endoplasmic reticulum, which may indicate that the development and meristematic activity maintenance of M. truncatula root nodules is associated with intracellular homeostasis of auxins level and their metabolism in the endoplasmic reticulum. PMID:27463709

  1. Flavone Synthases from Medicago truncatula Are Flavanone-2-Hydroxylases and Are Important for Nodulation1[W][OA

    PubMed Central

    Zhang, Juan; Subramanian, Senthil; Zhang, Yansheng; Yu, Oliver

    2007-01-01

    Flavones are important copigments found in the flowers of many higher plants and play a variety of roles in plant adaptation to stress. In Medicago species, flavones also act as signal molecules during symbiotic interaction with the diazotropic bacterium Sinorhizobium meliloti. They are the most potent nod gene inducers found in root exudates. However, flavone synthase II (FNS II), the key enzyme responsible for flavone biosynthesis, has not been characterized in Medicago species. We cloned two FNS II genes from Medicago truncatula using known FNS II sequences from other species and named them MtFNSII-1 and MtFNSII-2. Functional assays in yeast (Saccharomyces cerevisiae) suggested that the catalytic mechanisms of both cytochrome P450 monooxygenases were similar to the other known legume FNS II from licorice (Glycyrrhiza echinata). MtFNSII converted flavanones to 2-hydroxyflavanones instead of flavones whereas FNS II from the nonlegume Gerbera hybrida, converted flavanones to flavones directly. The two MtFNSII genes had distinct tissue-specific expression patterns. MtFNSII-1 was highly expressed in roots and seeds whereas MtFNSII-2 was highly expressed in flowers and siliques. In addition, MtFNSII-2 was inducible by S. meliloti and methyl jasmonate treatment, whereas MtFNSII-1 was not. Histochemical staining of transgenic hairy roots carrying the promoter-reporter constructs indicated that the MtFNSII-2 induction was tissue specific, mostly localized to vascular tissues and root hairs. RNA interference-mediated suppression of MtFNSII genes resulted in flavone depleted roots and led to significantly reduced nodulation when inoculated with S. meliloti. Our results provide genetic evidence supporting that flavones are important for nodulation in M. truncatula. PMID:17434990

  2. Comparative genomic sequence and expression analyses of Medicago truncatula and alfalfa subspecies falcata COLD-ACCLIMATION-SPECIFIC genes.

    PubMed

    Pennycooke, Joyce C; Cheng, Hongmei; Stockinger, Eric J

    2008-03-01

    In Arabidopsis (Arabidopsis thaliana) the low-temperature induction of genes encoding the C-REPEAT BINDING FACTOR (CBF) transcriptional activators is a key step in cold acclimation. CBFs in turn activate a battery of downstream genes known as the CBF regulon, which collectively act to increase tolerance to low temperatures. Fundamental questions are: What determines the size and scope of the CBF regulon, and is this is a major determinant of the low-temperature tolerance capacity of individual plant species? Here we have begun to address these questions through comparative analyses of Medicago truncatula and Medicago sativa subsp. falcata. M. truncatula survived to -4 degrees C but did not cold acclimate, whereas Medicago falcata cold acclimated and survived -14 degrees C. Both species possessed low-temperature-induced CBFs but differed in the expression of the COLD-ACCLIMATION-SPECIFIC (CAS) genes, which are candidate CBF targets. M. falcata CAS30 was robustly cold-responsive whereas the MtCAS31 homolog was not. M. falcata also possessed additional CAS30 homologs in comparison to the single CAS31 gene in M. truncatula. MfCAS30 possessed multiple pairs of closely spaced C-REPEAT/DEHYDRATION RESPONSIVE ELEMENT (CRT/DRE) motifs, the cognate CBF binding site in its upstream region whereas MtCAS31 lacked one CRT/DRE partner of the two proximal partner pairs. CAS genes also shared a promoter structure comprising modules proximal and distal to the coding sequence. CAS15, highly cold-responsive in both species, harbored numerous CRT/DRE motifs, but only in the distal module. However, fusion of the MtCAS15 promoter, including the distal module, to a reporter gene did not result in low-temperature responsiveness in stably transformed Arabidopsis. In contrast, both MtCAS31 and MfCAS30 promoter fusions were low-temperature responsive, although the MfCAS31 fusion was less robust than the MfCAS30 fusion. From these studies we conclude that CAS genes harbor CRT/DRE motifs, their

  3. Differential expression proteomics to investigate responses and resistance to Orobanche crenata in Medicago truncatula

    PubMed Central

    Castillejo, Ma Ángeles; Maldonado, Ana M; Dumas-Gaudot, Eliane; Fernández-Aparicio, Mónica; Susín, Rafael; Diego, Rubiales; Jorrín, Jesús V

    2009-01-01

    Background Parasitic angiosperm Orobanche crenata infection represents a major constraint for the cultivation of legumes worldwide. The level of protection achieved to date is either incomplete or ephemeral. Hence, an efficient control of the parasite requires a better understanding of its interaction and associated resistance mechanisms at molecular levels. Results In order to study the plant response to this parasitic plant and the molecular basis of the resistance we have used a proteomic approach. The root proteome of two accessions of the model legume Medicago truncatula displaying differences in their resistance phenotype, in control as well as in inoculated plants, over two time points (21 and 25 days post infection), has been compared. We report quantitative as well as qualitative differences in the 2-DE maps between early- (SA 27774) and late-resistant (SA 4087) genotypes after Coomassie and silver-staining: 69 differential spots were observed between non-inoculated genotypes, and 42 and 25 spots for SA 4087 and SA 27774 non-inoculated and inoculated plants, respectively. In all, 49 differential spots were identified by peptide mass fingerprinting (PMF) following MALDI-TOF/TOF mass spectrometry. Many of the proteins showing significant differences between genotypes and after parasitic infection belong to the functional category of defense and stress-related proteins. A number of spots correspond to proteins with the same function, and might represent members of a multigenic family or post-transcriptional forms of the same protein. Conclusion The results obtained suggest the existence of a generic defense mechanism operating during the early stages of infection and differing in both genotypes. The faster response to the infection observed in the SA 27774 genotype might be due to the action of proteins targeted against key elements needed for the parasite's successful infection, such as protease inhibitors. Our data are discussed and compared with those

  4. Salinity Adaptation and the Contribution of Parental Environmental Effects in Medicago truncatula

    PubMed Central

    Moriuchi, Ken S.; Friesen, Maren L.; Cordeiro, Matilde A.; Badri, Mounawer; Vu, Wendy T.; Main, Bradley J.; Aouani, Mohamed Elarbi; Nuzhdin, Sergey V.; Strauss, Sharon Y.; von Wettberg, Eric J. B.

    2016-01-01

    High soil salinity negatively influences plant growth and yield. Some taxa have evolved mechanisms for avoiding or tolerating elevated soil salinity, which can be modulated by the environment experienced by parents or offspring. We tested the contribution of the parental and offspring environments on salinity adaptation and their potential underlying mechanisms. In a two-generation greenhouse experiment, we factorially manipulated salinity concentrations for genotypes of Medicago truncatula that were originally collected from natural populations that differed in soil salinity. To compare population level adaptation to soil salinity and to test the potential mechanisms involved we measured two aspects of plant performance, reproduction and vegetative biomass, and phenological and physiological traits associated with salinity avoidance and tolerance. Saline-origin populations had greater biomass and reproduction under saline conditions than non-saline populations, consistent with local adaptation to saline soils. Additionally, parental environmental exposure to salt increased this difference in performance. In terms of environmental effects on mechanisms of salinity adaptation, parental exposure to salt spurred phenological differences that facilitated salt avoidance, while offspring exposure to salt resulted in traits associated with greater salt tolerance. Non-saline origin populations expressed traits associated with greater growth in the absence of salt while, for saline adapted populations, the ability to maintain greater performance in saline environments was also associated with lower growth potential in the absence of salt. Plastic responses induced by parental and offspring environments in phenology, leaf traits, and gas exchange contribute to salinity adaptation in M. truncatula. The ability of plants to tolerate environmental stress, such as high soil salinity, is likely modulated by a combination of parental effects and within-generation phenotypic

  5. Sexual Polyploidization in Medicago sativa L.: Impact on the Phenotype, Gene Transcription, and Genome Methylation.

    PubMed

    Rosellini, Daniele; Ferradini, Nicoletta; Allegrucci, Stefano; Capomaccio, Stefano; Zago, Elisa Debora; Leonetti, Paola; Balech, Bachir; Aversano, Riccardo; Carputo, Domenico; Reale, Lara; Veronesi, Fabio

    2016-01-01

    Polyploidization as the consequence of 2n gamete formation is a prominent mechanism in plant evolution. Studying its effects on the genome, and on genome expression, has both basic and applied interest. We crossed two diploid (2n = 2x = 16) Medicago sativa plants, a subsp. falcata seed parent, and a coerulea × falcata pollen parent that form a mixture of n and 2n eggs and pollen, respectively. Such a cross produced full-sib diploid and tetraploid (2n = 4x = 32) hybrids, the latter being the result of bilateral sexual polyploidization (BSP). These unique materials allowed us to investigate the effects of BSP, and to separate the effect of intraspecific hybridization from those of polyploidization by comparing 2x with 4x full sib progeny plants. Simple sequence repeat marker segregation demonstrated tetrasomic inheritance for all chromosomes but one, demonstrating that these neotetraploids are true autotetraploids. BSP brought about increased biomass, earlier flowering, higher seed set and weight, and larger leaves with larger cells. Microarray analyses with M. truncatula gene chips showed that several hundred genes, related to diverse metabolic functions, changed their expression level as a consequence of polyploidization. In addition, cytosine methylation increased in 2x, but not in 4x, hybrids. Our results indicate that sexual polyploidization induces significant transcriptional novelty, possibly mediated in part by DNA methylation, and phenotypic novelty that could underpin improved adaptation and reproductive success of tetraploid M. sativa with respect to its diploid progenitor. These polyploidy-induced changes may have promoted the adoption of tetraploid alfalfa in agriculture. PMID:26858330

  6. Integrated metabolomics and transcriptomics reveal enhanced specialized metabolism in Medicago truncatula root border cells.

    PubMed

    Watson, Bonnie S; Bedair, Mohamed F; Urbanczyk-Wochniak, Ewa; Huhman, David V; Yang, Dong Sik; Allen, Stacy N; Li, Wensheng; Tang, Yuhong; Sumner, Lloyd W

    2015-04-01

    Integrated metabolomics and transcriptomics of Medicago truncatula seedling border cells and root tips revealed substantial metabolic differences between these distinct and spatially segregated root regions. Large differential increases in oxylipin-pathway lipoxygenases and auxin-responsive transcript levels in border cells corresponded to differences in phytohormone and volatile levels compared with adjacent root tips. Morphological examinations of border cells revealed the presence of significant starch deposits that serve as critical energy and carbon reserves, as documented through increased β-amylase transcript levels and associated starch hydrolysis metabolites. A substantial proportion of primary metabolism transcripts were decreased in border cells, while many flavonoid- and triterpenoid-related metabolite and transcript levels were increased dramatically. The cumulative data provide compounding evidence that primary and secondary metabolism are differentially programmed in border cells relative to root tips. Metabolic resources normally destined for growth and development are redirected toward elevated accumulation of specialized metabolites in border cells, resulting in constitutively elevated defense and signaling compounds needed to protect the delicate root cap and signal motile rhizobia required for symbiotic nitrogen fixation. Elevated levels of 7,4'-dihydroxyflavone were further increased in border cells of roots exposed to cotton root rot (Phymatotrichopsis omnivora), and the value of 7,4'-dihydroxyflavone as an antimicrobial compound was demonstrated using in vitro growth inhibition assays. The cumulative and pathway-specific data provide key insights into the metabolic programming of border cells that strongly implicate a more prominent mechanistic role for border cells in plant-microbe signaling, defense, and interactions than envisioned previously.

  7. Speciation Matters: Bioavailability of Silver and Silver Sulfide Nanoparticles to Alfalfa (Medicago sativa).

    PubMed

    Stegemeier, John P; Schwab, Fabienne; Colman, Benjamin P; Webb, Samuel M; Newville, Matthew; Lanzirotti, Antonio; Winkler, Christopher; Wiesner, Mark R; Lowry, Gregory V

    2015-07-21

    Terrestrial crops are directly exposed to silver nanoparticles (Ag-NPs) and their environmentally transformed analog silver sulfide nanoparticles (Ag2S-NPs) when wastewater treatment biosolids are applied as fertilizer to agricultural soils. This leads to a need to understand their bioavailability to plants. In the present study, the mechanisms of uptake and distribution of silver in alfalfa (Medicago sativa) were quantified and visualized upon hydroponic exposure to Ag-NPs, Ag2S-NPs, and AgNO3 at 3 mg total Ag/L. Total silver uptake was measured in dried roots and shoots, and the spatial distribution of elements was investigated using transmission electron microscopy (TEM) and synchrotron-based X-ray imaging techniques. Despite large differences in release of Ag(+) ions from the particles, Ag-NPs, Ag2S-NPs, and Ag(+) became associated with plant roots to a similar degree, and exhibited similarly limited (<1%) amounts of translocation of silver into the shoot system. X-ray fluorescence (XRF) mapping revealed differences in the distribution of Ag into roots for each treatment. Silver nanoparticles mainly accumulated in the (columella) border cells and elongation zone, whereas Ag(+) accumulated more uniformly throughout the root. In contrast, Ag2S-NPs remained largely adhered to the root exterior, and the presence of cytoplasmic nano-SixOy aggregates was observed. Exclusively in roots exposed to particulate silver, NPs smaller than the originally dosed NPs were identified by TEM in the cell walls. The apparent accumulation of Ag in the root apoplast determined by XRF, and the presence of small NPs in root cell walls suggests uptake of partially dissolved NPs and translocation along the apoplast.

  8. Sexual Polyploidization in Medicago sativa L.: Impact on the Phenotype, Gene Transcription, and Genome Methylation

    PubMed Central

    Rosellini, Daniele; Ferradini, Nicoletta; Allegrucci, Stefano; Capomaccio, Stefano; Zago, Elisa Debora; Leonetti, Paola; Balech, Bachir; Aversano, Riccardo; Carputo, Domenico; Reale, Lara; Veronesi, Fabio

    2016-01-01

    Polyploidization as the consequence of 2n gamete formation is a prominent mechanism in plant evolution. Studying its effects on the genome, and on genome expression, has both basic and applied interest. We crossed two diploid (2n = 2x = 16) Medicago sativa plants, a subsp. falcata seed parent, and a coerulea × falcata pollen parent that form a mixture of n and 2n eggs and pollen, respectively. Such a cross produced full-sib diploid and tetraploid (2n = 4x = 32) hybrids, the latter being the result of bilateral sexual polyploidization (BSP). These unique materials allowed us to investigate the effects of BSP, and to separate the effect of intraspecific hybridization from those of polyploidization by comparing 2x with 4x full sib progeny plants. Simple sequence repeat marker segregation demonstrated tetrasomic inheritance for all chromosomes but one, demonstrating that these neotetraploids are true autotetraploids. BSP brought about increased biomass, earlier flowering, higher seed set and weight, and larger leaves with larger cells. Microarray analyses with M. truncatula gene chips showed that several hundred genes, related to diverse metabolic functions, changed their expression level as a consequence of polyploidization. In addition, cytosine methylation increased in 2x, but not in 4x, hybrids. Our results indicate that sexual polyploidization induces significant transcriptional novelty, possibly mediated in part by DNA methylation, and phenotypic novelty that could underpin improved adaptation and reproductive success of tetraploid M. sativa with respect to its diploid progenitor. These polyploidy-induced changes may have promoted the adoption of tetraploid alfalfa in agriculture. PMID:26858330

  9. Ensifer meliloti is the preferred symbiont of Medicago arborea in eastern Morocco soils.

    PubMed

    Guerrouj, Kamal; Pérez-Valera, Eduardo; Abdelmoumen, Hanaa; Bedmar, Eulogio J; Missbah El Idrissi, Mustapha

    2013-08-01

    Polyphasic characterization of 61 bacteria isolated from root nodules of Medicago arborea (Medic tree) plants growing in 4 arid soils of the arid eastern area of Morocco was studied. All the isolates characterized were fast growers. The phenotypic, symbiotic, and cultural characteristics analyzed allowed the description of a broad physiological diversity among the isolates. The results obtained suggest that the phenotype of these rhizobia might have evolved to adapt to the local conditions. The genetic characterization consisted of an analysis of the rep-PCR (repetitive extragenic palindromic polymerase chain reaction) fingerprints and a PCR-based RFLP (restriction fragment length polymorphism) of the 16S rDNA patterns. The diversity of the isolates was investigated by rep-PCR, giving a similarity of 62%, delineated into 3 clusters, 4 groups, and 6 subclusters. This wide diversity was also observed by a phenotypic approach, where the carbohydrate assimilation test was the most discriminating. The results show a relationship between rep-PCR fingerprinting and sugar assimilation, which are complementary in diversity investigation. The nearly complete 16S rRNA gene sequence from representative strains of each soil showed they are closely related to members of the genus Ensifer of the family Rhizobiaceae within the Alphaproteobacteria and shows the highest similitude values (99.93%/100%) with Ensifer meliloti LMG 6133(T) (X67222). Sequencing of the symbiotic nodC gene from 7 representative strains revealed they had 94.89% identity with the nodC sequence of the type strain E. meliloti LMG 6133(T) (EF428922). Therefore, the 61 M. arborea isolates from the 4 different soils have the same phylogenetic affiliation, which proves the restricted host specificity among M. arborea species.

  10. Cross-Family Translational Genomics of Abiotic Stress-Responsive Genes between Arabidopsis and Medicago truncatula

    PubMed Central

    Kim, Jin-Hyun; Yoo, Dongwoon; Seo, Young-Su; Jeong, Soon-Chun; Lee, Jai-Heon; Chung, Youngsoo; Jung, Ki-Hong; Cook, Douglas R.; Choi, Hong-kyu

    2014-01-01

    Cross-species translation of genomic information may play a pivotal role in applying biological knowledge gained from relatively simple model system to other less studied, but related, genomes. The information of abiotic stress (ABS)-responsive genes in Arabidopsis was identified and translated into the legume model system, Medicago truncatula. Various data resources, such as TAIR/AtGI DB, expression profiles and literatures, were used to build a genome-wide list of ABS genes. tBlastX/BlastP similarity search tools and manual inspection of alignments were used to identify orthologous genes between the two genomes. A total of 1,377 genes were finally collected and classified into 18 functional criteria of gene ontology (GO). The data analysis according to the expression cues showed that there was substantial level of interaction among three major types (i.e., drought, salinity and cold stress) of abiotic stresses. In an attempt to translate the ABS genes between these two species, genomic locations for each gene were mapped using an in-house-developed comparative analysis platform. The comparative analysis revealed that fragmental colinearity, represented by only 37 synteny blocks, existed between Arabidopsis and M. truncatula. Based on the combination of E-value and alignment remarks, estimated translation rate was 60.2% for this cross-family translation. As a prelude of the functional comparative genomic approaches, in-silico gene network/interactome analyses were conducted to predict key components in the ABS responses, and one of the sub-networks was integrated with corresponding comparative map. The results demonstrated that core members of the sub-network were well aligned with previously reported ABS regulatory networks. Taken together, the results indicate that network-based integrative approaches of comparative and functional genomics are important to interpret and translate genomic information for complex traits such as abiotic stresses. PMID:24675968

  11. Duration and intensity of shade differentially affects mycorrhizal growth- and phosphorus uptake responses of Medicago truncatula

    PubMed Central

    Konvalinková, Tereza; Püschel, David; Janoušková, Martina; Gryndler, Milan; Jansa, Jan

    2015-01-01

    Plant and fungal partners in arbuscular mycorrhizal symbiosis trade mineral nutrients for carbon, with the outcome of this relationship for plant growth and nutrition being highly context-dependent and changing with the availability of resources as well as with the specific requirements of the different partners. Here we studied how the model legume Medicago truncatula, inoculated or not with a mycorrhizal fungus Rhizophagus irregularis, responded to a gradient of light intensities applied over different periods of time, in terms of growth, phosphorus nutrition and the levels of root colonization by the mycorrhizal fungus. Short-term (6 d) shading, depending on its intensity, resulted in a rapid decline of phosphorus uptake to the shoots of mycorrhizal plants and simultaneous accumulation of phosphorus in the roots (most likely in the fungal tissues), as compared to the non-mycorrhizal controls. There was, however, no significant change in the levels of mycorrhizal colonization of roots due to short-term shading. Long-term (38 d) shading, depending on its intensity, provoked a multitude of plant compensatory mechanisms, which were further boosted by the mycorrhizal symbiosis. Mycorrhizal growth- and phosphorus uptake benefits, however, vanished at 10% of the full light intensity applied over a long-term. Levels of root colonization by the mycorrhizal fungus were significantly reduced by long-term shading. Our results indicate that even short periods of shade could have important consequences for the functioning of mycorrhizal symbiosis in terms of phosphorus transfer between the fungus and the plants, without any apparent changes in root colonization parameters or mycorrhizal growth response, and call for more focused research on temporal dynamics of mycorrhizal functioning under changing environmental conditions. PMID:25763002

  12. The alternative Medicago truncatula defense proteome of ROS—defective transgenic roots during early microbial infection

    PubMed Central

    Kiirika, Leonard M.; Schmitz, Udo; Colditz, Frank

    2014-01-01

    ROP-type GTPases of plants function as molecular switches within elementary signal transduction pathways such as the regulation of ROS synthesis via activation of NADPH oxidases (RBOH-respiratory burst oxidase homolog in plants). Previously, we reported that silencing of the Medicago truncatula GTPase MtROP9 led to reduced ROS production and suppressed induction of ROS-related enzymes in transgenic roots (MtROP9i) infected with pathogenic (Aphanomyces euteiches) and symbiotic microorganisms (Glomus intraradices, Sinorhizobium meliloti). While fungal infections were enhanced, S. meliloti infection was drastically impaired. In this study, we investigate the temporal proteome response of M. truncatula MtROP9i transgenic roots during the same microbial interactions under conditions of deprived potential to synthesize ROS. In comparison with control roots (Mtvector), we present a comprehensive proteomic analysis using sensitive MS protein identification. For four early infection time-points (1, 3, 5, 24 hpi), 733 spots were found to be different in abundance: 213 spots comprising 984 proteins (607 unique) were identified after S. meliloti infection, 230 spots comprising 796 proteins (580 unique) after G. intraradices infection, and 290 spots comprising 1240 proteins (828 unique) after A. euteiches infection. Data evaluation by GelMap in combination with a heatmap tool allowed recognition of key proteome changes during microbial interactions under conditions of hampered ROS synthesis. Overall, the number of induced proteins in MtROP9i was low as compared with controls, indicating a dual function of ROS in defense signaling as well as alternative response patterns activated during microbial infection. Qualitative analysis of induced proteins showed that enzymes linked to ROS production and scavenging were highly induced in control roots, while in MtROP9i the majority of proteins were involved in alternative defense pathways such as cell wall and protein degradation. PMID

  13. Two euAGAMOUS genes control C-function in Medicago truncatula.

    PubMed

    Serwatowska, Joanna; Roque, Edelín; Gómez-Mena, Concepción; Constantin, Gabriela D; Wen, Jiangqi; Mysore, Kirankumar S; Lund, Ole S; Johansen, Elisabeth; Beltrán, José Pío; Cañas, Luis A

    2014-01-01

    C-function MADS-box transcription factors belong to the AGAMOUS (AG) lineage and specify both stamen and carpel identity and floral meristem determinacy. In core eudicots, the AG lineage is further divided into two branches, the euAG and PLE lineages. Functional analyses across flowering plants strongly support the idea that duplicated AG lineage genes have different degrees of subfunctionalization of the C-function. The legume Medicago truncatula contains three C-lineage genes in its genome: two euAG genes (MtAGa and MtAGb) and one PLENA-like gene (MtSHP). This species is therefore a good experimental system to study the effects of gene duplication within the AG subfamily. We have studied the respective functions of each euAG genes in M. truncatula employing expression analyses and reverse genetic approaches. Our results show that the M. truncatula euAG- and PLENA-like genes are an example of subfunctionalization as a result of a change in expression pattern. MtAGa and MtAGb are the only genes showing a full C-function activity, concomitant with their ancestral expression profile, early in the floral meristem, and in the third and fourth floral whorls during floral development. In contrast, MtSHP expression appears late during floral development suggesting it does not contribute significantly to the C-function. Furthermore, the redundant MtAGa and MtAGb paralogs have been retained which provides the overall dosage required to specify the C-function in M. truncatula.

  14. Speciation Matters: Bioavailability of Silver and Silver Sulfide Nanoparticles to Alfalfa (Medicago sativa).

    PubMed

    Stegemeier, John P; Schwab, Fabienne; Colman, Benjamin P; Webb, Samuel M; Newville, Matthew; Lanzirotti, Antonio; Winkler, Christopher; Wiesner, Mark R; Lowry, Gregory V

    2015-07-21

    Terrestrial crops are directly exposed to silver nanoparticles (Ag-NPs) and their environmentally transformed analog silver sulfide nanoparticles (Ag2S-NPs) when wastewater treatment biosolids are applied as fertilizer to agricultural soils. This leads to a need to understand their bioavailability to plants. In the present study, the mechanisms of uptake and distribution of silver in alfalfa (Medicago sativa) were quantified and visualized upon hydroponic exposure to Ag-NPs, Ag2S-NPs, and AgNO3 at 3 mg total Ag/L. Total silver uptake was measured in dried roots and shoots, and the spatial distribution of elements was investigated using transmission electron microscopy (TEM) and synchrotron-based X-ray imaging techniques. Despite large differences in release of Ag(+) ions from the particles, Ag-NPs, Ag2S-NPs, and Ag(+) became associated with plant roots to a similar degree, and exhibited similarly limited (<1%) amounts of translocation of silver into the shoot system. X-ray fluorescence (XRF) mapping revealed differences in the distribution of Ag into roots for each treatment. Silver nanoparticles mainly accumulated in the (columella) border cells and elongation zone, whereas Ag(+) accumulated more uniformly throughout the root. In contrast, Ag2S-NPs remained largely adhered to the root exterior, and the presence of cytoplasmic nano-SixOy aggregates was observed. Exclusively in roots exposed to particulate silver, NPs smaller than the originally dosed NPs were identified by TEM in the cell walls. The apparent accumulation of Ag in the root apoplast determined by XRF, and the presence of small NPs in root cell walls suggests uptake of partially dissolved NPs and translocation along the apoplast. PMID:26106801

  15. Mineral accumulation in vegetative and reproductive tissues during seed development in Medicago truncatula

    PubMed Central

    Garcia, Christina B.; Grusak, Michael A.

    2015-01-01

    Enhancing nutrient density in legume seeds is one of several strategies being explored to improve the nutritional quality of the food supply. In order to develop crop varieties with increased seed mineral concentration, a more detailed understanding of mineral translocation within the plant is required. By studying mineral accumulation in different organs within genetically diverse members of the same species, it may be possible to identify variable traits that modulate seed mineral concentration. We utilized two ecotypes (A17 and DZA315.16) of the model legume, Medicago truncatula, to study dry mass and mineral accumulation in the leaves, pod walls, and seeds during reproductive development. The pod wall dry mass was significantly different between the two ecotypes beginning at 12 days after pollination, whereas there was no significant difference in the average dry mass of individual seeds between the two ecotypes at any time point. There were also no significant differences in leaf dry mass between ecotypes; however, we observed expansion of A17 leaves during the first 21 days of pod development, while DZA315.16 leaves did not display a significant increase in leaf area. Mineral profiling of the leaves, pod walls, and seeds highlighted differences in accumulation patterns among minerals within each tissue as well as genotypic differences with respect to individual minerals. Because there were differences in the average seed number per pod, the total seed mineral content per pod was generally higher in A17 than DZA315.16. In addition, mineral partitioning to the seeds tended to be higher in A17 pods. These data revealed that mineral retention within leaves and/or pod walls might attenuate mineral accumulation within the seeds. As a result, strategies to increase seed mineral content should include approaches that will enhance export from these tissues. PMID:26322063

  16. Salinity Adaptation and the Contribution of Parental Environmental Effects in Medicago truncatula.

    PubMed

    Moriuchi, Ken S; Friesen, Maren L; Cordeiro, Matilde A; Badri, Mounawer; Vu, Wendy T; Main, Bradley J; Aouani, Mohamed Elarbi; Nuzhdin, Sergey V; Strauss, Sharon Y; von Wettberg, Eric J B

    2016-01-01

    High soil salinity negatively influences plant growth and yield. Some taxa have evolved mechanisms for avoiding or tolerating elevated soil salinity, which can be modulated by the environment experienced by parents or offspring. We tested the contribution of the parental and offspring environments on salinity adaptation and their potential underlying mechanisms. In a two-generation greenhouse experiment, we factorially manipulated salinity concentrations for genotypes of Medicago truncatula that were originally collected from natural populations that differed in soil salinity. To compare population level adaptation to soil salinity and to test the potential mechanisms involved we measured two aspects of plant performance, reproduction and vegetative biomass, and phenological and physiological traits associated with salinity avoidance and tolerance. Saline-origin populations had greater biomass and reproduction under saline conditions than non-saline populations, consistent with local adaptation to saline soils. Additionally, parental environmental exposure to salt increased this difference in performance. In terms of environmental effects on mechanisms of salinity adaptation, parental exposure to salt spurred phenological differences that facilitated salt avoidance, while offspring exposure to salt resulted in traits associated with greater salt tolerance. Non-saline origin populations expressed traits associated with greater growth in the absence of salt while, for saline adapted populations, the ability to maintain greater performance in saline environments was also associated with lower growth potential in the absence of salt. Plastic responses induced by parental and offspring environments in phenology, leaf traits, and gas exchange contribute to salinity adaptation in M. truncatula. The ability of plants to tolerate environmental stress, such as high soil salinity, is likely modulated by a combination of parental effects and within-generation phenotypic

  17. Catalase and superoxide dismutase in alfalfa root nodules. [Medicago sativa L

    SciTech Connect

    Becana, M.; Aparicio-Tejo, P.M.; Sanchez-Diaz, M.

    1986-04-01

    Catalase and superoxide dismutase (SOD), in scavenging H/sub 2/ O/sub 2/ and O/sub 2/, respectively, have been recently proposed to play a role in leghemoglobin protection. The occurrence of catalase and SOD activities in alfalfa (Medicago sativa L.) nodule cytosol is reported here. Enzymes were extracted at 0-4/sup 0/C from 0.5 g fresh nodules with 12 ml of a medium containing K-phosphate buffer 50 mM, pH 7.8 and Na/sub 2/EDTA 0.1 mM. The homogenate was filtered and centrifuged at 18,000 xg for 10 min, and the resulting supernatant was used for catalase assay. A further precipitation of leghemoglobin was required to avoid interferences with SOD determination. Catalase was determined by back-titration with KMnO/sub 4/. SOD was assayed by measuring the inhibition of nitro blue tetrazolium reduction. The sensitivity of SOD activity to CN/sup -/ was tested by including 1 mM KCN in the reaction mixture. Catalase activity of alfalfa nodule cytosol was 237 +/- 1 units/mg protein, decreasing very significantly (P < 0.01, Duncan's multiple range test) at 20 mM NO/sub 3//sup -/. Typical specific SOD activities were 94 +/- 5 and 65 +/- 4 units/mg protein, without CN/sup -/ and with CN/sup -/, respectively. Both activities increased very significantly at 20 mM NO/sub 3//sup -/. SOD activities with CN/sup -/ were 70-80% those without CN/sup -/ within the range of NO/sub 3//sup -/ investigated (0-20 mM).

  18. Integrated Metabolomics and Transcriptomics Reveal Enhanced Specialized Metabolism in Medicago truncatula Root Border Cells1[OPEN

    PubMed Central

    Watson, Bonnie S.; Bedair, Mohamed F.; Urbanczyk-Wochniak, Ewa; Huhman, David V.; Yang, Dong Sik; Allen, Stacy N.; Li, Wensheng; Tang, Yuhong; Sumner, Lloyd W.

    2015-01-01

    Integrated metabolomics and transcriptomics of Medicago truncatula seedling border cells and root tips revealed substantial metabolic differences between these distinct and spatially segregated root regions. Large differential increases in oxylipin-pathway lipoxygenases and auxin-responsive transcript levels in border cells corresponded to differences in phytohormone and volatile levels compared with adjacent root tips. Morphological examinations of border cells revealed the presence of significant starch deposits that serve as critical energy and carbon reserves, as documented through increased β-amylase transcript levels and associated starch hydrolysis metabolites. A substantial proportion of primary metabolism transcripts were decreased in border cells, while many flavonoid- and triterpenoid-related metabolite and transcript levels were increased dramatically. The cumulative data provide compounding evidence that primary and secondary metabolism are differentially programmed in border cells relative to root tips. Metabolic resources normally destined for growth and development are redirected toward elevated accumulation of specialized metabolites in border cells, resulting in constitutively elevated defense and signaling compounds needed to protect the delicate root cap and signal motile rhizobia required for symbiotic nitrogen fixation. Elevated levels of 7,4′-dihydroxyflavone were further increased in border cells of roots exposed to cotton root rot (Phymatotrichopsis omnivora), and the value of 7,4′-dihydroxyflavone as an antimicrobial compound was demonstrated using in vitro growth inhibition assays. The cumulative and pathway-specific data provide key insights into the metabolic programming of border cells that strongly implicate a more prominent mechanistic role for border cells in plant-microbe signaling, defense, and interactions than envisioned previously. PMID:25667316

  19. Salinity Adaptation and the Contribution of Parental Environmental Effects in Medicago truncatula.

    PubMed

    Moriuchi, Ken S; Friesen, Maren L; Cordeiro, Matilde A; Badri, Mounawer; Vu, Wendy T; Main, Bradley J; Aouani, Mohamed Elarbi; Nuzhdin, Sergey V; Strauss, Sharon Y; von Wettberg, Eric J B

    2016-01-01

    High soil salinity negatively influences plant growth and yield. Some taxa have evolved mechanisms for avoiding or tolerating elevated soil salinity, which can be modulated by the environment experienced by parents or offspring. We tested the contribution of the parental and offspring environments on salinity adaptation and their potential underlying mechanisms. In a two-generation greenhouse experiment, we factorially manipulated salinity concentrations for genotypes of Medicago truncatula that were originally collected from natural populations that differed in soil salinity. To compare population level adaptation to soil salinity and to test the potential mechanisms involved we measured two aspects of plant performance, reproduction and vegetative biomass, and phenological and physiological traits associated with salinity avoidance and tolerance. Saline-origin populations had greater biomass and reproduction under saline conditions than non-saline populations, consistent with local adaptation to saline soils. Additionally, parental environmental exposure to salt increased this difference in performance. In terms of environmental effects on mechanisms of salinity adaptation, parental exposure to salt spurred phenological differences that facilitated salt avoidance, while offspring exposure to salt resulted in traits associated with greater salt tolerance. Non-saline origin populations expressed traits associated with greater growth in the absence of salt while, for saline adapted populations, the ability to maintain greater performance in saline environments was also associated with lower growth potential in the absence of salt. Plastic responses induced by parental and offspring environments in phenology, leaf traits, and gas exchange contribute to salinity adaptation in M. truncatula. The ability of plants to tolerate environmental stress, such as high soil salinity, is likely modulated by a combination of parental effects and within-generation phenotypic

  20. Proteomic Identification of Differentially Expressed Proteins during Alfalfa (Medicago sativa L.) Flower Development

    PubMed Central

    Chen, Lingling; Chen, Quanzhu; Zhu, Yanqiao; Hou, Longyu; Mao, Peisheng

    2016-01-01

    Flower development, pollination, and fertilization are important stages in the sexual reproduction process of plants; they are also critical steps in the control of seed formation and development. During alfalfa (Medicago sativa L.) seed production, some distinct phenomena such as a low seed setting ratio, serious flower falling, and seed abortion commonly occur. However, the causes of these phenomena are complicated and largely unknown. An understanding of the mechanisms that regulate alfalfa flowering is important in order to increase seed yield. Hence, proteomic technology was used to analyze changes in protein expression during the stages of alfalfa flower development. Flower samples were collected at pre-pollination (S1), pollination (S2), and the post-pollination senescence period (S3). Twenty-four differentially expressed proteins were successfully identified, including 17 down-regulated in pollinated flowers, one up-regulated in pollinated and senesced flowers, and six up-regulated in senesced flowers. The largest proportions of the identified proteins were involved in metabolism, signal transduction, defense response, oxidation reduction, cell death, and programmed cell death (PCD). Their expression profiles demonstrated that energy metabolism, carbohydrate metabolism, and amino acid metabolism provided the nutrient foundation for pollination in alfalfa. Furthermore, there were three proteins involved in multiple metabolic pathways: dual specificity kinase splA-like protein (kinase splALs), carbonic anhydrase, and NADPH: quinone oxidoreductase-like protein. Expression patterns of these proteins indicated that MAPK cascades regulated multiple processes, such as signal transduction, stress response, and cell death. PCD also played an important role in the alfalfa flower developmental process, and regulated both pollination and flower senescence. The current study sheds some light on protein expression profiles during alfalfa flower development and

  1. Ontogeny of embryogenic callus in Medicago truncatula: the fate of the pluripotent and totipotent stem cells

    PubMed Central

    Wang, Xin-Ding; Nolan, Kim E.; Irwanto, Rina R.; Sheahan, Michael B.; Rose, Ray J.

    2011-01-01

    Background and Aims Understanding the fate and dynamics of cells during callus formation is essential to understanding totipotency and the mechanisms of somatic embryogenesis. Here, the fate of leaf explant cells during the development of embryogenic callus was investigated in the model legume Medicago truncatula. Methods Callus development was examined from cultured leaf explants of the highly regenerable genotype Jemalong 2HA (2HA) and from mesophyll protoplasts of 2HA and wild-type Jemalong. Callus development was studied by histology, manipulation of the culture system, detection of early production of reactive oxygen species and visualization of SERK1 (SOMATIC EMBRYO RECEPTOR KINASE1) gene expression. Key Results Callus formation in leaf explants initiates at the cut surface and within veins of the explant. The ontogeny of callus development is dominated by the division and differentiation of cells derived from pluripotent procambial cells and from dedifferentiated mesophyll cells. Procambium-derived cells differentiated into vascular tissue and rarely formed somatic embryos, whereas dedifferentiated mesophyll cells were competent to form somatic embryos. Interestingly, explants incubated adaxial-side down had substantially less cell proliferation associated with veins yet produced similar numbers of somatic embryos to explants incubated abaxial-side down. Somatic embryos mostly formed on the explant surface originally in contact with the medium, while in protoplast microcalli, somatic embryos only fully developed once at the surface of the callus. Mesophyll protoplasts of 2HA formed embryogenic callus while Jemalong mesophyll protoplasts produced callus rich in vasculature. Conclusions The ontogeny of embryogenic callus in M. truncatula relates to explant orientation and is driven by the dynamics of pluripotent procambial cells, which proliferate and differentiate into vasculature. The ontogeny is also related to de-differentiated mesophyll cells that acquire

  2. Influence of pCO2 on carbon allocation in nodulated Medicago sativa L.

    NASA Astrophysics Data System (ADS)

    Pereyra, Gabriela; Hartmann, Henrik; Ziegler, Waldemar; Michalzik, Beate; Gonzalez-Meler, Miquel; Trumbore, Susan

    2016-04-01

    Atmospheric CO2 concentrations (pCO_2) have been related to changes in plant carbon (C) availability and photosynthetic capacity, yet there is no clear consensus as to the effect of pCO2 on the plant C balance and on nitrogen fixation in symbiotic systems. We investigated how different pCO2 (Pleistocene: 170 ppm, ambient: 400 ppm and projected future: 700 ppm) influence C allocation in nodulated Medicago sativa L. We labeled 17 week old plants with depleted 13C (-34.7±1.2‰) and traced the label over a 9-day period, to assess the redistribution of newly assimilated C across different sinks, including nodules. We analyzed N concentrations in plant tissues and found no significant differences in leaves and roots across treatments. However, growth and C fixation rates increased with pCO_2, and differences were greatest between 170 ppm and 700 ppm. Across pCO2 treatments we observed a 13C-enrichment in roots compared to leaves. We further observed the highest 13C depletion of non-structural carbohydrates (NSCs) and respired CO2 in tissues of plants grown at 700 ppm, especially in leaves and nodules. Our preliminary results suggest that sink organs like roots and nodules are fed with newly-assimilated NSCs from leaves to support respiration, and especially in 170 ppm plants represented a major respiratory loss of newly assimilated C (≈ 35{%} of the total plant respiration). Our results suggest that although plant metabolic processes like photosynthesis and respiration are affected by changes in pCO_2, nitrogen acquisition in such a symbiotic system is not.

  3. Medicago truncatula CYP716A12 Is a Multifunctional Oxidase Involved in the Biosynthesis of Hemolytic Saponins[W

    PubMed Central

    Carelli, Maria; Biazzi, Elisa; Panara, Francesco; Tava, Aldo; Scaramelli, Laura; Porceddu, Andrea; Graham, Neil; Odoardi, Miriam; Piano, Efisio; Arcioni, Sergio; May, Sean; Scotti, Carla; Calderini, Ornella

    2011-01-01

    Saponins, a group of glycosidic compounds present in several plant species, have aglycone moieties that are formed using triterpenoid or steroidal skeletons. In spite of their importance as antimicrobial compounds and their possible benefits for human health, knowledge of the genetic control of saponin biosynthesis is still poorly understood. In the Medicago genus, the hemolytic activity of saponins is related to the nature of their aglycone moieties. We have identified a cytochrome P450 gene (CYP716A12) involved in saponin synthesis in Medicago truncatula using a combined genetic and biochemical approach. Genetic loss-of-function analysis and complementation studies showed that CYP716A12 is responsible for an early step in the saponin biosynthetic pathway. Mutants in CYP716A12 were unable to produce hemolytic saponins and only synthetized soyasaponins, and were thus named lacking hemolytic activity (lha). In vitro enzymatic activity assays indicate that CYP716A12 catalyzes the oxidation of β-amyrin and erythrodiol at the C-28 position, yielding oleanolic acid. Transcriptome changes in the lha mutant showed a modulation in the main steps of triterpenic saponin biosynthetic pathway: squalene cyclization, β-amyrin oxidation, and glycosylation. The analysis of CYP716A12 expression in planta is reported together with the sapogenin content in different tissues and stages. This article provides evidence for CYP716A12 being a key gene in hemolytic saponin biosynthesis. PMID:21821776

  4. How grow-and-switch gravitropism generates root coiling and root waving growth responses in Medicago truncatula

    PubMed Central

    Tan, Tzer Han; Silverberg, Jesse L.; Floss, Daniela S.; Harrison, Maria J.; Henley, Christopher L.; Cohen, Itai

    2015-01-01

    Experimental studies show that plant root morphologies can vary widely from straight gravity-aligned primary roots to fractal-like root architectures. However, the opaqueness of soil makes it difficult to observe how environmental factors modulate these patterns. Here, we combine a transparent hydrogel growth medium with a custom built 3D laser scanner to directly image the morphology of Medicago truncatula primary roots. In our experiments, root growth is obstructed by an inclined plane in the growth medium. As the tilt of this rigid barrier is varied, we find Medicago transitions between randomly directed root coiling, sinusoidal root waving, and normal gravity-aligned morphologies. Although these root phenotypes appear morphologically distinct, our analysis demonstrates the divisions are less well defined, and instead, can be viewed as a 2D biased random walk that seeks the path of steepest decent along the inclined plane. Features of this growth response are remarkably similar to the widely known run-and-tumble chemotactic behavior of Escherichia coli bacteria, where biased random walks are used as optimal strategies for nutrient uptake. PMID:26432881

  5. Analysis of Cell Wall-Related Genes in Organs of Medicago sativa L. under Different Abiotic Stresses

    PubMed Central

    Behr, Marc; Legay, Sylvain; Hausman, Jean-Francois; Guerriero, Gea

    2015-01-01

    Abiotic constraints are a source of concern in agriculture, because they can have a strong impact on plant growth and development, thereby affecting crop yield. The response of plants to abiotic constraints varies depending on the type of stress, on the species and on the organs. Although many studies have addressed different aspects of the plant response to abiotic stresses, only a handful has focused on the role of the cell wall. A targeted approach has been used here to study the expression of cell wall-related genes in different organs of alfalfa plants subjected for four days to three different abiotic stress treatments, namely salt, cold and heat stress. Genes involved in different steps of cell wall formation (cellulose biosynthesis, monolignol biosynthesis and polymerization) have been analyzed in different organs of Medicago sativa L. Prior to this analysis, an in silico classification of dirigent/dirigent-like proteins and class III peroxidases has been performed in Medicago truncatula and M. sativa. The final goal of this study is to infer and compare the expression patterns of cell wall-related genes in response to different abiotic stressors in the organs of an important legume crop. PMID:26193255

  6. The Hybridization Barrier between Herbaceous Medicago sativa and Woody M. arborea Is Weakened by Selection of Seed Parents

    PubMed Central

    Bingham, Edwin; Armour, David; Irwin, John

    2013-01-01

    Medicago sativa, alfalfa or lucerne, and M. arborea were considered reproductively isolated until recently. Then, in 2003, an alfalfa genotype was identified that produced a few seeds and progeny with hybrid traits after a large number of pollinations by M. arborea. A derivative of this alfalfa genotype also produced a low frequency of progeny with hybrid traits. Thus, the hybridization barrier was weakened by selection of seed parents. Hybrids from both events expressed traits from M. arborea and M. arborea-specific DNA bands, although more of the M. sativa genome was retained, based on the DNA results. Thus, there was chromatin elimination during embryogenesis, resulting in partial hybrids (hereafter hybrids). However, more than 30 hybrids with an array of M. arborea traits have been obtained thus far, and research continues on the nature of the hybrids. Traits have been genetically transmitted in crosses, and selected traits are in use for alfalfa breeding. This paper reviews the first hybrids and then focuses on further weakening of the hybridization barrier with the discovery of a more efficient hybridizer derived from crossing Medicago sativa subspecies, sativa, coerulea and falcata. This genotype was found to have reproductive abnormalities associated with its complex subspecies origin that are best described as hybrid breakdown. In effect, this subspecies derivative is a bridge-cross parent that consistently produces hybrids. Reproductive abnormalities in the bridge-cross parent are reported and discussed. PMID:27137379

  7. How grow-and-switch gravitropism generates root coiling and root waving growth responses in Medicago truncatula.

    PubMed

    Tan, Tzer Han; Silverberg, Jesse L; Floss, Daniela S; Harrison, Maria J; Henley, Christopher L; Cohen, Itai

    2015-10-20

    Experimental studies show that plant root morphologies can vary widely from straight gravity-aligned primary roots to fractal-like root architectures. However, the opaqueness of soil makes it difficult to observe how environmental factors modulate these patterns. Here, we combine a transparent hydrogel growth medium with a custom built 3D laser scanner to directly image the morphology of Medicago truncatula primary roots. In our experiments, root growth is obstructed by an inclined plane in the growth medium. As the tilt of this rigid barrier is varied, we find Medicago transitions between randomly directed root coiling, sinusoidal root waving, and normal gravity-aligned morphologies. Although these root phenotypes appear morphologically distinct, our analysis demonstrates the divisions are less well defined, and instead, can be viewed as a 2D biased random walk that seeks the path of steepest decent along the inclined plane. Features of this growth response are remarkably similar to the widely known run-and-tumble chemotactic behavior of Escherichia coli bacteria, where biased random walks are used as optimal strategies for nutrient uptake.

  8. How grow-and-switch gravitropism generates root coiling and root waving growth responses in Medicago truncatula.

    PubMed

    Tan, Tzer Han; Silverberg, Jesse L; Floss, Daniela S; Harrison, Maria J; Henley, Christopher L; Cohen, Itai

    2015-10-20

    Experimental studies show that plant root morphologies can vary widely from straight gravity-aligned primary roots to fractal-like root architectures. However, the opaqueness of soil makes it difficult to observe how environmental factors modulate these patterns. Here, we combine a transparent hydrogel growth medium with a custom built 3D laser scanner to directly image the morphology of Medicago truncatula primary roots. In our experiments, root growth is obstructed by an inclined plane in the growth medium. As the tilt of this rigid barrier is varied, we find Medicago transitions between randomly directed root coiling, sinusoidal root waving, and normal gravity-aligned morphologies. Although these root phenotypes appear morphologically distinct, our analysis demonstrates the divisions are less well defined, and instead, can be viewed as a 2D biased random walk that seeks the path of steepest decent along the inclined plane. Features of this growth response are remarkably similar to the widely known run-and-tumble chemotactic behavior of Escherichia coli bacteria, where biased random walks are used as optimal strategies for nutrient uptake. PMID:26432881

  9. Multifaceted Investigation of Metabolites During Nitrogen Fixation in Medicago via High Resolution MALDI-MS Imaging and ESI-MS

    NASA Astrophysics Data System (ADS)

    Gemperline, Erin; Jayaraman, Dhileepkumar; Maeda, Junko; Ané, Jean-Michel; Li, Lingjun

    2015-01-01

    Legumes have developed the unique ability to establish a symbiotic relationship with soil bacteria known as rhizobia. This interaction results in the formation of root nodules in which rhizobia thrive and reduce atmospheric dinitrogen into plant-usable ammonium through biological nitrogen fixation (BNF). Owing to the availability of genetic information for both of the symbiotic partners, the Medicago truncatula- Sinorhizobium meliloti association is an excellent model for examining the BNF process. Although metabolites are important in this symbiotic association, few studies have investigated the array of metabolites that influence this process. Of these studies, most target only a few specific metabolites, the roles of which are either well known or are part of a well-characterized metabolic pathway. Here, we used a multifaceted mass spectrometric (MS) approach to detect and identify the key metabolites that are present during BNF using the Medicago truncatula- Sinorhizobium meliloti association as the model system. High mass accuracy and high resolution matrix-assisted laser desorption/ionization (MALDI) and electrospray ionization (ESI) Orbitrap instruments were used in this study and provide complementary results for more in-depth characterization of the nitrogen-fixation process. We used well-characterized plant and bacterial mutants to highlight differences between the metabolites that are present in functional versus nonfunctional nodules. Our study highlights the benefits of using a combination of mass spectrometric techniques to detect differences in metabolite composition and the distributions of these metabolites in plant biology.

  10. Stress tolerance and symbiotic and phylogenic features of root nodule bacteria associated with Medicago species in different bioclimatic regions of Tunisia.

    PubMed

    Djedidi, Salem; Yokoyama, Tadashi; Ohkama-Ohtsu, Naoko; Risal, Chandra Prasad; Abdelly, Chedly; Sekimoto, Hitoshi

    2011-01-01

    Thirty two rhizobial isolates were obtained from different bioclimatic regions of Tunisia using as trap plants, Medicago sativa, Medicago ciliaris, Medicago polymorpha and Medicago minima. To study their diversity and characterize them in relation to Mediterranean conditions, abiotic stress resistance, symbiotic properties and genetic diversity in terms of 16S rRNA and nodA sequences were assessed. Five isolates from M. sativa, three from M. ciliaris and three from M. minima could grow at 45°C. Only two isolates from M. sativa grew at 4% NaCl. The most stress tolerant isolates were obtained from arid soils. A phylogenetic analysis of 16S rRNA genes revealed 29 isolates to be closely related to Ensifer including one (Pl.3-9) that showed a 16S rRNA sequence similar to that of Ensifer meliloti and nodA sequence similar to that of Ensifer medicae. However, three isolates were categorized into Agrobacterium containing the nodA of Ensifer. Furthermore, these isolates developed nodules on original hosts. The results for the four isolates suggest horizontal gene transfer between the species.

  11. Effects of Fe deficiency on the protein profiles and lignin composition of stem tissues of Medicago truncatula in absence and presence of calcium carbonate

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Iron deficiency is a yield-limiting factor with major implications for crop production, especially in soils with high CaCO3. Because stems are essential for the delivery of nutrients to the shoots, the aim of this work was to study the effects of Fe deficiency on the stem proteome of Medicago trunca...

  12. Control of Dissected Leaf Morphology by a Cys(2)His(2) Zinc Finger Transcription Factor in the Model Legume Medicago Truncatula

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Different plants may have different leaf types. Diversity in leaf types contributes to a large degree of plant diversity in the natural environment. How different leaf morphology is determined is not yet understood. The leguminous plant Medicago truncatula exhibits dissected leaves with three leafl...

  13. Genetic transformation and analysis of rice OsAPx2 gene in Medicago sativa.

    PubMed

    Guan, Qingjie; Takano, Tetsuo; Liu, Shenkui

    2012-01-01

    The OsAPx2 gene from rice was cloned to produce PBI121::OsAPx2 dual-expression plants, of which expression level would be increasing under stressful conditions. The enzyme ascorbate peroxidase (APX) in the leaves and roots of the plants increased with increasing exposure time to different sodium chloride (NaCl) and hydrogen peroxide (H(2)O(2))concentrations, as indicated by protein gel blot analysis. The increased enzyme yield improved the ability of the plants to resist the stress treatments. The OsAPx2 gene was localized in the cytoplasm of epidermal onion cells as indicated by the instantaneous expression of green fluorescence. An 80% regeneration rate was observed in Medicago sativa L. plants transformed with the OsAPx2 gene using Agrobacterium tumefaciens, as indicated by specific primer PCR. The OsAPx2 gene was expressed at the mRNA level and the individual M. sativa (T#1,T#2,T#5) were obtained through assaying the generation of positive T2 using RNA gel blot analysis. When the seeds of the wild type (WT) and the T2 (T#1,T#5) were incubated in culture containing MS with NaCl for 7 days, the results as shown of following: the root length of transgenic plant was longer than WT plants, the H(2)O(2) content in roots of WT was more than of transgenic plants, the APX activity under stresses increased by 2.89 times compared with the WT, the malondialdehyde (MDA) content of the WT was higher than the transgenic plants, the leaves of the WT turned yellow, but those of the transgenic plants remained green and remained healthy. The chlorophyll content in the WT leaves was less than in the transgenic plants, after soaking in solutions of H(2)O(2), sodium sulfite (Na(2)SO(3)), and sodium bicarbonate (NaHCO(3)). Therefore, the OsAPx2 gene overexpression in transgenic M. sativa improves the removal of H(2)O(2) and the salt-resistance compared with WT plants. A novel strain of M. sativa carrying a salt-resistance gene was obtained. PMID:22848448

  14. Genetic transformation and analysis of rice OsAPx2 gene in Medicago sativa.

    PubMed

    Guan, Qingjie; Takano, Tetsuo; Liu, Shenkui

    2012-01-01

    The OsAPx2 gene from rice was cloned to produce PBI121::OsAPx2 dual-expression plants, of which expression level would be increasing under stressful conditions. The enzyme ascorbate peroxidase (APX) in the leaves and roots of the plants increased with increasing exposure time to different sodium chloride (NaCl) and hydrogen peroxide (H(2)O(2))concentrations, as indicated by protein gel blot analysis. The increased enzyme yield improved the ability of the plants to resist the stress treatments. The OsAPx2 gene was localized in the cytoplasm of epidermal onion cells as indicated by the instantaneous expression of green fluorescence. An 80% regeneration rate was observed in Medicago sativa L. plants transformed with the OsAPx2 gene using Agrobacterium tumefaciens, as indicated by specific primer PCR. The OsAPx2 gene was expressed at the mRNA level and the individual M. sativa (T#1,T#2,T#5) were obtained through assaying the generation of positive T2 using RNA gel blot analysis. When the seeds of the wild type (WT) and the T2 (T#1,T#5) were incubated in culture containing MS with NaCl for 7 days, the results as shown of following: the root length of transgenic plant was longer than WT plants, the H(2)O(2) content in roots of WT was more than of transgenic plants, the APX activity under stresses increased by 2.89 times compared with the WT, the malondialdehyde (MDA) content of the WT was higher than the transgenic plants, the leaves of the WT turned yellow, but those of the transgenic plants remained green and remained healthy. The chlorophyll content in the WT leaves was less than in the transgenic plants, after soaking in solutions of H(2)O(2), sodium sulfite (Na(2)SO(3)), and sodium bicarbonate (NaHCO(3)). Therefore, the OsAPx2 gene overexpression in transgenic M. sativa improves the removal of H(2)O(2) and the salt-resistance compared with WT plants. A novel strain of M. sativa carrying a salt-resistance gene was obtained.

  15. The utilization of chopped and pelleted lucerne (Medicago sativa) by growing lambs.

    PubMed

    Thomson, D J; Cammell, S B

    1979-03-01

    1. The efficiency of utilization of the dietary energy and nitrogen contained in a dried lucerne (Medicago sativa cv. Chartainvilliers) given either chopped (CL) or ground (1.96 mm screen) and pelleted (PL), was measured in a comparative slaughter experiment. Growing lambs were given equal amounts of digestible energy in the chopped or pelleted form at each of the three planes of nutrition for a period of 100 d. 2. The initial energy, fat and protein content of both the carcass and the total body of the test lambs was estimated from regression equations between fasted (18 h) live weight and these components, derived from a group of twenty-three comparable lambs. The final energy, fat and protein content of the test lambs was determined directly by chemical analyses. 3. The metabolizable energy (ME) content of the diets was derived at each plane of nutrition from measured faecal and urinary losses and estimated methane losses. The depression in ME content with grinding and pelleting the dried lucerne was small (CL 8.69 MJ/kg dry matter (DM), PL 8.42 MG/kg DM). 4. The efficiency of utilization of the ME of the dried lucerne for growth and fattening was higher (P less than 0.01) when given in the ground pelleted form (0.533), than in the chopped form (0.284). The net energy value of the PL (3.5 MJ/kg DM) was higher than that of CL (2.2 MJ/kg DM). 5. Thus lambs fed on PL grew faster and had a higher caracass weight gain, carcass protein and fat retention than lambs fed on CL. The composition of the carcass was not altered by the physical processing treatment. 6. Digestion studies with these same CL and PL diets had shown that grinding and pelleting depressed digestion in the forestomachs and increased digestion in the small intestine compared with the chopped form. The increased efficiency of utilization of the gross energy and ME and the higher net energy value of PL was attributed primarily to a change in the site of digestion within the alimentary tract. Associated

  16. Genetic analysis of tolerance to the root lesion nematode Pratylenchus neglectus in the legume Medicago littoralis

    PubMed Central

    2014-01-01

    Background The nematode Pratylenchus neglectus has a wide host range and is able to feed on the root systems of cereals, oilseeds, grain and pasture legumes. Under the Mediterranean low rainfall environments of Australia, annual Medicago pasture legumes are used in rotation with cereals to fix atmospheric nitrogen and improve soil parameters. Considerable efforts are being made in breeding programs to improve resistance and tolerance to Pratylenchus neglectus in the major crops wheat and barley, which makes it vital to develop appropriate selection tools in medics. Results A strong source of tolerance to root damage by the root lesion nematode (RLN) Pratylenchus neglectus had previously been identified in line RH-1 (strand medic, M. littoralis). Using RH-1, we have developed a single seed descent (SSD) population of 138 lines by crossing it to the intolerant cultivar Herald. After inoculation, RLN-associated root damage clearly segregated in the population. Genetic analysis was performed by constructing a genetic map using simple sequence repeat (SSR) and gene-based SNP markers. A highly significant quantitative trait locus (QTL), QPnTolMl.1, was identified explaining 49% of the phenotypic variation in the SSD population. All SSRs and gene-based markers in the QTL region were derived from chromosome 1 of the sequenced genome of the closely related species M. truncatula. Gene-based markers were validated in advanced breeding lines derived from the RH-1 parent and also a second RLN tolerance source, RH-2 (M. truncatula ssp. tricycla). Comparative analysis to sequenced legume genomes showed that the physical QTL interval exists as a synteny block in Lotus japonicus, common bean, soybean and chickpea. Furthermore, using the sequenced genome information of M. truncatula, the QTL interval contains 55 genes out of which five are discussed as potential candidate genes responsible for the mapped tolerance. Conclusion The closely linked set of SNP-based PCR markers is

  17. Genome-wide transcriptional analysis of super-embryogenic Medicago truncatula explant cultures

    PubMed Central

    Imin, Nijat; Goffard, Nicolas; Nizamidin, Mahira; Rolfe, Barry G

    2008-01-01

    Background The Medicago truncatula (M. truncatula) line 2HA has a 500-fold greater capacity to regenerate plants in culture by somatic embryogenesis than its wild type progenitor Jemalong. To understand the molecular basis for the regeneration capacity of this super-embryogenic line 2HA, using Affymetrix GeneChip®, we have compared transcriptomes of explant leaf cultures of these two lines that were grown on media containing the auxin NAA (1-naphthaleneacetic acid) and the cytokinin BAP (6-benzylaminopurine) for two weeks, an early time point for tissue culture proliferation. Results Using Affymetrix GeneChip®, GCRMA normalisation and statistical analysis, we have shown that more than 196 and 49 probe sets were significantly (p < 0.05) up- or down-regulated respectively more than 2 fold in expression. We have utilised GeneBins, a database for classifying gene expression data to distinguish differentially displayed pathways among these two cultures which showed changes in number of biochemical pathways including carbon and flavonoid biosynthesis, phytohormone biosynthesis and signalling. The up-regulated genes in the embryogenic 2HA culture included nodulins, transporters, regulatory genes, embryogenesis related arabinogalactans and genes involved in redox homeostasis, the transition from vegetative growth to reproductive growth and cytokinin signalling. Down-regulated genes included protease inhibitors, wound-induced proteins, and genes involved in biosynthesis and signalling of phytohormones auxin, gibberellin and ethylene. These changes indicate essential differences between the super-embryogenic line 2HA and Jemalong not only in many aspects of biochemical pathways but also in their response to auxin and cytokinin. To validate the GeneChip results, we used quantitative real-time RT-PCR to examine the expression of the genes up-regulated in 2HA such as transposase, RNA-directed DNA polymerase, glycoside hydrolase, RESPONSE REGULATOR 10, AGAMOUS-LIKE 20, flower

  18. Genetic Transformation and Analysis of Rice OsAPx2 Gene in Medicago sativa

    PubMed Central

    Guan, Qingjie; Takano, Tetsuo; Liu, Shenkui

    2012-01-01

    The OsAPx2 gene from rice was cloned to produce PBI121::OsAPx2 dual-expression plants, of which expression level would be increasing under stressful conditions. The enzyme ascorbate peroxidase (APX) in the leaves and roots of the plants increased with increasing exposure time to different sodium chloride (NaCl) and hydrogen peroxide (H2O2)concentrations, as indicated by protein gel blot analysis. The increased enzyme yield improved the ability of the plants to resist the stress treatments. The OsAPx2 gene was localized in the cytoplasm of epidermal onion cells as indicated by the instantaneous expression of green fluorescence. An 80% regeneration rate was observed in Medicago sativa L. plants transformed with the OsAPx2 gene using Agrobacterium tumefaciens, as indicated by specific primer PCR. The OsAPx2 gene was expressed at the mRNA level and the individual M. sativa (T#1,T#2,T#5) were obtained through assaying the generation of positive T2 using RNA gel blot analysis. When the seeds of the wild type (WT) and the T2 (T#1,T#5) were incubated in culture containing MS with NaCl for 7 days, the results as shown of following: the root length of transgenic plant was longer than WT plants, the H2O2 content in roots of WT was more than of transgenic plants, the APX activity under stresses increased by 2.89 times compared with the WT, the malondialdehyde (MDA) content of the WT was higher than the transgenic plants, the leaves of the WT turned yellow, but those of the transgenic plants remained green and remained healthy. The chlorophyll content in the WT leaves was less than in the transgenic plants, after soaking in solutions of H2O2, sodium sulfite (Na2SO3), and sodium bicarbonate (NaHCO3). Therefore, the OsAPx2 gene overexpression in transgenic M. sativa improves the removal of H2O2 and the salt-resistance compared with WT plants. A novel strain of M. sativa carrying a salt-resistance gene was obtained. PMID:22848448

  19. A nitrogen-dependent switch in the high affinity ammonium transport in Medicago truncatula.

    PubMed

    Straub, Daniel; Ludewig, Uwe; Neuhäuser, Benjamin

    2014-11-01

    Ammonium transporters (AMTs) are crucial for the high affinity primary uptake and translocation of ammonium in plants. In the model legume Medicago truncatula, the genomic set of AMT-type ammonium transporters comprises eight members. Only four genes were abundantly expressed in young seedlings, both in roots and shoots. While the expression of all AMTs in the shoot was not affected by the nitrogen availability, the dominating MtAMT1;1 gene was repressed by nitrogen in roots, despite that cellular nitrogen concentrations were far above deficiency levels. A contrasting de-repression by nitrogen was observed for MtAMT1;4 and MtAMT2;1, which were both expressed at intermediate level. Weak expression was found for MtAMT1;2 and MtAMT2;3, while the other AMTs were not detected in young seedlings. When expressed from their endogenous promoters, translational fusion proteins of MtAMT1;1 and MtAMT2;1 with green fluorescent protein were co-localized in the plasma membrane of rhizodermal cells, but also detected in cortical root layers. Both transporter proteins similarly functionally complemented a yeast strain that is deficient in high affinity ammonium transport, both at acidic and neutral pH. The uptake into yeast mediated by these transporters saturated with Km AMT1;1 = 89 µM and Km AMT2;1 = 123 µM, respectively. When expressed in oocytes, MtAMT1;1 mediated much larger (15)N-ammonium uptake than MtAMT2;1, but NH4 (+) currents were only recorded for MtAMT1;1. These currents saturated with a voltage-dependent Km = 90 µM at -80 mV. The cellular localization and regulation of the AMTs suggests that MtAMT1;1 encodes the major high affinity ammonium transporter gene in low nitrogen grown young M. truncatula roots and despite the similar localization and substrate affinity, MtAMT2;1 appears functionally distinct and more important at higher nitrogen supply. PMID:25164101

  20. The Medicago FLOWERING LOCUS T Homolog, MtFTa1, Is a Key Regulator of Flowering Time1[C][W][OA

    PubMed Central

    Laurie, Rebecca E.; Diwadkar, Payal; Jaudal, Mauren; Zhang, Lulu; Hecht, Valérie; Wen, Jiangqi; Tadege, Million; Mysore, Kirankumar S.; Putterill, Joanna; Weller, James L.; Macknight, Richard C.

    2011-01-01

    FLOWERING LOCUS T (FT) genes encode proteins that function as the mobile floral signal, florigen. In this study, we characterized five FT-like genes from the model legume, Medicago (Medicago truncatula). The different FT genes showed distinct patterns of expression and responses to environmental cues. Three of the FT genes (MtFTa1, MtFTb1, and MtFTc) were able to complement the Arabidopsis (Arabidopsis thaliana) ft-1 mutant, suggesting that they are capable of functioning as florigen. MtFTa1 is the only one of the FT genes that is up-regulated by both long days (LDs) and vernalization, conditions that promote Medicago flowering, and transgenic Medicago plants overexpressing the MtFTa1 gene flowered very rapidly. The key role MtFTa1 plays in regulating flowering was demonstrated by the identification of fta1 mutants that flowered significantly later in all conditions examined. fta1 mutants do not respond to vernalization but are still responsive to LDs, indicating that the induction of flowering by prolonged cold acts solely through MtFTa1, whereas photoperiodic induction of flowering involves other genes, possibly MtFTb1, which is only expressed in leaves under LD conditions and therefore might contribute to the photoperiodic regulation of flowering. The role of the MtFTc gene is unclear, as the ftc mutants did not have any obvious flowering-time or other phenotypes. Overall, this work reveals the diversity of the regulation and function of the Medicago FT family. PMID:21685176

  1. MuTAnT: a family of Mutator-like transposable elements targeting TA microsatellites in Medicago truncatula.

    PubMed

    Stawujak, Krzysztof; Startek, Michał; Gambin, Anna; Grzebelus, Dariusz

    2015-08-01

    Transposable elements (TEs) are mobile DNA segments, abundant and dynamic in plant genomes. Because their mobility can be potentially deleterious to the host, a variety of mechanisms evolved limiting that negative impact, one of them being preference for a specific target insertion site. Here, we describe a family of Mutator-like DNA transposons in Medicago truncatula targeting TA microsatellites. We identified 218 copies of MuTAnTs and an element carrying a complete ORF encoding a mudrA-like transposase. Most insertion sites are flanked by a variable number of TA tandem repeats, indicating that MuTAnTs are specifically targeting TA microsatellites. Other TE families flanked by TA repeats (e.g. TAFT elements in maize) were described previously, however we identified the first putative autonomous element sharing that characteristics with a related group of short non-autonomous transposons.

  2. N-feedback regulation is synchronized with nodule carbon alteration in Medicago truncatula under excessive nitrate or low phosphorus conditions.

    PubMed

    Sulieman, Saad; Schulze, Joachim; Tran, Lam-Son Phan

    2014-03-15

    The aim of the present study was to test the hypothesis that the higher nodule amino acid content induced under certain treatments may play a role in the N-feedback regulation of nitrogenase (EC 1.18.6.1) activity by restricting the carbon supply to the functioning nodules. Growing Medicago truncatula plants under sub-optimal phosphorus conditions or upon exposure to large supply of nitrate caused significant asparagine accumulation in nodules of the treated plants. In addition, there was a remarkable decline in the nodule succinate content under phosphorus deprivation while malate was tended to increase. Interestingly, the relative share of succinate in the symbiotic tissues was totally inhibited, i.e. reached zero, by excessive nitrate application. These results provide evidence that succinate might be greatly affected by asparagine content of the nodule fraction, thereby restricting cellular carbon supply to the functioning bacteroids which leads to down-regulation of nodule metabolism and nitrogenase activity.

  3. Depletion of tyrosyl-DNA phosphodiesterase 1α (MtTdp1α) affects transposon expression in Medicago truncatula.

    PubMed

    Sabatini, Maria Elisa; Donà, Mattia; Leonetti, Paola; Minio, Andrea; Delledonne, Massimo; Carboneral, Daniela; Confalonieri, Massimo; Giraffa, Giorgio; Balestrazzi, Alma

    2016-07-01

    The role of plant tyrosyl-DNA phosphodiesterase 1α in genome stability is studied using a Medicago truncatula MtTdp1α-depleted line. Lack of MtTdp1α results in a 39% reduction of methylated cytosines as compared to control. RNA-Seq analyses revealed that 11 DNA transposons and 22 retrotransposons were differentially expressed in the Tdp1α-2a line. Among them all, DNA transposons (MuDR, hAT, DNA3-11_Mad) and seven retrotransposons (LTR (Long Terminal Repeat)/Gipsy, LTR/Copia, LTR and NonLTR/L1) were down-regulated, while the 15 retrotransposons were up-regulated. Results suggest that the occurrence of stress-responsive cis-elements as well as changes in the methylation pattern at the LTR promoters might be responsible for the enhanced retrotransposon transcription. PMID:26699667

  4. Haustorium Formation in Medicago truncatula Roots Infected by Phytophthora palmivora Does Not Involve the Common Endosymbiotic Program Shared by Arbuscular Mycorrhizal Fungi and Rhizobia.

    PubMed

    Huisman, Rik; Bouwmeester, Klaas; Brattinga, Marijke; Govers, Francine; Bisseling, Ton; Limpens, Erik

    2015-12-01

    In biotrophic plant-microbe interactions, microbes infect living plant cells, in which they are hosted in a novel membrane compartment, the host-microbe interface. To create a host-microbe interface, arbuscular mycorrhizal (AM) fungi and rhizobia make use of the same endosymbiotic program. It is a long-standing hypothesis that pathogens make use of plant proteins that are dedicated to mutualistic symbiosis to infect plants and form haustoria. In this report, we developed a Phytophthora palmivora pathosystem to study haustorium formation in Medicago truncatula roots. We show that P. palmivora does not require host genes that are essential for symbiotic infection and host-microbe interface formation to infect Medicago roots and form haustoria. Based on these findings, we conclude that P. palmivora does not hijack the ancient intracellular accommodation program used by symbiotic microbes to form a biotrophic host-microbe interface.

  5. Variation in rDNA locus number and position among legume species and detection of 2 linked rDNA loci in the model Medicago truncatula by FISH.

    PubMed

    Abirached-Darmency, Mona; Prado-Vivant, Emilce; Chelysheva, Liudmila; Pouthier, Thomas

    2005-06-01

    Within Fabaceae, legume species have a variable genome size, chromosome number, and ploidy level. The genome distribution of ribosomal genes, easily detectable by fluorescent in situ hybridization (FISH), is a good tool for anchoring physical and genetic comparative maps. The organisation of 45S rDNA and 5S loci was analysed by FISH in the 4 closely related species: Pisum sativum, Medicago truncatula, Medicago sativa (2 diploid taxa), and Lathyrus sativus. The 2 types of rDNA arrays displayed interspecific variation in locus number and location, but little intraspecific variation was detected. In the model legume, M. truncatula, the presence of 2 adjacent 45S rDNA loci was demonstrated, and the location of the rDNA loci was independent of the general evolution of the genome DNA. The different parameters relative to clustering of the rDNA loci in specific chromosome regions and the possible basis of rDNA instability are discussed. PMID:16121252

  6. High-quality permanent draft genome sequence of Ensifer medicae strain WSM244, a microsymbiont isolated from Medicago polymorpha growing in alkaline soil

    SciTech Connect

    Ardley, Julie; Tian, Rui; O’Hara, Graham; Seshadri, Rekha; Reddy, T. B. K.; Pati, Amrita; Woyke, Tanja; Markowitz, Victor; Ivanova, Natalia; Kyrpides, Nikos; Howieson, John; Reeve, Wayne

    2015-12-01

    We report that Ensifer medicae WSM244 is an aerobic, motile, Gram-negative, non-spore-forming rod that can exist as a soil saprophyte or as a legume microsymbiont of Medicago species. WSM244 was isolated in 1979 from a nodule recovered from the roots of the annual Medicago polymorpha L. growing in alkaline soil (pH 8.0) in Tel Afer, Iraq. WSM244 is the only acid-sensitive E. medicae strain that has been sequenced to date. It is effective at fixing nitrogen with M. polymorpha L., as well as with more alkaline-adapted Medicago spp. such as M. littoralis Loisel., M. scutellata (L.) Mill., M. tornata (L.) Mill. and M. truncatula Gaertn. This strain is also effective with the perennial M. sativa L. Here we describe the features of E. medicae WSM244, together with genome sequence information and its annotation. The 6,650,282 bp high-quality permanent draft genome is arranged into 91 scaffolds of 91 contigs containing 6,427 protein-coding genes and 68 RNA-only encoding genes, and is one of the rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project proposal.

  7. High-quality permanent draft genome sequence of Ensifer medicae strain WSM244, a microsymbiont isolated from Medicago polymorpha growing in alkaline soil

    DOE PAGES

    Ardley, Julie; Tian, Rui; O’Hara, Graham; Seshadri, Rekha; Reddy, T. B. K.; Pati, Amrita; Woyke, Tanja; Markowitz, Victor; Ivanova, Natalia; Kyrpides, Nikos; et al

    2015-12-01

    We report that Ensifer medicae WSM244 is an aerobic, motile, Gram-negative, non-spore-forming rod that can exist as a soil saprophyte or as a legume microsymbiont of Medicago species. WSM244 was isolated in 1979 from a nodule recovered from the roots of the annual Medicago polymorpha L. growing in alkaline soil (pH 8.0) in Tel Afer, Iraq. WSM244 is the only acid-sensitive E. medicae strain that has been sequenced to date. It is effective at fixing nitrogen with M. polymorpha L., as well as with more alkaline-adapted Medicago spp. such as M. littoralis Loisel., M. scutellata (L.) Mill., M. tornata (L.)more » Mill. and M. truncatula Gaertn. This strain is also effective with the perennial M. sativa L. Here we describe the features of E. medicae WSM244, together with genome sequence information and its annotation. The 6,650,282 bp high-quality permanent draft genome is arranged into 91 scaffolds of 91 contigs containing 6,427 protein-coding genes and 68 RNA-only encoding genes, and is one of the rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project proposal.« less

  8. The 2HA line of Medicago truncatula has characteristics of an epigenetic mutant that is weakly ethylene insensitive

    PubMed Central

    2014-01-01

    Background The Medicago truncatula 2HA seed line is highly embryogenic while the parental line Jemalong rarely produces embryos. The 2HA line was developed from one of the rare Jemalong regenerates and this method for obtaining a highly regenerable genotype in M. truncatula is readily reproducible suggesting an epigenetic mechanism. Microarray transcriptomic analysis showed down regulation of an ETHYLENE INSENSITIVE 3-like gene in 2HA callus which provided an approach to investigating epigenetic regulation of genes related to ethylene signalling and the 2HA phenotype. Ethylene is involved in many developmental processes including somatic embryogenesis (SE) and is associated with stress responses. Results Microarray transcriptomic analysis showed a significant number of up-regulated transcripts in 2HA tissue culture, including nodule and embryo specific genes and transposon-like genes, while only a few genes were down-regulated, including an EIN3-like gene we called MtEIL1. This reduced expression was associated with ethylene insensitivity of 2HA plants that was further investigated. The weak ethylene insensitivity affected root and nodule development. Sequencing of MtEIL1 found no difference between 2HA and wild-type plants. DNA methylation analysis of MtEIL1 revealed significant difference between 2HA and wild-type plants. Tiling arrays demonstrated an elevated level of miRNA in 2HA plants that hybridised to the antisense strand of the MtEIL1 gene. AFLP-like methylation profiling revealed more differences in DNA methylation between 2HA and wild-type. Segregation analysis demonstrated the recessive nature of the eil1 phenotype and the dominant nature of the SE trait. Conclusions We have demonstrated that EIL1 of Medicago truncatula (MtEIL1) is epigenetically silenced in the 2HA seed line. The possible cause is an elevated level of miRNA that targets its 3’UTR and is also associated with DNA methylation of MtEIL1. Down regulation of MtEIL1 makes it possible to

  9. Arbuscular mycorrhizal symbiosis can mitigate the negative effects of night warming on physiological traits of Medicago truncatula L.

    PubMed

    Hu, Yajun; Wu, Songlin; Sun, Yuqing; Li, Tao; Zhang, Xin; Chen, Caiyan; Lin, Ge; Chen, Baodong

    2015-02-01

    Elevated night temperature, one of the main climate warming scenarios, can have profound effects on plant growth and metabolism. However, little attention has been paid to the potential role of mycorrhizal associations in plant responses to night warming, although it is well known that symbiotic fungi can protect host plants against various environmental stresses. In the present study, physiological traits of Medicago truncatula L. in association with the arbuscular mycorrhizal (AM) fungus Rhizophagus irregularis were investigated under simulated night warming. A constant increase in night temperature of 1.53 °C significantly reduced plant shoot and root biomass, flower and seed number, leaf sugar concentration, and shoot Zn and root P concentrations. However, the AM association essentially mitigated these negative effects of night warming by improving plant growth, especially through increased root biomass, root to shoot ratio, and shoot Zn and root P concentrations. A significant interaction was observed between R. irregularis inoculation and night warming in influencing both root sucrose concentration and expression of sucrose synthase (SusS) genes, suggesting that AM symbiosis and increased night temperature jointly regulated plant sugar metabolism. Night warming stimulated AM fungal colonization but did not influence arbuscule abundance, symbiosis-related plant or fungal gene expression, or growth of extraradical mycelium, indicating little effect of night warming on the development or functioning of AM symbiosis. These findings highlight the importance of mycorrhizal symbiosis in assisting plant resilience to climate warming. PMID:25033924

  10. The NIN Transcription Factor Coordinates Diverse Nodulation Programs in Different Tissues of the Medicago truncatula Root[OPEN

    PubMed Central

    Kim, Jiyoung; Frances, Lisa; Ding, Yiliang; Sun, Jongho; Guan, Dian; de Carvalho-Niebel, Fernanda; Oldroyd, Giles E.D.

    2015-01-01

    Biological nitrogen fixation in legumes occurs in nodules that are initiated in the root cortex following Nod factor recognition at the root surface, and this requires coordination of diverse developmental programs in these different tissues. We show that while early Nod factor signaling associated with calcium oscillations is limited to the root surface, the resultant activation of Nodule Inception (NIN) in the root epidermis is sufficient to promote cytokinin signaling and nodule organogenesis in the inner root cortex. NIN or a product of its action must be associated with the transmission of a signal between the root surface and the cortical cells where nodule organogenesis is initiated. NIN appears to have distinct functions in the root epidermis and the root cortex. In the epidermis, NIN restricts the extent of Early Nodulin 11 (ENOD11) expression and does so through competitive inhibition of ERF Required for Nodulation (ERN1). In contrast, NIN is sufficient to promote the expression of the cytokinin receptor Cytokinin Response 1 (CRE1), which is restricted to the root cortex. Our work in Medicago truncatula highlights the complexity of NIN action and places NIN as a central player in the coordination of the symbiotic developmental programs occurring in differing tissues of the root that combined are necessary for a nitrogen-fixing symbiosis. PMID:26672071

  11. Concurrent synthesis and release of nod-gene-inducing flavonoids from alfalfa roots. [Medicago sativa L. ; Rhizobium meliloti

    SciTech Connect

    Maxwell, C.A.; Phillips, D.A. )

    1990-08-01

    Flavonoid signals from alfalfa (Medicago sativa L.) induce transcription of nodulation (nod) genes in Rhizobium meliloti. Alfalfa roots release three major nod-gene inducers: 4{prime},7-dihydroxyflavanone, 4{prime},7-dihydroxyflavone, and 4,4{prime}-dihydroxy-2{prime}-methoxychalcone. The objective of the present study was to define temporal relationships between synthesis and exudation for those flavonoids. Requirements for concurrent flavonoid biosynthesis were assessed by treating roots of intact alfalfa seedlings with (U-{sup 14}C)-L-phenylalanine in the presence or absence of the phenylalanine ammonia-lyase inhibitor L-2-aminoxy-3-phenylpropionic acid (AOPP). In the absence of AOPP, each of the three flavonoids in exudates contained {sup 14}C. In the presence of AOPP, {sup 14}C labeling and release of all the exuded nod-gene inducers were reduced significantly. AOPP inhibited labeling and release of the strongest nod-gene inducer, methoxychalcone, by more than 90%. The release process responsible for exudation of nod-gene inducers appears to be specific rather than a general phenomenon such as a sloughing off of cells during root growth.

  12. Transcriptomic and metabolomic analyses identify a role for chlorophyll catabolism and phytoalexin during Medicago nonhost resistance against Asian soybean rust.

    PubMed

    Ishiga, Yasuhiro; Uppalapati, Srinivasa Rao; Gill, Upinder S; Huhman, David; Tang, Yuhong; Mysore, Kirankumar S

    2015-08-12

    Asian soybean rust (ASR) caused by Phakopsora pachyrhizi is a devastating foliar disease affecting soybean production worldwide. Understanding nonhost resistance against ASR may provide an avenue to engineer soybean to confer durable resistance against ASR. We characterized a Medicago truncatula-ASR pathosystem to study molecular mechanisms of nonhost resistance. Although urediniospores formed appressoria and penetrated into epidermal cells of M. truncatula, P. pachyrhizi failed to sporulate. Transcriptomic analysis revealed the induction of phenylpropanoid, flavonoid and isoflavonoid metabolic pathway genes involved in the production of phytoalexin medicarpin in M. truncatula upon infection with P. pachyrhizi. Furthermore, genes involved in chlorophyll catabolism were induced during nonhost resistance. We further characterized one of the chlorophyll catabolism genes, Stay-green (SGR), and demonstrated that the M. truncatula sgr mutant and alfalfa SGR-RNAi lines showed hypersensitive-response-like enhanced cell death upon inoculation with P. pachyrhizi. Consistent with transcriptomic analysis, metabolomic analysis also revealed the accumulation of medicarpin and its intermediate metabolites. In vitro assay showed that medicarpin inhibited urediniospore germination and differentiation. In addition, several triterpenoid saponin glycosides accumulated in M. truncatula upon inoculation with P. pachyrhizi. In summary, using multi-omic approaches, we identified a correlation between phytoalexin production and M. truncatula defense responses against ASR.

  13. Systemic signaling of the plant nitrogen status triggers specific transcriptome responses depending on the nitrogen source in Medicago truncatula.

    PubMed

    Ruffel, Sandrine; Freixes, Sandra; Balzergue, Sandrine; Tillard, Pascal; Jeudy, Christian; Martin-Magniette, Marie Laure; van der Merwe, Margaretha J; Kakar, Klementina; Gouzy, Jerôme; Fernie, Alisdair R; Udvardi, Michael; Salon, Christophe; Gojon, Alain; Lepetit, Marc

    2008-04-01

    Legumes can acquire nitrogen (N) from NO(3)(-), NH(4)(+), and N(2) (through symbiosis with Rhizobium bacteria); however, the mechanisms by which uptake and assimilation of these N forms are coordinately regulated to match the N demand of the plant are currently unknown. Here, we find by use of the split-root approach in Medicago truncatula plants that NO(3)(-) uptake, NH(4)(+) uptake, and N(2) fixation are under general control by systemic signaling of plant N status. Indeed, irrespective of the nature of the N source, N acquisition by one side of the root system is repressed by high N supply to the other side. Transcriptome analysis facilitated the identification of over 3,000 genes that were regulated by systemic signaling of the plant N status. However, detailed scrutiny of the data revealed that the observation of differential gene expression was highly dependent on the N source. Localized N starvation results, in the unstarved roots of the same plant, in a strong compensatory up-regulation of NO(3)(-) uptake but not of either NH(4)(+) uptake or N(2) fixation. This indicates that the three N acquisition pathways do not always respond similarly to a change in plant N status. When taken together, these data indicate that although systemic signals of N status control root N acquisition, the regulatory gene networks targeted by these signals, as well as the functional response of the N acquisition systems, are predominantly determined by the nature of the N source.

  14. Medicago truncatula and Glycine max: Different Drought Tolerance and Similar Local Response of the Root Nodule Proteome.

    PubMed

    Gil-Quintana, Erena; Lyon, David; Staudinger, Christiana; Wienkoop, Stefanie; González, Esther M

    2015-12-01

    Legume crops present important agronomical and environmental advantages mainly due to their capacity to reduce atmospheric N2 to ammonium via symbiotic nitrogen fixation (SNF). This process is very sensitive to abiotic stresses such as drought, but the mechanism underlying this response is not fully understood. The goal of the current work is to compare the drought response of two legumes with high economic impact and research importance, Medicago truncatula and Glycine max, by characterizing their root nodule proteomes. Our results show that, although M. truncatula exhibits lower water potential values under drought conditions compared to G. max, SNF declined analogously in the two legumes. Both of their nodule proteomes are very similar, and comparable down-regulation responses in the diverse protein functional groups were identified (mainly proteins related to the metabolism of carbon, nitrogen, and sulfur). We suggest lipoxygenases and protein turnover as newly recognized players in SNF regulation. Partial drought conditions applied to a split-root system resulted in the local down-regulation of the entire proteome of drought-stressed nodules in both legumes. The high degree of similarity between both legume proteomes suggests that the vast amount of research conducted on M. truncatula could be applied to economically important legume crops, such as soybean.

  15. Toxic Effects of Copper-based Nanoparticles or Compounds to Lettuce (Lactuca sativa) and Alfalfa (Medicago sativa)

    PubMed Central

    Hong, Jie; Rico, Cyren; Zhao, Lijuan; Adeleye, Adeyemi S.; Keller, Arturo A.; Peralta-Videa, Jose R.; Gardea-Torresdey, Jorge L.

    2014-01-01

    The increased production and use of nanoparticles (NPs) has generated concerns about their impact on living organisms. In this study, nCu, bulk Cu, nCuO, bulk CuO, Cu(OH)2 (CuPRO 2005, Kocide 3000), and CuCl2 were exposed for 15 days to 10 day-old hydroponically grown lettuce (Lactuca sativa) and alfalfa (Medicago sativa). Each compound was applied at 0, 5, 10, and 20 mg/L. At harvest, we measured the size of the plants and determined the concentration of Cu, macro and microelements by using ICP-OES. Catalase and ascorbate peroxidase activity was also determined. Results showed that all Cu NPs/compounds reduced the root length by 49% in both plant species. All Cu NPs/compounds increased Cu, P, and S (>100%, >50%, and >20%, respectively) in alfalfa shoots and decreased P and Fe in lettuce shoot (>50% and >50%, respectively, excluding Fe in CuCl2 treatment). Biochemical assays showed reduced catalase activity in alfalfa (root and shoot) and increased ascorbate peroxidase activity in roots of both plant species. Results suggest that Cu NPs/compounds not only reduced the size of the plants but altered nutrient content and enzyme activity in both plant species. PMID:25474419

  16. The Transcriptional Repressor MYB2 Regulates Both Spatial and Temporal Patterns of Proanthocyandin and Anthocyanin Pigmentation in Medicago truncatula.

    PubMed

    Jun, Ji Hyung; Liu, Chenggang; Xiao, Xirong; Dixon, Richard A

    2015-10-01

    Accumulation of anthocyanins and proanthocyanidins (PAs) is limited to specific cell types and developmental stages, but little is known about how antagonistically acting transcriptional regulators work together to determine temporal and spatial patterning of pigmentation at the cellular level, especially for PAs. Here, we characterize MYB2, a transcriptional repressor regulating both anthocyanin and PA biosynthesis in the model legume Medicago truncatula. MYB2 was strongly upregulated by MYB5, a major regulator of PA biosynthesis in M. truncatula and a component of MYB-basic helix loop helix-WD40 (MBW) activator complexes. Overexpression of MYB2 abolished anthocyanin and PA accumulation in M. truncatula hairy roots and Arabidopsis thaliana seeds, respectively. Anthocyanin deposition was expanded in myb2 mutant seedlings and flowers accompanied by increased anthocyanin content. PA mainly accumulated in the epidermal layer derived from the outer integument in the M. truncatula seed coat, starting from the hilum area. The area of PA accumulation and ANTHOCYANIDIN REDUCTASE expression was expanded into the seed body at the early stage of seed development in the myb2 mutant. Genetic, biochemical, and cell biological evidence suggests that MYB2 functions as part of a multidimensional regulatory network to define the temporal and spatial pattern of anthocyanin and PA accumulation linked to developmental processes.

  17. Drought stress provokes the down-regulation of methionine and ethylene biosynthesis pathways in Medicago truncatula roots and nodules.

    PubMed

    Larrainzar, Estíbaliz; Molenaar, Johanna A; Wienkoop, Stefanie; Gil-Quintana, Erena; Alibert, Bénédicte; Limami, Anis M; Arrese-Igor, Cesar; González, Esther M

    2014-09-01

    Symbiotic nitrogen fixation is one of the first physiological processes inhibited in legume plants under water-deficit conditions. Despite the progress made in the last decades, the molecular mechanisms behind this regulation are not fully understood yet. Recent proteomic work carried out in the model legume Medicago truncatula provided the first indications of a possible involvement of nodule methionine (Met) biosynthesis and related pathways in response to water-deficit conditions. To better understand this involvement, the drought-induced changes in expression and content of enzymes involved in the biosynthesis of Met, S-adenosyl-L-methionine (SAM) and ethylene in M. truncatula root and nodules were analyzed using targeted approaches. Nitrogen-fixing plants were subjected to a progressive water deficit and a subsequent recovery period. Besides the physiological characterization of the plants, the content of total sulphur, sulphate and main S-containing metabolites was measured. Results presented here show that S availability is not a limiting factor in the drought-induced decline of nitrogen fixation rates in M. truncatula plants and provide evidences for a down-regulation of the Met and ethylene biosynthesis pathways in roots and nodules in response to water-deficit conditions.

  18. GRAS Proteins Form a DNA Binding Complex to Induce Gene Expression during Nodulation Signaling in Medicago truncatula[W

    PubMed Central

    Hirsch, Sibylle; Kim, Jiyoung; Muñoz, Alfonso; Heckmann, Anne B.; Downie, J. Allan; Oldroyd, Giles E.D.

    2009-01-01

    The symbiotic association of legumes with rhizobia involves bacterially derived Nod factor, which is sufficient to activate the formation of nodules on the roots of the host plant. Perception of Nod factor by root hair cells induces calcium oscillations that are a component of the Nod factor signal transduction pathway. Perception of the calcium oscillations is a function of a calcium- and calmodulin-dependent protein kinase, and this activates nodulation gene expression via two GRAS domain transcriptional regulators, Nodulation Signaling Pathway1 (NSP1) and NSP2, and an ERF transcription factor required for nodulation. Here, we show that NSP1 and NSP2 form a complex that is associated with the promoters of early nodulin genes. We show that NSP1 binds directly to ENOD promoters through the novel cis-element AATTT. While NSP1 shows direct binding to the ENOD11 promoter in vitro, this association in vivo requires NSP2. The NSP1-NSP2 association with the ENOD11 promoter is enhanced following Nod factor elicitation. Mutations in the domain of NSP2 responsible for its interaction with NSP1 highlight the significance of the NSP1-NSP2 heteropolymer for nodulation signaling. Our work reveals direct binding of a GRAS protein complex to DNA and highlights the importance of the NSP1-NSP2 complex for efficient nodulation in the model legume Medicago truncatula. PMID:19252081

  19. Inhibition of nitrogen fixation in symbiotic Medicago truncatula upon Cd exposure is a local process involving leghaemoglobin

    PubMed Central

    Marino, Daniel

    2013-01-01

    Leguminous biological nitrogen fixation (BNF) is very sensitive to environmental fluctuations. It is still contentious how BNF is regulated under stress conditions. The local or systemic control of BNF and the role played by reactive oxygen species (ROS) in such regulation have still not been elucidated completely. Cadmium, which belongs to the so-called heavy metals, is one of the most toxic substances released into the environment. The mechanisms involved in Cd toxicity are still not completely understood but the overproduction of ROS is one of its characteristic symptoms. In this work, we used a split-root system approach to study nodule BNF and the antioxidant machinery’s response to the application of a mild Cd treatment on one side of a nodulated Medicago truncatula root system. Cd induced the majority of nodule antioxidants without generating any oxidative damage. Cd treatment also provoked BNF inhibition exclusively in nodules directly exposed to Cd, without provoking any effect on plant shoot biomass or chlorophyll content. The overall data suggest that the decline in BNF was not due to a generalized breakdown of the plant but to control exerted through leghaemoglobin/oxygen availability, affecting nitrogenase function. PMID:24151304

  20. The structures of cytosolic and plastid-located glutamine synthetases from Medicago truncatula reveal a common and dynamic architecture

    PubMed Central

    Torreira, Eva; Seabra, Ana Rita; Marriott, Hazel; Zhou, Min; Llorca, Óscar; Robinson, Carol V.; Carvalho, Helena G.; Fernández-Tornero, Carlos; Pereira, Pedro José Barbosa

    2014-01-01

    The first step of nitrogen assimilation in higher plants, the energy-driven incorporation of ammonia into glutamate, is catalyzed by glutamine synthetase. This central process yields the readily metabolizable glutamine, which in turn is at the basis of all subsequent biosynthesis of nitrogenous compounds. The essential role performed by glutamine synthetase makes it a prime target for herbicidal compounds, but also a suitable intervention point for the improvement of crop yields. Although the majority of crop plants are dicotyledonous, little is known about the structural organization of glutamine synthetase in these organisms and about the functional differences between the different isoforms. Here, the structural characterization of two glutamine synthetase isoforms from the model legume Medicago truncatula is reported: the crystallographic structure of cytoplasmic GSII-1a and an electron cryomicroscopy reconstruction of plastid-located GSII-2a. Together, these structural models unveil a decameric organization of dicotyledonous glutamine synthetase, with two pentameric rings weakly connected by inter-ring loops. Moreover, rearrangement of these dynamic loops changes the relative orientation of the rings, suggesting a zipper-like mechanism for their assembly into a decameric enzyme. Finally, the atomic structure of M. truncatula GSII-1a provides important insights into the structural determinants of herbicide resistance in this family of enzymes, opening new avenues for the development of herbicide-resistant plants. PMID:24699643

  1. Flavonoids and Auxin Transport Inhibitors Rescue Symbiotic Nodulation in the Medicago truncatula Cytokinin Perception Mutant cre1

    PubMed Central

    Ng, Jason Liang Pin; Hassan, Samira; Truong, Thy T.; Hocart, Charles H.; Laffont, Carole; Frugier, Florian; Mathesius, Ulrike

    2015-01-01

    Initiation of symbiotic nodules in legumes requires cytokinin signaling, but its mechanism of action is largely unknown. Here, we tested whether the failure to initiate nodules in the Medicago truncatula cytokinin perception mutant cre1 (cytokinin response1) is due to its altered ability to regulate auxin transport, auxin accumulation, and induction of flavonoids. We found that in the cre1 mutant, symbiotic rhizobia cannot locally alter acro- and basipetal auxin transport during nodule initiation and that these mutants show reduced auxin (indole-3-acetic acid) accumulation and auxin responses compared with the wild type. Quantification of flavonoids, which can act as endogenous auxin transport inhibitors, showed a deficiency in the induction of free naringenin, isoliquiritigenin, quercetin, and hesperetin in cre1 roots compared with wild-type roots 24 h after inoculation with rhizobia. Coinoculation of roots with rhizobia and the flavonoids naringenin, isoliquiritigenin, and kaempferol, or with the synthetic auxin transport inhibitor 2,3,5,-triiodobenzoic acid, rescued nodulation efficiency in cre1 mutants and allowed auxin transport control in response to rhizobia. Our results suggest that CRE1-dependent cytokinin signaling leads to nodule initiation through the regulation of flavonoid accumulation required for local alteration of polar auxin transport and subsequent auxin accumulation in cortical cells during the early stages of nodulation. PMID:26253705

  2. Relationship between photosynthetic capacity, nitrogen assimilation and nodule metabolism in alfalfa (Medicago sativa) grown with sewage sludge.

    PubMed

    Antolín, M Carmen; Fiasconaro, M Laura; Sánchez-Díaz, Manuel

    2010-10-15

    Sewage sludge has been used as N fertilizer because it contains some of inorganic N, principally as nitrate and ammonium ions. However, sewage sludge addition to legumes could result in impaired nodule metabolism due to the presence of inorganic N from sludge. A greenhouse experiment was conducted to examine the effects of sewage sludge on growth, photosynthesis, nitrogen assimilation and nodule metabolism in alfalfa (Medicago sativa L. cv. Aragón). Plants were grown in pots with a mixture of perlite and vermiculite (2:1, v/v). The experiment included three treatments: (1) plants inoculated with rhizobia and amended with sewage sludge at rate of 10% (w/w) (RS); (2) plants inoculated with rhizobia without any amendment (R); and (3) non-inoculated plants fed with ammonium nitrate (N). N(2)-fixing plants had lower growth and sucrose phosphate synthase activity but higher photosynthesis than nitrate-fed plants because they compensated the carbon cost of the rhizobia. However, sewage sludge-treated plants evidenced a loss of carbon sink strength due to N(2) fixation by means of decreased photosynthetic capacity, leaf chlorophylls and N concentration in comparison to untreated plants. Sewage sludge did no affect nodulation but decreased nodule enzyme activities involved in carbon and N metabolisms that may lead to accumulation of toxic N-compounds. PMID:20591568

  3. Arbuscular mycorrhizal symbiosis influences arsenic accumulation and speciation in Medicago truncatula L. in arsenic-contaminated soil.

    PubMed

    Zhang, Xin; Ren, Bai-Hui; Wu, Song-Lin; Sun, Yu-Qing; Lin, Ge; Chen, Bao-Dong

    2015-01-01

    In two pot experiments, wild type and a non-mycorrhizal mutant (TR25:3-1) of Medicago truncatula were grown in arsenic (As)-contaminated soil to investigate the influences of arbuscular mycorrhizal fungi (AMF) on As accumulation and speciation in host plants. The results indicated that the plant biomass of M. truncatula was dramatically increased by AM symbiosis. Mycorrhizal colonization significantly increased phosphorus concentrations and decreased As concentrations in plants. Moreover, mycorrhizal colonization generally increased the percentage of arsenite in total As both in shoots and roots, while dimethylarsenic acid (DMA) was only detected in shoots of mycorrhizal plants. The results suggested that AMF are most likely to get involved in the methylating of inorganic As into less toxic organic DMA and also in the reduction of arsenate to arsenite. The study allowed a deeper insight into the As detoxification mechanisms in AM associations. By using the mutant M. truncatula, we demonstrated the importance of AMF in plant As tolerance under natural conditions. PMID:25016555

  4. Identification of transcriptome involved in atrazine detoxification and degradation in alfalfa (Medicago sativa) exposed to realistic environmental contamination.

    PubMed

    Zhang, Jing Jing; Lu, Yi Chen; Zhang, Shu Hao; Lu, Feng Fan; Yang, Hong

    2016-08-01

    Plants are constantly exposed to a variety of toxic compounds (or xenobiotics) such as pesticides (or herbicides). Atrazine (ATZ) as herbicide has become one of the environmental contaminants due to its intensive use during crop production. Plants have evolved strategies to cope with the adverse impact of ATZ. However, the mechanism for ATZ degradation and detoxification in plants is largely unknown. Here we employed a global RNA-sequencing (RNA-Seq) strategy to dissect transcriptome variation in alfalfa (Medicago sativa) exposed to ATZ. Four libraries were constructed including Root-ATZ (root control, ATZ-free), Shoot-ATZ, Root+ATZ (root treated with ATZ) and Shoot+ATZ. Hierarchical clustering was performed to display the expression patterns for all differentially expressed genes (DEGs) under ATZ exposure. Transcripts involved in ATZ detoxification, stress responses (e.g. oxidation and reduction, conjugation and hydrolytic reactions), and regulations of cysteine biosynthesis were identified. Several genes encoding glycosyltransferases, glutathione S-transferases or ABC transporters were up-regulated notably. Also, many other genes involved in oxidation-reduction, conjugation, and hydrolysis for herbicide degradation were differentially expressed. These results suggest that ATZ in alfalfa can be detoxified or degraded through different pathways. The expression patterns of some DEGs by high-throughput sequencing were well confirmed by qRT-PCR. Our results not only highlight the transcriptional complexity in alfalfa exposed to ATZ but represent a major improvement for analyzing transcriptional changes on a large scale as well.

  5. Genetic Mapping of a Major Resistance Gene to Pea Aphid (Acyrthosipon pisum) in the Model Legume Medicago truncatula

    PubMed Central

    Kamphuis, Lars G.; Guo, Su-Min; Gao, Ling-Ling; Singh, Karam B.

    2016-01-01

    Resistance to the Australian pea aphid (PA; Acyrthosiphon pisum) biotype in cultivar Jester of the model legume Medicago truncatula is mediated by a single dominant gene and is phloem-mediated. The genetic map position for this resistance gene, APR (Acyrthosiphon pisum resistance), is provided and shows that APR maps 39 centiMorgans (cM) distal of the A. kondoi resistance (AKR) locus, which mediates resistance to a closely related species of the same genus bluegreen aphid (A. kondoi). The APR region on chromosome 3 is dense in classical nucleotide binding site leucine-rich repeats (NLRs) and overlaps with the region harbouring the RAP1 gene which confers resistance to a European PA biotype in the accession Jemalong A17. Further screening of a core collection of M. truncatula accessions identified seven lines with strong resistance to PA. Allelism experiments showed that the single dominant resistance to PA in M. truncatula accessions SA10481 and SA1516 are allelic to SA10733, the donor of the APR locus in cultivar Jester. While it remains unclear whether there are multiple PA resistance genes in an R-gene cluster or the resistance loci identified in the other M. truncatula accessions are allelic to APR, the introgression of APR into current M. truncatula cultivars will provide more durable resistance to PA. PMID:27483247

  6. Application of sodium nitroprusside results in distinct antioxidant gene expression patterns in leaves of mature and senescing Medicago truncatula plants.

    PubMed

    Fotopoulos, Vasileios; Antoniou, Chrystalla; Filippou, Panagiota; Mylona, Photini; Fasoula, Dionysia; Ioannides, Ioannis; Polidoros, Alexios

    2014-07-01

    Sodium nitroprusside (SNP) represents one of the most commonly used NO donors in biological sciences, which acts as a signal molecule in plants responsible for the regulation of the expression of many defense-related enzymes. This study attempts to provide novel insight into the effect of application of low (100 μΜ) and high (2.5 mM) concentrations of SNP on antioxidant gene expression (cAPX, GST, FeSOD, CAT, and AOX) in mature (40 day) and senescing (65 day) Medicago truncatula plants. Quantitative real-time RT-PCR suggests that low concentration of SNP applied in mature leaves leads to an overall induction of antioxidant gene expression, while increasing concentration results in suppression of these genes. Conversely, older plants demonstrate a much more variable regulation which appears to be time dependent. The observed transcriptional regulation pattern in mature M. truncatula plants comes in support of the previously documented protective or damaging effect of SNP depending on concentration applied, whereas senescing M. truncatula plants demonstrated a general suppression in antioxidant gene expression levels regardless of SNP concentration, indicative of reduced overall plant defense capacity against free radicals.

  7. Deep-sequencing transcriptome analysis of field-grown Medicago sativa L. crown buds acclimated to freezing stress.

    PubMed

    Song, Lili; Jiang, Lin; Chen, Yue; Shu, Yongjun; Bai, Yan; Guo, Changhong

    2016-09-01

    Medicago sativa L. (alfalfa) 'Zhaodong' is an important forage legume that can safely survive in northern China where winter temperatures reach as low as -30 °C. Survival of alfalfa following freezing stress depends on the amount and revival ability of crown buds. In order to investigate the molecular mechanisms of frost tolerance in alfalfa, we used transcriptome sequencing technology and bioinformatics strategies to analyze crown buds of field-grown alfalfa during winter. We statistically identified a total of 5605 differentially expressed genes (DEGs) involved in freezing stress including 1900 upregulated and 3705 downregulated DEGs. We validated 36 candidate DEGs using qPCR to confirm the accuracy of the RNA-seq data. Unlike other recent studies, this study employed alfalfa plants grown in the natural environment. Our results indicate that not only the CBF orthologs but also membrane proteins, hormone signal transduction pathways, and ubiquitin-mediated proteolysis pathways indicate the presence of a special freezing adaptation mechanism in alfalfa. The antioxidant defense system may rapidly confer freezing tolerance to alfalfa. Importantly, biosynthesis of secondary metabolites and phenylalanine metabolism, which is of potential importance in coordinating freezing tolerance with growth and development, were downregulated in subzero temperatures. The adaptive mechanism for frost tolerance is a complex multigenic process that is not well understood. This systematic analysis provided an in-depth view of stress tolerance mechanisms in alfalfa. PMID:27272950

  8. Flavonoids and Auxin Transport Inhibitors Rescue Symbiotic Nodulation in the Medicago truncatula Cytokinin Perception Mutant cre1.

    PubMed

    Ng, Jason Liang Pin; Hassan, Samira; Truong, Thy T; Hocart, Charles H; Laffont, Carole; Frugier, Florian; Mathesius, Ulrike

    2015-08-01

    Initiation of symbiotic nodules in legumes requires cytokinin signaling, but its mechanism of action is largely unknown. Here, we tested whether the failure to initiate nodules in the Medicago truncatula cytokinin perception mutant cre1 (cytokinin response1) is due to its altered ability to regulate auxin transport, auxin accumulation, and induction of flavonoids. We found that in the cre1 mutant, symbiotic rhizobia cannot locally alter acro- and basipetal auxin transport during nodule initiation and that these mutants show reduced auxin (indole-3-acetic acid) accumulation and auxin responses compared with the wild type. Quantification of flavonoids, which can act as endogenous auxin transport inhibitors, showed a deficiency in the induction of free naringenin, isoliquiritigenin, quercetin, and hesperetin in cre1 roots compared with wild-type roots 24 h after inoculation with rhizobia. Coinoculation of roots with rhizobia and the flavonoids naringenin, isoliquiritigenin, and kaempferol, or with the synthetic auxin transport inhibitor 2,3,5,-triiodobenzoic acid, rescued nodulation efficiency in cre1 mutants and allowed auxin transport control in response to rhizobia. Our results suggest that CRE1-dependent cytokinin signaling leads to nodule initiation through the regulation of flavonoid accumulation required for local alteration of polar auxin transport and subsequent auxin accumulation in cortical cells during the early stages of nodulation.

  9. Identification of transcriptome involved in atrazine detoxification and degradation in alfalfa (Medicago sativa) exposed to realistic environmental contamination.

    PubMed

    Zhang, Jing Jing; Lu, Yi Chen; Zhang, Shu Hao; Lu, Feng Fan; Yang, Hong

    2016-08-01

    Plants are constantly exposed to a variety of toxic compounds (or xenobiotics) such as pesticides (or herbicides). Atrazine (ATZ) as herbicide has become one of the environmental contaminants due to its intensive use during crop production. Plants have evolved strategies to cope with the adverse impact of ATZ. However, the mechanism for ATZ degradation and detoxification in plants is largely unknown. Here we employed a global RNA-sequencing (RNA-Seq) strategy to dissect transcriptome variation in alfalfa (Medicago sativa) exposed to ATZ. Four libraries were constructed including Root-ATZ (root control, ATZ-free), Shoot-ATZ, Root+ATZ (root treated with ATZ) and Shoot+ATZ. Hierarchical clustering was performed to display the expression patterns for all differentially expressed genes (DEGs) under ATZ exposure. Transcripts involved in ATZ detoxification, stress responses (e.g. oxidation and reduction, conjugation and hydrolytic reactions), and regulations of cysteine biosynthesis were identified. Several genes encoding glycosyltransferases, glutathione S-transferases or ABC transporters were up-regulated notably. Also, many other genes involved in oxidation-reduction, conjugation, and hydrolysis for herbicide degradation were differentially expressed. These results suggest that ATZ in alfalfa can be detoxified or degraded through different pathways. The expression patterns of some DEGs by high-throughput sequencing were well confirmed by qRT-PCR. Our results not only highlight the transcriptional complexity in alfalfa exposed to ATZ but represent a major improvement for analyzing transcriptional changes on a large scale as well. PMID:27092973

  10. Jasmonate Signalling and Defence Responses in the Model Legume Medicago truncatula-A Focus on Responses to Fusarium Wilt Disease.

    PubMed

    Thatcher, Louise F; Gao, Ling-Ling; Singh, Karam B

    2016-02-05

    Jasmonate (JA)-mediated defences play important roles in host responses to pathogen attack, in particular to necrotrophic fungal pathogens that kill host cells in order to extract nutrients and live off the dead plant tissue. The root-infecting fungal pathogen Fusarium oxysporum initiates a necrotrophic growth phase towards the later stages of its lifecycle and is responsible for devastating Fusarium wilt disease on numerous legume crops worldwide. Here we describe the use of the model legume Medicago truncatula to study legume-F. oxysporum interactions and compare and contrast this against knowledge from other model pathosystems, in particular Arabidopsis thaliana-F. oxysporum interactions. We describe publically-available genomic, transcriptomic and genetic (mutant) resources developed in M. truncatula that enable dissection of host jasmonate responses and apply aspects of these herein during the M. truncatula--F. oxysporum interaction. Our initial results suggest not all components of JA-responses observed in M. truncatula are shared with Arabidopsis in response to F. oxysporum infection.

  11. Jasmonate Signalling and Defence Responses in the Model Legume Medicago truncatula—A Focus on Responses to Fusarium Wilt Disease

    PubMed Central

    Thatcher, Louise F.; Gao, Ling-Ling; Singh, Karam B.

    2016-01-01

    Jasmonate (JA)-mediated defences play important roles in host responses to pathogen attack, in particular to necrotrophic fungal pathogens that kill host cells in order to extract nutrients and live off the dead plant tissue. The root-infecting fungal pathogen Fusarium oxysporum initiates a necrotrophic growth phase towards the later stages of its lifecycle and is responsible for devastating Fusarium wilt disease on numerous legume crops worldwide. Here we describe the use of the model legume Medicago truncatula to study legume–F. oxysporum interactions and compare and contrast this against knowledge from other model pathosystems, in particular Arabidopsis thaliana–F. oxysporum interactions. We describe publically-available genomic, transcriptomic and genetic (mutant) resources developed in M. truncatula that enable dissection of host jasmonate responses and apply aspects of these herein during the M. truncatula-–F. oxysporum interaction. Our initial results suggest not all components of JA-responses observed in M. truncatula are shared with Arabidopsis in response to F. oxysporum infection. PMID:27135231

  12. Identification of molecular markers associated with Verticillium wilt resistance in alfalfa (Medicago sativa L.) using high-resolution melting.

    PubMed

    Zhang, Tiejun; Yu, Long-Xi; McCord, Per; Miller, David; Bhamidimarri, Suresh; Johnson, David; Monteros, Maria J; Ho, Julie; Reisen, Peter; Samac, Deborah A

    2014-01-01

    Verticillium wilt, caused by the soilborne fungus, Verticillium alfalfae, is one of the most serious diseases of alfalfa (Medicago sativa L.) worldwide. To identify loci associated with resistance to Verticillium wilt, a bulk segregant analysis was conducted in susceptible or resistant pools constructed from 13 synthetic alfalfa populations, followed by association mapping in two F1 populations consisted of 352 individuals. Simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) markers were used for genotyping. Phenotyping was done by manual inoculation of the pathogen to replicated cloned plants of each individual and disease severity was scored using a standard scale. Marker-trait association was analyzed by TASSEL. Seventeen SNP markers significantly associated with Verticillium wilt resistance were identified and they were located on chromosomes 1, 2, 4, 7 and 8. SNP markers identified on chromosomes 2, 4 and 7 co-locate with regions of Verticillium wilt resistance loci reported in M. truncatula. Additional markers identified on chromosomes 1 and 8 located the regions where no Verticillium resistance locus has been reported. This study highlights the value of SNP genotyping by high resolution melting to identify the disease resistance loci in tetraploid alfalfa. With further validation, the markers identified in this study could be used for improving resistance to Verticillium wilt in alfalfa breeding programs.

  13. Identification of Molecular Markers Associated with Verticillium Wilt Resistance in Alfalfa (Medicago Sativa L.) Using High-Resolution Melting

    PubMed Central

    Zhang, Tiejun; Yu, Long-Xi; McCord, Per; Miller, David; Bhamidimarri, Suresh; Johnson, David; Monteros, Maria J.; Ho, Julie; Reisen, Peter; Samac, Deborah A.

    2014-01-01

    Verticillium wilt, caused by the soilborne fungus, Verticillium alfalfae, is one of the most serious diseases of alfalfa (Medicago sativa L.) worldwide. To identify loci associated with resistance to Verticillium wilt, a bulk segregant analysis was conducted in susceptible or resistant pools constructed from 13 synthetic alfalfa populations, followed by association mapping in two F1 populations consisted of 352 individuals. Simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) markers were used for genotyping. Phenotyping was done by manual inoculation of the pathogen to replicated cloned plants of each individual and disease severity was scored using a standard scale. Marker-trait association was analyzed by TASSEL. Seventeen SNP markers significantly associated with Verticillium wilt resistance were identified and they were located on chromosomes 1, 2, 4, 7 and 8. SNP markers identified on chromosomes 2, 4 and 7 co-locate with regions of Verticillium wilt resistance loci reported in M. truncatula. Additional markers identified on chromosomes 1 and 8 located the regions where no Verticillium resistance locus has been reported. This study highlights the value of SNP genotyping by high resolution melting to identify the disease resistance loci in tetraploid alfalfa. With further validation, the markers identified in this study could be used for improving resistance to Verticillium wilt in alfalfa breeding programs. PMID:25536106

  14. DASH transcription factor impacts Medicago truncatula seed size by its action on embryo morphogenesis and auxin homeostasis

    PubMed Central

    Noguero, Mélanie; Le Signor, Christine; Vernoud, Vanessa; Bandyopadhyay, Kaustav; Sanchez, Myriam; Fu, Chunxiang; Torres-Jerez, Ivone; Wen, Jiangqi; Mysore, Kirankumar S; Gallardo, Karine; Udvardi, Michael; Thompson, Richard; Verdier, Jerome

    2015-01-01

    The endosperm plays a pivotal role in the integration between component tissues of molecular signals controlling seed development. It has been shown to participate in the regulation of embryo morphogenesis and ultimately seed size determination. However, the molecular mechanisms that modulate seed size are still poorly understood especially in legumes. DASH (DOF Acting in Seed embryogenesis and Hormone accumulation) is a DOF transcription factor (TF) expressed during embryogenesis in the chalazal endosperm of the Medicago truncatula seed. Phenotypic characterization of three independent dash mutant alleles revealed a role for this TF in the prevention of early seed abortion and the determination of final seed size. Strong loss-of-function alleles cause severe defects in endosperm development and lead to embryo growth arrest at the globular stage. Transcriptomic analysis of dash pods versus wild-type (WT) pods revealed major transcriptional changes and highlighted genes that are involved in auxin transport and perception as mainly under-expressed in dash mutant pods. Interestingly, the exogenous application of auxin alleviated the seed-lethal phenotype, whereas hormonal dosage revealed a much higher auxin content in dash pods compared with WT. Together these results suggested that auxin transport/signaling may be affected in the dash mutant and that aberrant auxin distribution may contribute to the defect in embryogenesis resulting in the final seed size phenotype. PMID:25492260

  15. Allelopathic Potential of Switchgrass ( Panicum virgatum L.) on Perennial Ryegrass ( Lolium perenne L.) and Alfalfa ( Medicago sativa L.)

    NASA Astrophysics Data System (ADS)

    Shui, Junfeng; An, Yu; Ma, Yongqing; Ichizen, Nobumasa

    2010-10-01

    This study investigated allelopathy and its chemical basis in nine switchgrass ( Panicum virgatum L.) accessions. Perennial ryegrass ( Lolium perenne L.) and alfalfa ( Medicago sativa L.) were used as test species. Undiluted aqueous extracts (5 g plant tissue in 50 ml water) from the shoots and roots of most of the switchgrass accessions inhibited the germination and growth of the test species. However, the allelopathic effect of switchgrass declined when extracts were diluted 5- or 50-fold. Seedling growth was more sensitive than seed germination as an indicator of allelopathic effect. Allelopathic effect was related to switchgrass ecotype but not related to ploidy level. Upland accessions displayed stronger allelopathic potential than lowland accessions. The aqueous extract from one switchgrass accession was separated into phenols, organic acids, neutral chemicals, and alkaloids, and then these fractions were bioassayed to test for allelopathic potential. Alkaloids had the strongest allelopathic effect among the four chemical fractions. In summary, the results indicated that switchgrass has allelopathic potential; however, there is not enough evidence to conclude that allelopathic advantage is the main factor that has contributed to the successful establishment of switchgrass on China’s Loess Plateau.

  16. The structures of cytosolic and plastid-located glutamine synthetases from Medicago truncatula reveal a common and dynamic architecture.

    PubMed

    Torreira, Eva; Seabra, Ana Rita; Marriott, Hazel; Zhou, Min; Llorca, Óscar; Robinson, Carol V; Carvalho, Helena G; Fernández-Tornero, Carlos; Pereira, Pedro José Barbosa

    2014-04-01

    The first step of nitrogen assimilation in higher plants, the energy-driven incorporation of ammonia into glutamate, is catalyzed by glutamine synthetase. This central process yields the readily metabolizable glutamine, which in turn is at the basis of all subsequent biosynthesis of nitrogenous compounds. The essential role performed by glutamine synthetase makes it a prime target for herbicidal compounds, but also a suitable intervention point for the improvement of crop yields. Although the majority of crop plants are dicotyledonous, little is known about the structural organization of glutamine synthetase in these organisms and about the functional differences between the different isoforms. Here, the structural characterization of two glutamine synthetase isoforms from the model legume Medicago truncatula is reported: the crystallographic structure of cytoplasmic GSII-1a and an electron cryomicroscopy reconstruction of plastid-located GSII-2a. Together, these structural models unveil a decameric organization of dicotyledonous glutamine synthetase, with two pentameric rings weakly connected by inter-ring loops. Moreover, rearrangement of these dynamic loops changes the relative orientation of the rings, suggesting a zipper-like mechanism for their assembly into a decameric enzyme. Finally, the atomic structure of M. truncatula GSII-1a provides important insights into the structural determinants of herbicide resistance in this family of enzymes, opening new avenues for the development of herbicide-resistant plants. PMID:24699643

  17. Multiple mutualist effects on genomewide expression in the tripartite association between Medicago truncatula, nitrogen-fixing bacteria and mycorrhizal fungi.

    PubMed

    Afkhami, Michelle E; Stinchcombe, John R

    2016-10-01

    While all species interact with multiple mutualists, the fitness consequences and molecular mechanisms underlying these interactions remain largely unknown. We combined factorial ecological experiments with genomewide expression analyses to examine the phenotypic and transcriptomic responses of model legume Medicago truncatula to rhizobia and mycorrhizal fungi. We found synergistic effects of these mutualists on plant performance and examined unique features of plant gene expression responses to multiple mutualists. There were genomewide signatures of mutualists and multiple mutualists on expression, with partners often affecting unique sets of genes. Mycorrhizal fungi had stronger effects on plant expression than rhizobia, with 70% of differentially expressed genes affected by fungi. Fungal and bacterial mutualists had joint effects on 10% of differentially expressed genes, including unexpected, nonadditive effects on some genes with important functions such as nutrient metabolism. For a subset of genes, interacting with multiple mutualists even led to reversals in the direction of expression (shifts from up to downregulation) compared to interacting with single mutualists. Rhizobia also affected the expression of several mycorrhizal genes, including those involved in nutrient transfer to host plants, indicating that partner species can also impact each other's molecular phenotypes. Collectively, these data illustrate the diverse molecular mechanisms and transcriptional responses associated with the synergistic benefits of multiple mutualists.

  18. MALDI mass spectrometry-assisted molecular imaging of metabolites during nitrogen fixation in the Medicago truncatula-Sinorhizobium meliloti symbiosis.

    PubMed

    Ye, Hui; Gemperline, Erin; Venkateshwaran, Muthusubramanian; Chen, Ruibing; Delaux, Pierre-Marc; Howes-Podoll, Maegen; Ané, Jean-Michel; Li, Lingjun

    2013-07-01

    Symbiotic associations between leguminous plants and nitrogen-fixing rhizobia culminate in the formation of specialized organs called root nodules, in which the rhizobia fix atmospheric nitrogen and transfer it to the plant. Efficient biological nitrogen fixation depends on metabolites produced by and exchanged between both partners. The Medicago truncatula-Sinorhizobium meliloti association is an excellent model for dissecting this nitrogen-fixing symbiosis because of the availability of genetic information for both symbiotic partners. Here, we employed a powerful imaging technique - matrix-assisted laser desorption/ionization (MALDI)/mass spectrometric imaging (MSI) - to study metabolite distribution in roots and root nodules of M. truncatula during nitrogen fixation. The combination of an efficient, novel MALDI matrix [1,8-bis(dimethyl-amino) naphthalene, DMAN] with a conventional matrix 2,5-dihydroxybenzoic acid (DHB) allowed detection of a large array of organic acids, amino acids, sugars, lipids, flavonoids and their conjugates with improved coverage. Ion density maps of representative metabolites are presented and correlated with the nitrogen fixation process. We demonstrate differences in metabolite distribution between roots and nodules, and also between fixing and non-fixing nodules produced by plant and bacterial mutants. Our study highlights the benefits of using MSI for detecting differences in metabolite distributions in plant biology.

  19. Different cytokinin histidine kinase receptors regulate nodule initiation as well as later nodule developmental stages in Medicago truncatula.

    PubMed

    Boivin, Stéphane; Kazmierczak, Théophile; Brault, Mathias; Wen, Jiangqi; Gamas, Pascal; Mysore, Kirankumar S; Frugier, Florian

    2016-10-01

    Legume plants adapt to low nitrogen by developing an endosymbiosis with nitrogen-fixing soil bacteria to form a new specific organ: the nitrogen-fixing nodule. In the Medicago truncatula model legume, the MtCRE1 cytokinin receptor is essential for this symbiotic interaction. As three other putative CHASE-domain containing histidine kinase (CHK) cytokinin receptors exist in M. truncatula, we determined their potential contribution to this symbiotic interaction. The four CHKs have extensive redundant expression patterns at early nodulation stages but diverge in differentiated nodules, even though MtCHK1/MtCRE1 has the strongest expression at all stages. Mutant and knock-down analyses revealed that other CHKs than MtCHK1/CRE1 are positively involved in nodule initiation, which explains the delayed nodulation phenotype of the chk1/cre1 mutant. In addition, cre1 nodules exhibit an increased growth, whereas other chk mutants have no detectable phenotype, and the maintained nitrogen fixation capacity in cre1 requires other CHK genes. Interestingly, an AHK4/CRE1 genomic locus from the aposymbiotic Arabidopsis plant rescues nodule initiation but not the nitrogen fixation capacity. This indicates that different CHK cytokinin signalling pathways regulate not only nodule initiation but also later developmental stages, and that legume-specific determinants encoded by the MtCRE1 gene are required for later nodulation stages than initiation. PMID:27341695

  20. Deep-sequencing transcriptome analysis of field-grown Medicago sativa L. crown buds acclimated to freezing stress.

    PubMed

    Song, Lili; Jiang, Lin; Chen, Yue; Shu, Yongjun; Bai, Yan; Guo, Changhong

    2016-09-01

    Medicago sativa L. (alfalfa) 'Zhaodong' is an important forage legume that can safely survive in northern China where winter temperatures reach as low as -30 °C. Survival of alfalfa following freezing stress depends on the amount and revival ability of crown buds. In order to investigate the molecular mechanisms of frost tolerance in alfalfa, we used transcriptome sequencing technology and bioinformatics strategies to analyze crown buds of field-grown alfalfa during winter. We statistically identified a total of 5605 differentially expressed genes (DEGs) involved in freezing stress including 1900 upregulated and 3705 downregulated DEGs. We validated 36 candidate DEGs using qPCR to confirm the accuracy of the RNA-seq data. Unlike other recent studies, this study employed alfalfa plants grown in the natural environment. Our results indicate that not only the CBF orthologs but also membrane proteins, hormone signal transduction pathways, and ubiquitin-mediated proteolysis pathways indicate the presence of a special freezing adaptation mechanism in alfalfa. The antioxidant defense system may rapidly confer freezing tolerance to alfalfa. Importantly, biosynthesis of secondary metabolites and phenylalanine metabolism, which is of potential importance in coordinating freezing tolerance with growth and development, were downregulated in subzero temperatures. The adaptive mechanism for frost tolerance is a complex multigenic process that is not well understood. This systematic analysis provided an in-depth view of stress tolerance mechanisms in alfalfa.

  1. Genetic Mapping of a Major Resistance Gene to Pea Aphid (Acyrthosipon pisum) in the Model Legume Medicago truncatula.

    PubMed

    Kamphuis, Lars G; Guo, Su-Min; Gao, Ling-Ling; Singh, Karam B

    2016-01-01

    Resistance to the Australian pea aphid (PA; Acyrthosiphon pisum) biotype in cultivar Jester of the model legume Medicago truncatula is mediated by a single dominant gene and is phloem-mediated. The genetic map position for this resistance gene, APR (Acyrthosiphon pisum resistance), is provided and shows that APR maps 39 centiMorgans (cM) distal of the A. kondoi resistance (AKR) locus, which mediates resistance to a closely related species of the same genus bluegreen aphid (A. kondoi). The APR region on chromosome 3 is dense in classical nucleotide binding site leucine-rich repeats (NLRs) and overlaps with the region harbouring the RAP1 gene which confers resistance to a European PA biotype in the accession Jemalong A17. Further screening of a core collection of M. truncatula accessions identified seven lines with strong resistance to PA. Allelism experiments showed that the single dominant resistance to PA in M. truncatula accessions SA10481 and SA1516 are allelic to SA10733, the donor of the APR locus in cultivar Jester. While it remains unclear whether there are multiple PA resistance genes in an R-gene cluster or the resistance loci identified in the other M. truncatula accessions are allelic to APR, the introgression of APR into current M. truncatula cultivars will provide more durable resistance to PA. PMID:27483247

  2. Increase in 4-coumaryl alcohol units during lignification in alfalfa (Medicago sativa) alters the extractability and molecular weight of lignin.

    PubMed

    Ziebell, Angela; Gracom, Kristen; Katahira, Rui; Chen, Fang; Pu, Yunqiao; Ragauskas, Art; Dixon, Richard A; Davis, Mark

    2010-12-10

    The lignin content of biomass can impact the ease and cost of biomass processing. Lignin reduction through breeding and genetic modification therefore has potential to reduce costs in biomass-processing industries (e.g. pulp and paper, forage, and lignocellulosic ethanol). We investigated compositional changes in two low-lignin alfalfa (Medicago sativa) lines with antisense down-regulation of p-coumarate 3-hydroxylase (C3H) or hydroxycinnamoyl-CoA:shikimate hydroxycinnamoyltransferase (HCT). We investigated whether the difference in reactivity during lignification of 4-coumaryl alcohol (H) monomers versus the naturally dominant sinapyl alcohol and coniferyl alcohol lignin monomers alters the lignin structure. Sequential base extraction readily reduced the H monomer content of the transgenic lines, leaving a residual lignin greatly enriched in H subunits; the extraction profile highlighted the difference between the control and transgenic lines. Gel permeation chromatography of isolated ball-milled lignin indicated significant changes in the weight average molecular weight distribution of the control versus transgenic lines (CTR1a, 6000; C3H4a, 5500; C3H9a, 4000; and HCT30a, 4000).

  3. Characterization of the plant growth promoting bacterium, Enterobacter cloacae MSR1, isolated from roots of non-nodulating Medicago sativa

    PubMed Central

    Khalifa, Ashraf Y.Z.; Alsyeeh, Abdel-Moneium; Almalki, Mohammed A.; Saleh, Farag A.

    2015-01-01

    The aim of the present study was to characterize the endophytic bacterial strain designated MSR1 that was isolated from inside the non-nodulating roots of Medicago sativa after surface-sterilization. MSR1 was identified as Enterobacter cloacae using both 16S rDNA gene sequence analysis and API20E biochemical identification system (Biomerieux, France). Furthermore, this bacterium was characterized using API50CH kit (Biomerieux, France) and tested for antibacterial activities against some food borne pathogens. The results showed that E. cloacae consumed certain carbohydrates such as glycerol, d-xylose, d-maltose and esculin melibiose as a sole carbon source and certain amino acids such as arginine, tryptophan ornithine as nitrogen source. Furthermore, MSR1 possessed multiple plant-growth promoting characteristics; phosphate solubility, production of phytohormones acetoin and bioactive compounds. Inoculation of Pisum sativum with MSR1 significantly improved the growth parameters (the length and dry weight) of this economically important grain legume compared to the non-treated plants. To our knowledge, this is the first report addressing E. cloacae which exist in roots of alfalfa growing in Al-Ahsaa region. The results confirmed that E. cloacae exhibited traits for plant growth promoting and could be developed as an eco-friendly biofertilizer for P. sativum and probably for other important plant species in future. PMID:26858542

  4. RNA-Seq analysis discloses early senescence and nucleolar dysfunction triggered by Tdp1α depletion in Medicago truncatula.

    PubMed

    Donà, Mattia; Confalonieri, Massimo; Minio, Andrea; Biggiogera, Marco; Buttafava, Armando; Raimondi, Elena; Delledonne, Massimo; Ventura, Lorenzo; Sabatini, Maria Elisa; Macovei, Anca; Giraffa, Giorgio; Carbonera, Daniela; Balestrazzi, Alma

    2013-04-01

    An intron-spliced hairpin RNA approach was used for the targeted silencing of the MtTdp1α gene encoding the αisoform of tyrosyl-DNA phosphodiesterase 1 in Medicago truncatula Gaertn. Tyrosyl-DNA phosphodiesterase 1, involved in the repair of DNA topoisomerase I-mediated DNA damage, has been poorly investigated in plants. RNA-Seq analysis, carried out in the MtTdp1α-depleted plants, revealed different levels of transcriptional modulation (up- and down-regulation, alternative splicing, activation of alternative promoter) in genes involved in DNA damage sensing, DNA repair, and chromatin remodelling. It is suggested that the MtTdp1α gene has new, previously undetected roles in maintaining genome integrity. Up-regulation of senescence-associated genes and telomere shortening were observed. Moreover, impaired ribosome biogenesis indicated that the MtTdp1α gene is required for the nucleolar function. In agreement with the RNA-Seq data, transmission electron microscopy detected an altered nucleolar architecture in the MtTdp1α-depleted cells. Based on the reported data, a working hypothesis related to the occurrence of a nucleolar checkpoint in plant cells is proposed. PMID:23467834

  5. Characterization of the plant growth promoting bacterium, Enterobacter cloacae MSR1, isolated from roots of non-nodulating Medicago sativa.

    PubMed

    Khalifa, Ashraf Y Z; Alsyeeh, Abdel-Moneium; Almalki, Mohammed A; Saleh, Farag A

    2016-01-01

    The aim of the present study was to characterize the endophytic bacterial strain designated MSR1 that was isolated from inside the non-nodulating roots of Medicago sativa after surface-sterilization. MSR1 was identified as Enterobacter cloacae using both 16S rDNA gene sequence analysis and API20E biochemical identification system (Biomerieux, France). Furthermore, this bacterium was characterized using API50CH kit (Biomerieux, France) and tested for antibacterial activities against some food borne pathogens. The results showed that E. cloacae consumed certain carbohydrates such as glycerol, d-xylose, d-maltose and esculin melibiose as a sole carbon source and certain amino acids such as arginine, tryptophan ornithine as nitrogen source. Furthermore, MSR1 possessed multiple plant-growth promoting characteristics; phosphate solubility, production of phytohormones acetoin and bioactive compounds. Inoculation of Pisum sativum with MSR1 significantly improved the growth parameters (the length and dry weight) of this economically important grain legume compared to the non-treated plants. To our knowledge, this is the first report addressing E. cloacae which exist in roots of alfalfa growing in Al-Ahsaa region. The results confirmed that E. cloacae exhibited traits for plant growth promoting and could be developed as an eco-friendly biofertilizer for P. sativum and probably for other important plant species in future. PMID:26858542

  6. Multiple mutualist effects on genomewide expression in the tripartite association between Medicago truncatula, nitrogen-fixing bacteria and mycorrhizal fungi.

    PubMed

    Afkhami, Michelle E; Stinchcombe, John R

    2016-10-01

    While all species interact with multiple mutualists, the fitness consequences and molecular mechanisms underlying these interactions remain largely unknown. We combined factorial ecological experiments with genomewide expression analyses to examine the phenotypic and transcriptomic responses of model legume Medicago truncatula to rhizobia and mycorrhizal fungi. We found synergistic effects of these mutualists on plant performance and examined unique features of plant gene expression responses to multiple mutualists. There were genomewide signatures of mutualists and multiple mutualists on expression, with partners often affecting unique sets of genes. Mycorrhizal fungi had stronger effects on plant expression than rhizobia, with 70% of differentially expressed genes affected by fungi. Fungal and bacterial mutualists had joint effects on 10% of differentially expressed genes, including unexpected, nonadditive effects on some genes with important functions such as nutrient metabolism. For a subset of genes, interacting with multiple mutualists even led to reversals in the direction of expression (shifts from up to downregulation) compared to interacting with single mutualists. Rhizobia also affected the expression of several mycorrhizal genes, including those involved in nutrient transfer to host plants, indicating that partner species can also impact each other's molecular phenotypes. Collectively, these data illustrate the diverse molecular mechanisms and transcriptional responses associated with the synergistic benefits of multiple mutualists. PMID:27543961

  7. Rhizobial symbiosis effect on the growth, metal uptake, and antioxidant responses of Medicago lupulina under copper stress.

    PubMed

    Kong, Zhaoyu; Mohamad, Osama Abdalla; Deng, Zhenshan; Liu, Xiaodong; Glick, Bernard R; Wei, Gehong

    2015-08-01

    The effects of rhizobial symbiosis on the growth, metal uptake, and antioxidant responses of Medicago lupulina in the presence of 200 mg kg(-1) Cu(2+) throughout different stages of symbiosis development were studied. The symbiosis with Sinorhizobium meliloti CCNWSX0020 induced an increase in plant growth and nitrogen content irrespective of the presence of Cu(2+). The total amount of Cu uptake of inoculated plants significantly increased by 34.0 and 120.4% in shoots and roots, respectively, compared with non-inoculated plants. However, although the rhizobial symbiosis promoted Cu accumulation both in shoots and roots, the increase in roots was much higher than in shoots, thus decreasing the translocation factor and helping Cu phytostabilization. The rate of lipid peroxidation was significantly decreased in both shoots and roots of inoculated vs. non-inoculated plants when measured either 8, 13, or 18 days post-inoculation. In comparison with non-inoculated plants, the activities of superoxide dismutase and ascorbate peroxidase of shoots of inoculated plants exposed to excess Cu were significantly elevated at different stages of symbiosis development; similar increases occurred in the activities of superoxide dismutase, catalase, and glutathione reductase of inoculated roots. The symbiosis with S. meliloti CCNWSX0020 also upregulated the corresponding genes involved in antioxidant responses in the plants treated with excess Cu. The results indicated that the rhizobial symbiosis with S. meliloti CCNWSX0020 not only enhanced plant growth and metal uptake but also improved the responses of plant antioxidant defense to excess Cu stress.

  8. Arbuscular mycorrhizal symbiosis can mitigate the negative effects of night warming on physiological traits of Medicago truncatula L.

    PubMed

    Hu, Yajun; Wu, Songlin; Sun, Yuqing; Li, Tao; Zhang, Xin; Chen, Caiyan; Lin, Ge; Chen, Baodong

    2015-02-01

    Elevated night temperature, one of the main climate warming scenarios, can have profound effects on plant growth and metabolism. However, little attention has been paid to the potential role of mycorrhizal associations in plant responses to night warming, although it is well known that symbiotic fungi can protect host plants against various environmental stresses. In the present study, physiological traits of Medicago truncatula L. in association with the arbuscular mycorrhizal (AM) fungus Rhizophagus irregularis were investigated under simulated night warming. A constant increase in night temperature of 1.53 °C significantly reduced plant shoot and root biomass, flower and seed number, leaf sugar concentration, and shoot Zn and root P concentrations. However, the AM association essentially mitigated these negative effects of night warming by improving plant growth, especially through increased root biomass, root to shoot ratio, and shoot Zn and root P concentrations. A significant interaction was observed between R. irregularis inoculation and night warming in influencing both root sucrose concentration and expression of sucrose synthase (SusS) genes, suggesting that AM symbiosis and increased night temperature jointly regulated plant sugar metabolism. Night warming stimulated AM fungal colonization but did not influence arbuscule abundance, symbiosis-related plant or fungal gene expression, or growth of extraradical mycelium, indicating little effect of night warming on the development or functioning of AM symbiosis. These findings highlight the importance of mycorrhizal symbiosis in assisting plant resilience to climate warming.

  9. Jasmonate Signalling and Defence Responses in the Model Legume Medicago truncatula-A Focus on Responses to Fusarium Wilt Disease.

    PubMed

    Thatcher, Louise F; Gao, Ling-Ling; Singh, Karam B

    2016-01-01

    Jasmonate (JA)-mediated defences play important roles in host responses to pathogen attack, in particular to necrotrophic fungal pathogens that kill host cells in order to extract nutrients and live off the dead plant tissue. The root-infecting fungal pathogen Fusarium oxysporum initiates a necrotrophic growth phase towards the later stages of its lifecycle and is responsible for devastating Fusarium wilt disease on numerous legume crops worldwide. Here we describe the use of the model legume Medicago truncatula to study legume-F. oxysporum interactions and compare and contrast this against knowledge from other model pathosystems, in particular Arabidopsis thaliana-F. oxysporum interactions. We describe publically-available genomic, transcriptomic and genetic (mutant) resources developed in M. truncatula that enable dissection of host jasmonate responses and apply aspects of these herein during the M. truncatula--F. oxysporum interaction. Our initial results suggest not all components of JA-responses observed in M. truncatula are shared with Arabidopsis in response to F. oxysporum infection. PMID:27135231

  10. Medicago truncatula and Glycine max: Different Drought Tolerance and Similar Local Response of the Root Nodule Proteome

    PubMed Central

    2015-01-01

    Legume crops present important agronomical and environmental advantages mainly due to their capacity to reduce atmospheric N2 to ammonium via symbiotic nitrogen fixation (SNF). This process is very sensitive to abiotic stresses such as drought, but the mechanism underlying this response is not fully understood. The goal of the current work is to compare the drought response of two legumes with high economic impact and research importance, Medicago truncatula and Glycine max, by characterizing their root nodule proteomes. Our results show that, although M. truncatula exhibits lower water potential values under drought conditions compared to G. max, SNF declined analogously in the two legumes. Both of their nodule proteomes are very similar, and comparable down-regulation responses in the diverse protein functional groups were identified (mainly proteins related to the metabolism of carbon, nitrogen, and sulfur). We suggest lipoxygenases and protein turnover as newly recognized players in SNF regulation. Partial drought conditions applied to a split-root system resulted in the local down-regulation of the entire proteome of drought-stressed nodules in both legumes. The high degree of similarity between both legume proteomes suggests that the vast amount of research conducted on M. truncatula could be applied to economically important legume crops, such as soybean. PMID:26503705

  11. Adjustment of host cells for accommodation of symbiotic bacteria: vacuole defunctionalization, HOPS suppression, and TIP1g retargeting in Medicago.

    PubMed

    Gavrin, Aleksandr; Kaiser, Brent N; Geiger, Dietmar; Tyerman, Stephen D; Wen, Zhengyu; Bisseling, Ton; Fedorova, Elena E

    2014-09-01

    In legume-rhizobia symbioses, the bacteria in infected cells are enclosed in a plant membrane, forming organelle-like compartments called symbiosomes. Symbiosomes remain as individual units and avoid fusion with lytic vacuoles of host cells. We observed changes in the vacuole volume of infected cells and thus hypothesized that microsymbionts may cause modifications in vacuole formation or function. To examine this, we quantified the volumes and surface areas of plant cells, vacuoles, and symbiosomes in root nodules of Medicago truncatula and analyzed the expression and localization of VPS11 and VPS39, members of the HOPS vacuole-tethering complex. During the maturation of symbiosomes to become N2-fixing organelles, a developmental switch occurs and changes in vacuole features are induced. For example, we found that expression of VPS11 and VPS39 in infected cells is suppressed and host cell vacuoles contract, permitting the expansion of symbiosomes. Trafficking of tonoplast-targeted proteins in infected symbiotic cells is also altered, as shown by retargeting of the aquaporin TIP1g from the tonoplast membrane to the symbiosome membrane. This retargeting appears to be essential for the maturation of symbiosomes. We propose that these alterations in the function of the vacuole are key events in the adaptation of the plant cell to host intracellular symbiotic bacteria.

  12. Transcriptomic and metabolomic analyses identify a role for chlorophyll catabolism and phytoalexin during Medicago nonhost resistance against Asian soybean rust

    PubMed Central

    Ishiga, Yasuhiro; Rao Uppalapati, Srinivasa; Gill, Upinder S.; Huhman, David; Tang, Yuhong; Mysore, Kirankumar S.

    2015-01-01

    Asian soybean rust (ASR) caused by Phakopsora pachyrhizi is a devastating foliar disease affecting soybean production worldwide. Understanding nonhost resistance against ASR may provide an avenue to engineer soybean to confer durable resistance against ASR. We characterized a Medicago truncatula-ASR pathosystem to study molecular mechanisms of nonhost resistance. Although urediniospores formed appressoria and penetrated into epidermal cells of M. truncatula, P. pachyrhizi failed to sporulate. Transcriptomic analysis revealed the induction of phenylpropanoid, flavonoid and isoflavonoid metabolic pathway genes involved in the production of phytoalexin medicarpin in M. truncatula upon infection with P. pachyrhizi. Furthermore, genes involved in chlorophyll catabolism were induced during nonhost resistance. We further characterized one of the chlorophyll catabolism genes, Stay-green (SGR), and demonstrated that the M. truncatula sgr mutant and alfalfa SGR-RNAi lines showed hypersensitive-response-like enhanced cell death upon inoculation with P. pachyrhizi. Consistent with transcriptomic analysis, metabolomic analysis also revealed the accumulation of medicarpin and its intermediate metabolites. In vitro assay showed that medicarpin inhibited urediniospore germination and differentiation. In addition, several triterpenoid saponin glycosides accumulated in M. truncatula upon inoculation with P. pachyrhizi. In summary, using multi-omic approaches, we identified a correlation between phytoalexin production and M. truncatula defense responses against ASR. PMID:26267598

  13. Three Medicago MtFUL genes have distinct and overlapping expression patterns during vegetative and reproductive development and 35S:MtFULb accelerates flowering and causes a terminal flower phenotype in Arabidopsis.

    PubMed

    Jaudal, Mauren; Zhang, Lulu; Che, Chong; Putterill, Joanna

    2015-01-01

    The timing of the transition to flowering is carefully controlled by plants in order to optimize sexual reproduction and the ensuing production of seeds, grains, and fruits. The genetic networks that regulate floral induction are best characterized in the temperate eudicot Arabidopsis in which the florigen gene FT plays a major role in promoting the transition to flowering. Legumes are an important plant group, but less is known about the regulation of their flowering time. In the model legume Medicago truncatula (Medicago), a temperate annual plant like Arabidopsis, flowering is induced by prolonged cold (vernalization) followed by long day lengths (LD). Recent molecular-genetic experiments have revealed that a FT-like gene, MtFTa1, is a central regulator of flowering time in Medicago. Here, we characterize the three Medicago FRUITFULL (FUL) MADS transcription factors, MtFULa, MtFULb, and MtFULc using phylogenetic analyses, gene expression profiling through developmental time courses, and functional analyses in transgenic plants. MtFULa and MtFULb have similarity in sequence and expression profiles under inductive environmental conditions during both vegetative and reproductive development while MtFULc is only up regulated in the apex after flowering in LD conditions. Sustained up regulation of MtFULs requires functional MtFTa1 but their transcript levels are not affected during cold treatment. Overexpression of MtFULa and MtFULb promotes flowering in transgenic Arabidopsis plants with an additional terminal flower phenotype on some 35S:MtFULb plants. An increase in transcript levels of the MtFULs was also observed in Medicago plants overexpressing MtFTa1. Our results suggest that the MtFULs are targets of MtFTa1. Overall, this work highlights the conserved functions of FUL-like genes in promoting flowering and other roles in plant development and thus contributes to our understanding of the genetic control of the flowering process in Medicago.

  14. Aphanomyces euteiches Cell Wall Fractions Containing Novel Glucan-Chitosaccharides Induce Defense Genes and Nuclear Calcium Oscillations in the Plant Host Medicago truncatula

    PubMed Central

    Nars, Amaury; Lafitte, Claude; Chabaud, Mireille; Drouillard, Sophie; Mélida, Hugo; Danoun, Saïda; Le Costaouëc, Tinaig; Rey, Thomas; Benedetti, Julie; Bulone, Vincent; Barker, David George; Bono, Jean-Jacques; Dumas, Bernard; Jacquet, Christophe; Heux, Laurent; Fliegmann, Judith; Bottin, Arnaud

    2013-01-01

    N-acetylglucosamine-based saccharides (chitosaccharides) are components of microbial cell walls and act as molecular signals during host-microbe interactions. In the legume plant Medicago truncatula, the perception of lipochitooligosaccharide signals produced by symbiotic rhizobia and arbuscular mycorrhizal fungi involves the Nod Factor Perception (NFP) lysin motif receptor-like protein and leads to the activation of the so-called common symbiotic pathway. In rice and Arabidopsis, lysin motif receptors are involved in the perception of chitooligosaccharides released by pathogenic fungi, resulting in the activation of plant immunity. Here we report the structural characterization of atypical chitosaccharides from the oomycete pathogen Aphanomyces euteiches, and their biological activity on the host Medicago truncatula. Using a combination of biochemical and biophysical approaches, we show that these chitosaccharides are linked to β-1,6-glucans, and contain a β-(1,3;1,4)-glucan backbone whose β-1,3-linked glucose units are substituted on their C-6 carbon by either glucose or N-acetylglucosamine residues. This is the first description of this type of structural motif in eukaryotic cell walls. Glucan-chitosaccharide fractions of A. euteiches induced the expression of defense marker genes in Medicago truncatula seedlings independently from the presence of a functional Nod Factor Perception protein. Furthermore, one of the glucan-chitosaccharide fractions elicited calcium oscillations in the nucleus of root cells. In contrast to the asymmetric oscillatory calcium spiking induced by symbiotic lipochitooligosaccharides, this response depends neither on the Nod Factor Perception protein nor on the common symbiotic pathway. These findings open new perspectives in oomycete cell wall biology and elicitor recognition and signaling in legumes. PMID:24086432

  15. Trophic structure of amoeba communities near roots of Medicago sativa after contamination with fuel oil no. 6.

    PubMed

    Cortés-Pérez, Sandra; Rodríguez-Zaragoza, Salvador; Mendoza-López, Ma Remedios

    2014-02-01

    Root exudation increases microbial activity, selecting bacterial and fungal communities that metabolize organic matter such as hydrocarbons. However, a strong contamination pulse of hydrocarbons around plant roots may reorganize the soil's microbial trophic structure toward amoebae feeding on bacteria. We conducted a microcosm experiment to elucidate the effect of Medicago sativa on the trophic structure of naked amoebae after a strong pulse of pollution (50,000 ppm of fuel oil no. 6, which is a mixture of long chains ranging from C10 to C28). Plants were seeded 24 h after contamination and species of amoebae in the microcosms were identified at 1, 30, and 60 days after pollution. Several species from three trophic groups of naked amoeba were still alive 24 h after the hydrocarbon pulse. Non-planted microcosms harbored three trophic groups after 60 days, while planted ones nourished four groups. The bacterivore group was the most diverse in all microcosms, followed by protist-eaters and omnivores. The quantity of amoebae was significantly higher (3.4×10(3) organisms/g soil) in the planted pots than in the non-planted ones (1.3×10(3) organisms/g soil after 30 days of pollution (P ≤ 0.01). The shortest hydrocarbon chains (C10-C14) disappeared or diminished in all microcosms, and the longest ones increased in the planted ones. M. sativa thus exerted a positive effect on species richness, quantity, and the composition of amoebae trophic groups in contaminated soil. This indirect effect on bacterial predators is another key factor underlying hydrocarbon assimilation by living organisms during phytoremediation.

  16. Using a model-based framework for analysing genetic diversity during germination and heterotrophic growth of Medicago truncatula

    PubMed Central

    Brunel, S.; Teulat-Merah, B.; Wagner, M.-H.; Huguet, T.; Prosperi, J. M.; Dürr, C.

    2009-01-01

    Background and Aims The framework provided by an emergence model was used: (1) for phenotyping germination and heterotrophic growth of Medicago truncatula in relation to two major environmental factors, temperature and water potential; and (2) to evaluate the extent of genetic differences in emergence-model parameters. Methods Eight cultivars and natural accessions of M. trunculata were studied. Germination was recorded from 5 to 30 °C and from 0 to −0·75 MPa, and seedling growth from 10 to 20 °C. Key Results Thermal time to reach 50 % germination was very short (15 °Cd) and almost stable between genotypes, while base temperature (2–3 °C) and base water potential for germination (−0·7 to −1·3 MPa) varied between genotypes. Only 35 °Cd after germination were required to reach 30 mm hypocotyl length with significant differences among genotypes. Base temperature for elongation varied from 5·5 to 7·5 °C. Low temperatures induced a general shortening of the seedling, with some genotypes more responsive than others. No relationship with initial seed mass or seed reserve distribution was observed, which might have explained differences between genotypes and the effects of low temperatures. Conclusions The study provides a set of reference values for M. trunculata users. The use of the ecophysiological model allows comparison of these values between such non-crop species and other crops. It has enabled phenotypic variability in response to environmental conditions related to the emergence process to be identified. The model will allow simulation of emergence differences between genotypes in a range of environments using these parameter values. Genomic tools available for the model species M. trunculata will make it possible to analyse the genetic and molecular determinants of these differences. PMID:19251713

  17. Structural Investigations of N-carbamoylputrescine Amidohydrolase from Medicago truncatula: Insights into the Ultimate Step of Putrescine Biosynthesis in Plants

    PubMed Central

    Sekula, Bartosz; Ruszkowski, Milosz; Malinska, Maura; Dauter, Zbigniew

    2016-01-01

    Putrescine, 1,4-diaminobutane, is an intermediate in the biosynthesis of more complexed polyamines, spermidine and spermine. Unlike other eukaryotes, plants have evolved a multistep pathway for putrescine biosynthesis that utilizes arginine. In the final reaction, N-carbamoylputrescine is hydrolyzed to putrescine by N-carbamoylputrescine amidohydrolase (CPA, EC 3.5.1.53). During the hydrolysis, consecutive nucleophilic attacks on the substrate by Cys158 and water lead to formation of putrescine and two by-products, ammonia and carbon dioxide. CPA from the model legume plant, Medicago truncatula (MtCPA), was investigated in this work. Four crystal structures were determined: the wild-type MtCPA in complex with the reaction intermediate, N-(dihydroxymethyl)putrescine as well as with cadaverine, which is a longer analog of putrescine; and also structures of MtCPA-C158S mutant unliganded and with putrescine. MtCPA assembles into octamers, which resemble an incomplete left-handed helical twist. The active site of MtCPA is funnel-like shaped, and its entrance is walled with a contribution of the neighboring protein subunits. Deep inside the catalytic cavity, Glu48, Lys121, and Cys158 form the catalytic triad. In this studies, we have highlighted the key residues, highly conserved among the plant kingdom, responsible for the activity and selectivity of MtCPA toward N-carbamoylputrescine. Moreover, since, according to previous reports, a close MtCPA relative from Arabidopsis thaliana, along with several other nitrilase-like proteins, are subjected to allosteric regulation by substrates, we have used the structural information to indicate a putative secondary binding site. Based on the docking experiment, we postulate that this site is adjacent to the entrance to the catalytic pocket. PMID:27066023

  18. Distribution of hydrogen-metabolizing bacteria in alfalfa field soil. [Medicago sativa L. ; Convolvulus arvensis L. ; Rhizobium meliloti

    SciTech Connect

    Cunningham, S.D.; Kapulnik, Y.; Phillips, D.A.

    1986-11-01

    H/sub 2/ evolved by alfalfa root nodules during the process of N/sub 2/ fixation may be an important factor influencing the distribution of soil bacteria. To test this hypothesis under field conditions, over 700 bacterial isolates were obtained from fallow soil or from the 3-mm layer of soil surrounding alfalfa (Medicago sativa L.) root nodules, alfalfa roots, or bindweed (Convolvulus arvensis L.) roots. Bacteria were isolated under either aerobic or microaerophilic conditions and were tested for their capacity to metabolize H/sub 2/. Isolates showing net H/sub 2/ uptake and /sup 3/H/sub 2/ incorporation activity under laboratory conditions were assigned a Hup/sup +/ phenotype, whereas organisms with significant H/sub 2/ output capacity were designated as a Hout/sup +/ phenotype. Under aerobic isolation conditions two Hup/sup +/ isolates were obtained, whereas under microaerophilic conditions five Hup/sup +/ and two Hout/sup +/ isolates were found. The nine isolates differed on the basis of 24 standard bacteriological characteristics or fatty acid composition. Five of the nine organisms were isolated from soil around root nodules, whereas the other four were found distributed among the other three soil environments. On the basis of the microaerophilic isolations, 4.8% of the total procaryotic isolates from soil around root nodules were capable of oxidizing H/sub 2/, and 1.2% could produce H/sub 2/. Two of the Hup/sup +/ isolates were identified as Rhizobium meliloti by root nodulation tests, but the fact that none of the isolates reduced C/sub 2/H/sub 2/ under the assay conditions suggested that the H/sub 2/ metabolism traits were associated with various hydrogenase systems rather than with nitrogenase activity.

  19. Possible role of glutamine synthetase of the prokaryotic type (GSI-like) in nitrogen signaling in Medicago truncatula.

    PubMed

    Silva, Liliana S; Seabra, Ana R; Leitão, José N; Carvalho, Helena G

    2015-11-01

    Genes containing domains related to glutamine synthetase of the prokaryotic type (GSI-like) are widespread in higher plants, but their function is currently unknown. To gain insights into the possible role of GSI-like proteins, we characterized the GSI-like gene family of Medicago truncatula and investigated the functionality of the encoded proteins. M. truncatula contains two-expressed GSI-like genes, MtGSIa and MtGSIb, encoding polypeptides of 454 and 453 amino acids, respectively. Heterologous complementation assays of a bacterial glnA mutant indicate that the proteins are not catalytically functional for glutamine synthesis. Gene expression was investigated by qRT-PCR and western blot analysis in different organs of the plant and under different nitrogen (N) regimes, revealing that both genes are preferentially expressed in roots and root nodules, and that their expression is influenced by the N-status of the plant. Analysis of transgenic plants expressing MtGSI-like-promoter-gusA fusion, indicate that the two genes are strongly expressed in the root pericycle, and interestingly, the expression is enhanced at the sites of nodule emergence being particularly strong in specific cells located in front of the protoxylem poles. Taken together, the results presented here support a role of GSI-like proteins in N sensing and/or signaling, probably operating at the interface between perception of the N-status and the developmental processes underlying both root nodule and lateral root formation. This study indicates that GSI-like genes may represent a novel class of molecular players of the N-mediated signaling events. PMID:26475191

  20. The bHLH Transcription Factors TSAR1 and TSAR2 Regulate Triterpene Saponin Biosynthesis in Medicago truncatula.

    PubMed

    Mertens, Jan; Pollier, Jacob; Vanden Bossche, Robin; Lopez-Vidriero, Irene; Franco-Zorrilla, José Manuel; Goossens, Alain

    2016-01-01

    Plants respond to stresses by producing a broad spectrum of bioactive specialized metabolites. Hormonal elicitors, such as jasmonates, trigger a complex signaling circuit leading to the concerted activation of specific metabolic pathways. However, for many specialized metabolic pathways, the transcription factors involved remain unknown. Here, we report on two homologous jasmonate-inducible transcription factors of the basic helix-loop-helix family, TRITERPENE SAPONIN BIOSYNTHESIS ACTIVATING REGULATOR1 (TSAR1) and TSAR2, which direct triterpene saponin biosynthesis in Medicago truncatula. TSAR1 and TSAR2 are coregulated with and transactivate the genes encoding 3-HYDROXY-3-METHYLGLUTARYL-COENZYME A REDUCTASE1 (HMGR1) and MAKIBISHI1, the rate-limiting enzyme for triterpene biosynthesis and an E3 ubiquitin ligase that controls HMGR1 levels, respectively. Transactivation is mediated by direct binding of TSARs to the N-box in the promoter of HMGR1. In transient expression assays in tobacco (Nicotiana tabacum) protoplasts, TSAR1 and TSAR2 exhibit different patterns of transactivation of downstream triterpene saponin biosynthetic genes, hinting at distinct functionalities within the regulation of the pathway. Correspondingly, overexpression of TSAR1 or TSAR2 in M. truncatula hairy roots resulted in elevated transcript levels of known triterpene saponin biosynthetic genes and strongly increased the accumulation of triterpene saponins. TSAR2 overexpression specifically boosted hemolytic saponin biosynthesis, whereas TSAR1 overexpression primarily stimulated nonhemolytic soyasaponin biosynthesis. Both TSARs also activated all genes of the precursor mevalonate pathway but did not affect sterol biosynthetic genes, pointing to their specific role as regulators of specialized triterpene metabolism in M. truncatula.

  1. [Development of Arbuscular Mycorrhiza in Highly Responsive and Mycotrophic Host Plant-Black Medick (Medicago lupulina L.)].

    PubMed

    Yurkova, A P; Jacobi, L M; Gapeeva, N E; Stepanova, G V; Shishova, M F

    2015-01-01

    The main phases of arbuscular mycorrhiza (AM) development were analyzed in black medick (Medicago lupulina) with Glomus intraradices. Methods of light and transmission electron microscopy were used to investigate AM. The first mycorrhization was identified on the seventh day after sowing. M. lupulina with AM-fungus Glomus intraradices formed Arum type of AM. Roots of black medick at fruiting stage (on the 88th day) were characterized by the development of forceful mycelium. The thickness of mycelium was comparable with the vascular system of root central cylinder. The development of vesicules into intraradical spores was shown. Micelium, arbuscules, and vesicules developed in close vicinity to the division zone of root tip. This might be evidence of an active symbiotic interaction between partners. All stages of fungal development and breeding, including intraradical spores (in inter-cellular matrix of root cortex), were identified in the roots of black medick, which indicated an active utilization of host plant nutrient substrates by the mycosymbiont. Plant cell cytoplasm extension was identified around young arbuscular branches but not for intracellular hyphae. The presence of active symbiosis was confirmed by increased accumulation of phosphorus in M. lupulina root tissues under conditions of G. intraradices inoculation and low phosphorus level in the soil. Thus, black medick cultivar-population can be characterized as an ecologically obligate mycotrophic plant under conditions of low level of available phosphorus in the soil. Specific features of AM development in intensively mycotrophic black medick, starting from the stage of the first true leaf until host plant fruiting, were evaluated. The obtained plant-microbe system is a perspective model object for further ultracytological and molecular genetic studies of the mechanisms controlling arbuscular mycorrhiza symbiotic efficiency, including selection and investigation of new symbiotic plant mutants.

  2. A tandem Kunitz protease inhibitor (KPI106)-serine carboxypeptidase (SCP1) controls mycorrhiza establishment and arbuscule development in Medicago truncatula.

    PubMed

    Rech, Stefanie S; Heidt, Sven; Requena, Natalia

    2013-09-01

    Plant proteases and protease inhibitors are involved in plant developmental processes including those involving interactions with microbes. Here we show that a tandem between a Kunitz protease inhibitor (KPI106) and a serine carboxypeptidase (SCP1) controls arbuscular mycorrhiza development in the root cortex of Medicago truncatula. Both proteins are only induced during mycorrhiza formation and belong to large families whose members are also mycorrhiza-specific. Furthermore, the interaction between KPI106 and SCP1 analysed using the yeast two-hybrid system is specific, indicating that each family member might have a defined counterpart. In silico docking analysis predicted a putative P1 residue in KPI106 (Lys173) that fits into the catalytic pocket of SCP1, suggesting that KPI106 might inhibit the enzyme activity by mimicking the protease substrate. In vitro mutagenesis of the Lys173 showed that this residue is important in determining the strength and specificity of the interaction. The RNA interference (RNAi) inactivation of the serine carboxypeptidase SCP1 produces aberrant mycorrhizal development with an increased number of septated hyphae and degenerate arbuscules, a phenotype also observed when overexpressing KPI106. Protease and inhibitor are both secreted as observed when expressed in Nicotiana benthamiana epidermal cells. Taken together we envisage a model in which the protease SCP1 is secreted in the apoplast where it produces a peptide signal critical for proper fungal development within the root. KPI106 also at the apoplast would modulate the spatial and/or temporal activity of SCP1 by competing with the protease substrate.

  3. Drought and Recovery: Independently Regulated Processes Highlighting the Importance of Protein Turnover Dynamics and Translational Regulation in Medicago truncatula.

    PubMed

    Lyon, David; Castillejo, Maria Angeles; Mehmeti-Tershani, Vlora; Staudinger, Christiana; Kleemaier, Christoph; Wienkoop, Stefanie

    2016-06-01

    Climate change in conjunction with population growth necessitates a systems biology approach to characterize plant drought acclimation as well as a more thorough understanding of the molecular mechanisms of stress recovery. Plants are exposed to a continuously changing environment. Extremes such as several weeks of drought are followed by rain. This requires a molecular plasticity of the plant enabling drought acclimation and the necessity of deacclimation processes for recovery and continuous growth.During drought stress and subsequent recovery, the metabolome and proteome are regulated through a sequence of molecular processes including synthesis and degradation and molecular interaction networks are part of this regulatory process. In order to study this complex regulatory network, a comprehensive analysis is presented for the first time, investigating protein turnover and regulatory classes of proteins and metabolites during a stress recovery scenario in the model legume Medicago truncatula The data give novel insights into the molecular capacity and differential processes required for acclimation and deacclimation of severe drought stressed plants.Functional cluster and network analyses unraveled independent regulatory mechanisms for stress and recovery with different dynamic phases that during the course of recovery define the plants deacclimation from stress. The combination of relative abundance levels and turnover analysis revealed an early transition phase that seems key for recovery initiation through water resupply and is independent from renutrition. Thus, a first indication for a metabolite and protein-based load capacity was observed necessary for the recovery from drought, an important but thus far ignored possible feature toward tolerance. The data indicate that apart from the plants molecular stress response mechanisms, plasticity may be related to the nutritional status of the plant prior to stress initiation. A new perspective and possible new

  4. Possible role of glutamine synthetase of the prokaryotic type (GSI-like) in nitrogen signaling in Medicago truncatula.

    PubMed

    Silva, Liliana S; Seabra, Ana R; Leitão, José N; Carvalho, Helena G

    2015-11-01

    Genes containing domains related to glutamine synthetase of the prokaryotic type (GSI-like) are widespread in higher plants, but their function is currently unknown. To gain insights into the possible role of GSI-like proteins, we characterized the GSI-like gene family of Medicago truncatula and investigated the functionality of the encoded proteins. M. truncatula contains two-expressed GSI-like genes, MtGSIa and MtGSIb, encoding polypeptides of 454 and 453 amino acids, respectively. Heterologous complementation assays of a bacterial glnA mutant indicate that the proteins are not catalytically functional for glutamine synthesis. Gene expression was investigated by qRT-PCR and western blot analysis in different organs of the plant and under different nitrogen (N) regimes, revealing that both genes are preferentially expressed in roots and root nodules, and that their expression is influenced by the N-status of the plant. Analysis of transgenic plants expressing MtGSI-like-promoter-gusA fusion, indicate that the two genes are strongly expressed in the root pericycle, and interestingly, the expression is enhanced at the sites of nodule emergence being particularly strong in specific cells located in front of the protoxylem poles. Taken together, the results presented here support a role of GSI-like proteins in N sensing and/or signaling, probably operating at the interface between perception of the N-status and the developmental processes underlying both root nodule and lateral root formation. This study indicates that GSI-like genes may represent a novel class of molecular players of the N-mediated signaling events.

  5. Genome-Wide Analysis of the AP2/ERF Superfamily Genes and their Responses to Abiotic Stress in Medicago truncatula

    PubMed Central

    Shu, Yongjun; Liu, Ying; Zhang, Jun; Song, Lili; Guo, Changhong

    2016-01-01

    The AP2/ERF superfamily is a large, plant-specific transcription factor family that is involved in many important processes, including plant growth, development, and stress responses. Using Medicago truncatula genome information, we identified and characterized 123 putative AP2/ERF genes, which were named as MtERF1–123. These genes were classified into four families based on phylogenetic analysis, which is consistent with the results of other plant species. MtERF genes are distributed throughout all chromosomes but are clustered on various chromosomes due to genomic tandem and segmental duplication. Using transcriptome, high-throughput sequencing data, and qRT-PCR analysis, we assessed the expression patterns of the MtERF genes in tissues during development and under abiotic stresses. In total, 87 MtERF genes were expressed in plant tissues, most of which were expressed in specific tissues during development or under specific abiotic stress treatments. These results support the notion that MtERF genes are involved in developmental regulation and environmental responses in M. truncatula. Furthermore, a cluster of DREB subfamily members on chromosome 6 was induced by both cold and freezing stress, representing a positive gene regulatory response under low temperature stress, which suggests that these genes might contribute to freezing tolerance to M. truncatula. In summary, our genome-wide characterization, evolutionary analysis, and expression pattern analysis of MtERF genes in M. truncatula provides valuable information for characterizing the molecular functions of these genes and utilizing them to improve stress tolerance in plants. PMID:26834762

  6. Morphological and genetic changes induced by excess Zn in roots of Medicago truncatula A17 and a Zn accumulating mutant

    PubMed Central

    2012-01-01

    Background Nutrient fluxes associated with legume-rhizobia symbioses are poorly understood and little is known regarding the influence of abiotic stresses on development and maintenance of N-fixing nodules and root system architecture (RSA). We examined effects of Zn on nodule development and structure, root architecture, and expression of nodulation-related miRNAs in Medicago truncatula and the mutant, raz (requires additional Zn). Findings Excess Zn increased root and shoot associated Zn in both genotypes, however, raz plants had lower root associated Zn than WT plants. Roots of raz plants exposed to excess Zn had less volume, surface area, and total length compared to WT plants. Raz plants had lower lateral root number than WT plants. Excess Zn was found to increase root diameter in both genotypes. The Mn Translocation Factor (TfMn) increased in response to Zn in both genotypes; this was more pronounced in raz plants. TfZn was higher in raz plants and reduced in both genotypes in response to Zn. Nodulation was not influenced by Zn treatment or plant genotype. MicroRNA166 was upregulated under excess Zn in WT plants. Conclusions Neither the raz mutation nor Zn treatment affected nodulation, however, raz plants had altered RSA compared with WT and responded differently to Zn, implying the mutation potentially modulates RSA responses to Zn but doesn’t play a direct role in nodulation. MicroRNA166 was significantly induced in WT plants by excess Zn, warranting further investigation into the potential role it plays in controlling RSA. PMID:23191938

  7. Improvement of phosphate solubilization and Medicago plant yield by an indole-3-acetic acid-overproducing strain of Sinorhizobium meliloti.

    PubMed

    Bianco, Carmen; Defez, Roberto

    2010-07-01

    Nitrogen (N) and phosphorus (P) are the most limiting factors for plant growth. Some microorganisms improve the uptake and availability of N and P, minimizing chemical fertilizer dependence. It has been published that the RD64 strain, a Sinorhizobium meliloti 1021 strain engineered to overproduce indole-3-acetic acid (IAA), showed improved nitrogen fixation ability compared to the wild-type 1021 strain. Here, we present data showing that RD64 is also highly effective in mobilizing P from insoluble sources, such as phosphate rock (PR). Under P-limiting conditions, the higher level of P-mobilizing activity of RD64 than of the 1021 wild-type strain is connected with the upregulation of genes coding for the high-affinity P transport system, the induction of acid phosphatase activity, and the increased secretion into the growth medium of malic, succinic, and fumaric acids. Medicago truncatula plants nodulated by RD64 (Mt-RD64), when grown under P-deficient conditions, released larger amounts of another P-solubilizing organic acid, 2-hydroxyglutaric acid, than plants nodulated by the wild-type strain (Mt-1021). It has already been shown that Mt-RD64 plants exhibited higher levels of dry-weight production than Mt-1021 plants. Here, we also report that P-starved Mt-RD64 plants show significant increases in both shoot and root fresh weights when compared to P-starved Mt-1021 plants. We discuss how, in a Rhizobium-legume model system, a balanced interplay of different factors linked to bacterial IAA overproduction rather than IAA production per se stimulates plant growth under stressful environmental conditions and, in particular, under P starvation. PMID:20511434

  8. Medicago truncatula Mutants Demonstrate the Role of Plant Calcium Oxalate Crystals as an Effective Defense against Chewing Insects1

    PubMed Central

    Korth, Kenneth L.; Doege, Sarah J.; Park, Sang-Hyuck; Goggin, Fiona L.; Wang, Qin; Gomez, S. Karen; Liu, Guangjie; Jia, Lingling; Nakata, Paul A.

    2006-01-01

    Calcium oxalate is the most abundant insoluble mineral found in plants and its crystals have been reported in more than 200 plant families. In the barrel medic Medicago truncatula Gaertn., these crystals accumulate predominantly in a sheath surrounding secondary veins of leaves. Mutants of M. truncatula with decreased levels of calcium oxalate crystals were used to assess the defensive role of this mineral against insects. Caterpillar larvae of the beet armyworm Spodoptera exigua Hübner show a clear feeding preference for tissue from calcium oxalate-defective (cod) mutant lines cod5 and cod6 in choice test comparisons with wild-type M. truncatula. Compared to their performance on mutant lines, larvae feeding on wild-type plants with abundant calcium oxalate crystals suffer significantly reduced growth and increased mortality. Induction of wound-responsive genes appears to be normal in cod5 and cod6, indicating that these lines are not deficient in induced insect defenses. Electron micrographs of insect mouthparts indicate that the prismatic crystals in M. truncatula leaves act as physical abrasives during feeding. Food utilization measurements show that, after consumption, calcium oxalate also interferes with the conversion of plant material into insect biomass during digestion. In contrast to their detrimental effects on a chewing insect, calcium oxalate crystals do not negatively affect the performance of the pea aphid Acyrthosiphon pisum Harris, a sap-feeding insect with piercing-sucking mouthparts. The results confirm a long-held hypothesis for the defensive function of these crystals and point to the potential value of genes controlling crystal formation and localization in crop plants. PMID:16514014

  9. A switch in Ca2+ spiking signature is concomitant with endosymbiotic microbe entry into cortical root cells of Medicago truncatula.

    PubMed

    Sieberer, Björn J; Chabaud, Mireille; Fournier, Joëlle; Timmers, Antonius C J; Barker, David G

    2012-03-01

    Ca(2+) spiking is a central component of a common signaling pathway that is activated in the host epidermis during initial recognition of endosymbiotic microbes. However, it is not known to what extent Ca(2+) signaling also plays a role during subsequent root colonization involving apoplastic transcellular infection. Live-tissue imaging using calcium cameleon reporters expressed in Medicago truncatula roots has revealed that distinct Ca(2+) oscillatory profiles correlate with specific stages of transcellular cortical infection by both rhizobia and arbuscular mycorrhizal fungi. Outer cortical cells exhibit low-frequency Ca(2+) spiking during the extensive intracellular remodeling that precedes infection. This appears to be a prerequisite for the formation of either pre-infection threads or the pre-penetration apparatus, both of which are fully reversible processes. A transition from low- to high-frequency spiking is concomitant with the initial stages of apoplastic cell entry by both microbes. This high-frequency spiking is of limited duration in the case of rhizobial infection and is completely switched off by the time transcellular infection by both microsymbionts is completed. The Ca(2+) spiking profiles associated with both rhizobial and arbuscular mycorrhizal cell entry are remarkably similar in terms of periodicity, suggesting that microbe specificity is unlikely to be encoded by the Ca(2+) signature during this particular stage of host infection in the outer cortex. Together, these findings lead to the proposal that tightly regulated Ca(2+) -mediated signal transduction is a key player in reprogramming root cell development at the critical stage of commitment to endosymbiotic infection.

  10. Local and distal effects of arbuscular mycorrhizal colonization on direct pathway Pi uptake and root growth in Medicago truncatula.

    PubMed

    Watts-Williams, Stephanie J; Jakobsen, Iver; Cavagnaro, Timothy R; Grønlund, Mette

    2015-07-01

    Two pathways exist for plant Pi uptake from soil: via root epidermal cells (direct pathway) or via associations with arbuscular mycorrhizal (AM) fungi, and the two pathways interact in a complex manner. This study investigated distal and local effects of AM colonization on direct root Pi uptake and root growth, at different soil P levels. Medicago truncatula was grown at three soil P levels in split-pots with or without AM fungal inoculation and where one root half grew into soil labelled with (33)P. Plant genotypes included the A17 wild type and the mtpt4 mutant. The mtpt4 mutant, colonized by AM fungi, but with no functional mycorrhizal pathway for Pi uptake, was included to better understand effects of AM colonization per se. Colonization by AM fungi decreased expression of direct Pi transporter genes locally, but not distally in the wild type. In mtpt4 mutant plants, direct Pi transporter genes and the Pi starvation-induced gene Mt4 were more highly expressed than in wild-type roots. In wild-type plants, less Pi was taken up via the direct pathway by non-colonized roots when the other root half was colonized by AM fungi, compared with non-mycorrhizal plants. Colonization by AM fungi strongly influenced root growth locally and distally, and direct root Pi uptake activity locally, but had only a weak influence on distal direct pathway activity. The responses to AM colonization in the mtpt4 mutant suggested that in the wild type, the increased P concentration of colonized roots was a major factor driving the effects of AM colonization on direct root Pi uptake. PMID:25944927

  11. Local and distal effects of arbuscular mycorrhizal colonization on direct pathway Pi uptake and root growth in Medicago truncatula.

    PubMed

    Watts-Williams, Stephanie J; Jakobsen, Iver; Cavagnaro, Timothy R; Grønlund, Mette

    2015-07-01

    Two pathways exist for plant Pi uptake from soil: via root epidermal cells (direct pathway) or via associations with arbuscular mycorrhizal (AM) fungi, and the two pathways interact in a complex manner. This study investigated distal and local effects of AM colonization on direct root Pi uptake and root growth, at different soil P levels. Medicago truncatula was grown at three soil P levels in split-pots with or without AM fungal inoculation and where one root half grew into soil labelled with (33)P. Plant genotypes included the A17 wild type and the mtpt4 mutant. The mtpt4 mutant, colonized by AM fungi, but with no functional mycorrhizal pathway for Pi uptake, was included to better understand effects of AM colonization per se. Colonization by AM fungi decreased expression of direct Pi transporter genes locally, but not distally in the wild type. In mtpt4 mutant plants, direct Pi transporter genes and the Pi starvation-induced gene Mt4 were more highly expressed than in wild-type roots. In wild-type plants, less Pi was taken up via the direct pathway by non-colonized roots when the other root half was colonized by AM fungi, compared with non-mycorrhizal plants. Colonization by AM fungi strongly influenced root growth locally and distally, and direct root Pi uptake activity locally, but had only a weak influence on distal direct pathway activity. The responses to AM colonization in the mtpt4 mutant suggested that in the wild type, the increased P concentration of colonized roots was a major factor driving the effects of AM colonization on direct root Pi uptake.

  12. Arbuscular Mycorrhizal Colonization Alters Subcellular Distribution and Chemical Forms of Cadmium in Medicago sativa L. and Resists Cadmium Toxicity

    PubMed Central

    Gao, Yanzheng

    2012-01-01

    Some plants can tolerate and even detoxify soils contaminated with heavy metals. This detoxification ability may depend on what chemical forms of metals are taken up by plants and how the plants distribute the toxins in their tissues. This, in turn, may have an important impact on phytoremediation. We investigated the impact of arbuscular mycorrhizal (AM) fungus, Glomus intraradices, on the subcellular distribution and chemical forms of cadmium (Cd) in alfalfa (Medicago sativa L.) that were grown in Cd-added soils. The fungus significantly colonized alfalfa roots by day 25 after planting. Colonization of alfalfa by G. intraradices in soils contaminated with Cd ranged from 17% to 69% after 25–60 days and then decreased to 43%. The biomass of plant shoots with AM fungi showed significant 1.7-fold increases compared to no AM fungi addition under the treatment of 20 mg·kg−1 Cd. Concentrations of Cd in the shoots of alfalfa under 0.5, 5, and 20 mg·kg−1 Cd without AM fungal inoculation are 1.87, 2.92, and 2.38 times higher, respectively, than those of fungi-inoculated plants. Fungal inoculation increased Cd (37.2–80.5%) in the cell walls of roots and shoots and decreased in membranes after 80 days of incubation compared to untreated plants. The proportion of the inactive forms of Cd in roots was higher in fungi-treated plants than in controls. Furthermore, although fungi-treated plants had less overall Cd in subcellular fragments in shoots, they had more inactive Cd in shoots than did control plants. These results provide a basis for further research on plant-microbe symbioses in soils contaminated with heavy metals, which may potentially help us develop management regimes for phytoremediation. PMID:23139811

  13. Selective recruitment of mRNAs and miRNAs to polyribosomes in response to rhizobia infection in Medicago truncatula.

    PubMed

    Reynoso, Mauricio Alberto; Blanco, Flavio Antonio; Bailey-Serres, Julia; Crespi, Martín; Zanetti, María Eugenia

    2013-01-01

    Translation of mRNAs is a key regulatory step that contributes to the coordination and modulation of eukaryotic gene expression during development or adaptation to the environment. mRNA stability or translatability can be regulated by the action of small regulatory RNAs (sRNAs), which control diverse biological processes. Under low nitrogen conditions, leguminous plants associate with soil bacteria and develop a new organ specialized in nitrogen fixation: the nodule. To gain insight into the translational regulation of mRNAs during nodule formation, the association of mRNAs and sRNAs to polysomes was characterized in roots of the model legume Medicago truncatula during the symbiotic interaction with Sinorhizobium meliloti. Quantitative comparison of steady-state and polysomal mRNAs for 15 genes involved in nodulation identified a group of transcripts with slight or no change in total cellular abundance that were significantly upregulated at the level of association with polysomes in response to rhizobia. This group included mRNAs encoding receptors like kinases required either for nodule organogenesis, bacterial infection or both, and transcripts encoding GRAS and NF-Y transcription factors (TFs). Quantitative analysis of sRNAs in total and polysomal RNA samples revealed that mature microRNAs (miRNAs) were associated with the translational machinery, notably, miR169 and miR172, which target the NF-YA/HAP2 and AP2 TFs, respectively. Upon inoculation, levels of miR169 pronouncedly decreased in polysomal complexes, concomitant with the increased accumulation of the NF-YA/HAP2 protein. These results indicate that both mRNAs and miRNAs are subject to differential recruitment to polysomes, and expose the importance of selective mRNA translation during root nodule symbiosis.

  14. Expression of coordinately regulated defence response genes and analysis of their role in disease resistance in Medicago truncatula.

    PubMed

    Samac, Deborah A; Peñuela, Silvia; Schnurr, Judy A; Hunt, E Nicole; Foster-Hartnett, Dawn; Vandenbosch, Kathryn A; Gantt, J Stephen

    2011-10-01

    Microarray technology was used to identify the genes associated with disease defence responses in the model legume Medicago truncatula. Transcript profiles from M. truncatula cv. Jemalong genotype A17 leaves inoculated with Colletotrichum trifolii and Erysiphe pisi and roots infected with Phytophthora medicaginis were compared to identify the genes expressed in response to all three pathogens and genes unique to an interaction. The A17 genotype is resistant to C. trifolii and E. pisi, exhibiting a hypersensitive response after inoculation, and is moderately susceptible to P. medicaginis. Among the most strongly up-regulated genes in all three interactions were those encoding a hevein-like protein, thaumatin-like protein (TLP) and members of the pathogenesis response (PR)10 family. Transcripts of genes for enzymes in the phenylpropanoid pathway leading to the production of isoflavonoid phytoalexins increased dramatically in response to inoculation with the foliar pathogens. In P. medicaginis-inoculated roots, transcripts of genes in the phenylpropanoid pathway peaked at 5 days post-inoculation, when symptoms became visible. Transcript accumulation of three PR10 family members, a TLP and chalcone synthase (CHS) was assessed in M. truncatula genotype R108 plants. The R108 plants are resistant to C. trifolii and moderately susceptible to E. pisi and P. medicaginis. Transcript accumulation paralleled the stages of pathogen development. To evaluate the role of a TLP, a PR10 family member and CHS in disease resistance, transgenic R108 plants containing interfering RNA (RNAi) constructs were produced. Reduced expression of PR10 and TLP had no effect on the disease phenotype, whereas reduced expression of CHS resulted in increased susceptibility to necrotrophic pathogens.

  15. Pleiotropic effect of fluoranthene on anthocyanin synthesis and nodulation of Medicago sativa is reversed by the plant flavone luteolin

    SciTech Connect

    Wetzel, A.; Parniske, M.; Werner, D.

    1995-05-01

    The symbiosis between leguminous plants and soil bacteria of the genus Rhizobium is of considerable agronominal importance. Recently it has been found, that polycyclic aromatic hydrocarbons (PAHs; e.g. anthracene, phenanthrene, fluoranthene), occurring as ubiquitous environmental contaminants can inhibit nodulation of Medicago sativa. Fluoranthene is one of the dominant PAHs found in urban particulate matter, sewage sludge or beside motorways. Several organisms have been shown to be able to metabolize and mineralize fluoranthene but the uptake of fluoranthene is limited due to low solubility of fluoranthene in water and strong adsorption to humic substances in soil. Rhizobium meliloti cannot degrade fluoranthene. Toxic effects of fluoranthene on bacterial growth have never been observed. In contrast to their rhizobial symbiotic partners, alfalfa plants grown on a solidified fluoranthene-containing medium, exhibited symptoms of toxicity. They showed a dose-responsive decrease in shoot length and, if inoculated with R. meliloti, inhibition of nodule formation. Growth retardation is accompanied by a decrease in anthocyanin pigmentation of shoots, and an atypical accumulation of anthocyanins in roots. Plant flavonoids are known to play a central role in the signal exchange of the Legume-Rhizobium symbiosis. Phenylpropane derived compounds and flavonoids have been implicated in nodule development. Since fluoranthene impairs nodulation and induces the production of anthocyanins, it is possible that these events are causally linked via phenylpropanoid metabolism. These experiments attempt to overcome the inhibitory effects of fluoranthene by exogeneous application of the flavonoid luteolin. This paper demonstrates that luteolin antagonizes the fluoranthene mediated inhibition of nodule formation and prevents the accumulation of anthocyanins in roots. 29 refs., 4 figs., 1 tab.

  16. Occurrence of Transgenic Feral Alfalfa (Medicago sativa subsp. sativa L.) in Alfalfa Seed Production Areas in the United States

    PubMed Central

    Greene, Stephanie L.; Kesoju, Sandya R.; Martin, Ruth C.; Kramer, Matthew

    2015-01-01

    The potential environmental risks of transgene exposure are not clear for alfalfa (Medicago sativa subsp. sativa), a perennial crop that is cross-pollinated by insects. We gathered data on feral alfalfa in major alfalfa seed-production areas in the western United States to (1) evaluate evidence that feral transgenic plants spread transgenes and (2) determine environmental and agricultural production factors influencing the location of feral alfalfa, especially transgenic plants. Road verges in Fresno, California; Canyon, Idaho; and Walla Walla, Washington were surveyed in 2011 and 2012 for feral plants, and samples were tested for the CP4 EPSPS protein that conveys resistance to glyphosate. Of 4580 sites surveyed, feral plants were observed at 404 sites. Twenty-seven percent of these sites had transgenic plants. The frequency of sites having transgenic feral plants varied among our study areas. Transgenic plants were found in 32.7%, 21.4.7% and 8.3% of feral plant sites in Fresno, Canyon and Walla Walla, respectively. Spatial analysis suggested that feral populations started independently and tended to cluster in seed and hay production areas, places where seed tended to drop. Significant but low spatial auto correlation suggested that in some instances, plants colonized nearby locations. Neighboring feral plants were frequently within pollinator foraging range; however, further research is needed to confirm transgene flow. Locations of feral plant clusters were not well predicted by environmental and production variables. However, the likelihood of seed spillage during production and transport had predictive value in explaining the occurrence of transgenic feral populations. Our study confirms that genetically engineered alfalfa has dispersed into the environment, and suggests that minimizing seed spillage and eradicating feral alfalfa along road sides would be effective strategies to minimize transgene dispersal. PMID:26699337

  17. Altered callose deposition during embryo sac formation of multi-pistil mutant (mp1) in Medicago sativa.

    PubMed

    Zhou, H C; Jin, L; Li, J; Wang, X J

    2016-01-01

    Whether callose deposition is the cause or result of ovule sterility in Medicago sativa remains controversial, because it is unclear when and where changes in callose deposition and dissolution occur during fertile and sterile embryo sac formation. Here, alfalfa spontaneous multi-pistil mutant (mp1) and wild-type plants were used to compare the dynamics of callose deposition during embryo sac formation using microscopy. The results showed that both mutant and wild-type plants experienced megasporogenesis and megagametogenesis, and there was no significant difference during megasporogenesis. In contrast to the wild-type plants, in which the mature embryo sac was observed after three continuous cycles of mitosis, functional megaspores of mutant plants developed abnormally after the second round of mitosis, leading to degeneration of synergid, central, and antipodal cells. Callose deposition in both mutant and wild-type plants was first observed in the walls of megasporocytes, and then in the megaspore tetrad walls. After meiosis, the callose wall began to degrade as the functional megaspore underwent mitosis, and almost no callose was observed in the mature embryo sac in wild-type plants. However, callose deposition was observed in mp1 plants around the synergid, and increased with the development of the embryo sac, and was mainly deposited at the micropylar end. Our results indicate that synergid, central, and antipodal cells, which are surrounded by callose, may degrade owing to lack of nutrition. Callose accumulation around the synergid and at the micropylar end may hinder signals required for the pollen tube to enter the embryo sac, leading to abortion. PMID:27323128

  18. Expression of a Medicago falcata small GTPase gene, MfARL1 enhanced tolerance to salt stress in Arabidopsis thaliana.

    PubMed

    Wang, Tian-Zuo; Xia, Xiu-Zhi; Zhao, Min-Gui; Tian, Qiu-Ying; Zhang, Wen-Hao

    2013-02-01

    To understand the role of small GTPases in response to abiotic stress, we isolated a gene encoding a small GTPase, designated MfARL1, from a subtracted cDNA library in Medicago falcata, a native legume species in semi-arid grassland in northern China. The function of MfARL1 in response to salt stress was studied by expressing MfARL1 in Arabidopsis. Wild-type (WT) and transgenic plants constitutively expressing MfARL1 showed comparable phenotype when grown under control conditions. Germination of seeds expressing MfARL1 was less suppressed by salt stress than that of WT seeds. Transgenic seedlings had higher survival rate than WT seedlings under salt stress, suggesting that expression of MfARL1 confers tolerance to salt stress. The physiological and molecular mechanisms underlying these phenomena were elucidated. Salt stress led to a significant decrease in chlorophyll contents in WT plants, but not in transgenic plants. Transgenic plants accumulated less amounts of H(2)O(2) and malondialdehyde than their WT counterparts under salt stress, which can be accounted for by the higher catalase activities, lower activities of superoxide dismutase, and peroxidase in transgenic plants than in WT plants. Transgenic plants displayed lower Na(+)/K(+) ratio due to less accumulation of Na(+) than wild-type under salt stress conditions. The lower Na(+)/K(+) ratio may result from less accumulation of Na(+) due to reduced expression of AtHKT1 that encodes Na(+) transporter in transgenic plants under salt stress. These findings demonstrate that MfARL1 encodes a novel stress-responsive small GTPase that is involved in tolerance to salt stress.

  19. Dual involvement of a Medicago truncatula NAC transcription factor in root abiotic stress response and symbiotic nodule senescence.

    PubMed

    de Zélicourt, Axel; Diet, Anouck; Marion, Jessica; Laffont, Carole; Ariel, Federico; Moison, Michaël; Zahaf, Ons; Crespi, Martin; Gruber, Véronique; Frugier, Florian

    2012-04-01

    Legume crops related to the model plant Medicago truncatula can adapt their root architecture to environmental conditions, both by branching and by establishing a symbiosis with rhizobial bacteria to form nitrogen-fixing nodules. Soil salinity is a major abiotic stress affecting plant yield and root growth. Previous transcriptomic analyses identified several transcription factors linked to the M. truncatula response to salt stress in roots, including NAC (NAM/ATAF/CUC)-encoding genes. Over-expression of one of these transcription factors, MtNAC969, induced formation of a shorter and less-branched root system, whereas RNAi-mediated MtNAC969 inactivation promoted lateral root formation. The altered root system of over-expressing plants was able to maintain its growth under high salinity, and roots in which MtNAC969 was down-regulated showed improved growth under salt stress. Accordingly, expression of salt stress markers was decreased or induced in MtNAC969 over-expressing or RNAi roots, respectively, suggesting a repressive function for this transcription factor in the salt-stress response. Expression of MtNAC969 in central symbiotic nodule tissues was induced by nitrate treatment, and antagonistically affected by salt in roots and nodules, similarly to senescence markers. MtNAC969 RNAi nodules accumulated amyloplasts in the nitrogen-fixing zone, and were prematurely senescent. Therefore, the MtNAC969 transcription factor, which is differentially affected by environmental cues in root and nodules, participates in several pathways controlling adaptation of the M. truncatula root system to the environment.

  20. Meat quality of wether lambs grazed on either saltbush (Atriplex nummularia) plus supplements or lucerne (Medicago sativa).

    PubMed

    Hopkins, D L; Nicholson, A

    1999-01-01

    The meat quality of 42, 8 month-old cross-bred wether lambs that had been grazed on either predominantly saltbush (Atriplex nummularia) and supplemented with pasture hay (n=14) or oat grain (n=14) for 68 days was compared to lambs grazed predominantly on lucerne (Medicago sativa; n=14). The lambs used in the study ranged in un-fasted liveweight from 44.5 to 63.6kg pre-slaughter. There was no significant (p>0.05) difference between the treatments for liveweight, but there was a significant (p<0.05) treatment effect on hot carcass weight with those from the saltbush/hay group (SH) being lighter than those from the lucerne group. When the carcass measures of fatness or m. longissimus thoracis et lumborum area were adjusted to a common carcass weight of 22.4kg there was no difference between groups. There were no significant differences (p>0.05) between groups for pH or colour values (where L* indicates relative lightness, a* indicates relative redness and b* indicates relative yellowness). There was no significant difference (p>0.05) between groups for b(2)* values of subcutaneous fat. Treatment had a significant effect on aroma strength (p<0.05), samples from lambs in the SH group (n=10) and those in saltbush/grain (SG) group (n=10) having a stronger aroma than those from lambs grazed on lucerne (L; n=10). No treatment effect for liking of aroma was found. Flavour strength was not significantly (p>0.05) stronger for samples from groups SH and SG than for samples from group L. There was no effect of treatment on tenderness or juiciness and overall panellists ranked the samples similarly for acceptability. Finishing lambs on saltbush and either supplemented with hay or grain as used in this experiment did not present any apparent meat quality problems compared to lucerne fed lambs.

  1. Drought and Recovery: Independently Regulated Processes Highlighting the Importance of Protein Turnover Dynamics and Translational Regulation in Medicago truncatula*

    PubMed Central

    Lyon, David; Castillejo, Maria Angeles; Mehmeti-Tershani, Vlora; Staudinger, Christiana; Kleemaier, Christoph; Wienkoop, Stefanie

    2016-01-01

    Climate change in conjunction with population growth necessitates a systems biology approach to characterize plant drought acclimation as well as a more thorough understanding of the molecular mechanisms of stress recovery. Plants are exposed to a continuously changing environment. Extremes such as several weeks of drought are followed by rain. This requires a molecular plasticity of the plant enabling drought acclimation and the necessity of deacclimation processes for recovery and continuous growth. During drought stress and subsequent recovery, the metabolome and proteome are regulated through a sequence of molecular processes including synthesis and degradation and molecular interaction networks are part of this regulatory process. In order to study this complex regulatory network, a comprehensive analysis is presented for the first time, investigating protein turnover and regulatory classes of proteins and metabolites during a stress recovery scenario in the model legume Medicago truncatula. The data give novel insights into the molecular capacity and differential processes required for acclimation and deacclimation of severe drought stressed plants. Functional cluster and network analyses unraveled independent regulatory mechanisms for stress and recovery with different dynamic phases that during the course of recovery define the plants deacclimation from stress. The combination of relative abundance levels and turnover analysis revealed an early transition phase that seems key for recovery initiation through water resupply and is independent from renutrition. Thus, a first indication for a metabolite and protein-based load capacity was observed necessary for the recovery from drought, an important but thus far ignored possible feature toward tolerance. The data indicate that apart from the plants molecular stress response mechanisms, plasticity may be related to the nutritional status of the plant prior to stress initiation. A new perspective and possible

  2. Symbiosis between nitrogen-fixing bacteria and Medicago truncatula is not significantly affected by silver and silver sulfide nanomaterials.

    PubMed

    Judy, Jonathan D; Kirby, Jason K; McLaughlin, Mike J; McNear, David; Bertsch, Paul M

    2016-07-01

    Silver (Ag) engineered nanomaterials (ENMs) are being released into waste streams and are being discharged, largely as Ag2S aged-ENMs (a-ENMs), into agroecosystems receiving biosolids amendments. Recent research has demonstrated that biosolids containing an environmentally relevant mixture of ZnO, TiO2, and Ag ENMs and their transformation products, including Ag2S a-ENMs, disrupted the symbiosis between nitrogen-fixing bacteria and legumes. However, this study was unable to unequivocally determine which ENM or combination of ENMs and a-ENMs was responsible for the observed inhibition. Here, we examined further the effects of polyvinylpyrollidone (PVP) coated pristine Ag ENMs (PVP-Ag), Ag2S a-ENMs, and soluble Ag (as AgSO4) at 1, 10, and 100 mg Ag kg(-1) on the symbiosis between the legume Medicago truncatula and the nitrogen-fixing bacterium, Sinorhizobium melliloti in biosolids-amended soil. Nodulation frequency, nodule function, glutathione reductase production, and biomass were not significantly affected by any of the Ag treatments, even at 100 mg kg(-1), a concentration analogous to a worst-case scenario resulting from long-term, repeated biosolids amendments. Our results provide additional evidence that the disruption of the symbiosis between nitrogen-fixing bacteria and legumes in response to a mixture of ENMs in biosolids-amended soil reported previously may not be attributable to Ag ENMs or their transformation end-products. We anticipate these findings will provide clarity to regulators and industry regarding potential unintended consequences to terrestrial ecosystems resulting from of the use of Ag ENMs in consumer products. PMID:27149150

  3. Identification and Analysis of Medicago truncatula Auxin Transporter Gene Families Uncover their Roles in Responses to Sinorhizobium meliloti Infection.

    PubMed

    Shen, Chenjia; Yue, Runqing; Bai, Youhuang; Feng, Rong; Sun, Tao; Wang, Xiaofei; Yang, Yanjun; Tie, Shuanggui; Wang, Huizhong

    2015-10-01

    Auxin transport plays a pivotal role in the interaction between legume species and nitrogen-fixing bacteria to form symbioses. Auxin influx carriers auxin resistant 1/like aux 1 (AUX/LAX), efflux carriers pin-formed (PIN) and efflux/conditional P-glycoprotein (PGP/ABCB) are three major protein families participating in auxin polar transport. We used the latest Medicago truncatula genome sequence to characterize and analyze the M. truncatula LAX (MtLAX), M. truncatula PIN (MtPIN) and M. truncatula ABCB (MtABCB) families. Transient expression experiments indicated that three representative auxin transporters (MtLAX3, MtPIN7 and MtABCB1) showed cell plasma membrane localizations. The expression of most MtLAX, MtPIN and MtABCB genes was up-regulated in the roots and was down-regulated in the shoots by Sinorhizobium meliloti infection in the wild type (WT). However, the expression of these genes was down-regulated in both the roots and shoots of an infection-resistant mutant, dmi3. The different expression patterns between the WT and the mutant roots indicated that auxin relocation may be involved in rhizobial infection responses. Furthermore, IAA contents were significantly up-regulated in the shoots and down-regulated in the roots after Sinorhizobium meliloti infection in the WT. Inoculation of roots with rhizobia may reduce the auxin loading from shoots to roots by inhibiting the expression of most auxin transporter genes. However, the rate of change of gene expression and IAA contents in the dmi3 mutant were obviously lower than in the WT. The identification and expression analysis of auxin transporter genes helps us to understand the roles of auxin in the regulation of nodule formation in M. truncatula.

  4. The structures of cytosolic and plastid-located glutamine synthetases from Medicago truncatula reveal a common and dynamic architecture

    SciTech Connect

    Torreira, Eva; Seabra, Ana Rita; Marriott, Hazel; Zhou, Min; Llorca, Óscar; Robinson, Carol V.; Carvalho, Helena G.; Fernández-Tornero, Carlos; Pereira, Pedro José Barbosa

    2014-04-01

    The experimental models of dicotyledonous cytoplasmic and plastid-located glutamine synthetases unveil a conserved eukaryotic-type decameric architecture, with subtle structural differences in M. truncatula isoenzymes that account for their distinct herbicide resistance. The first step of nitrogen assimilation in higher plants, the energy-driven incorporation of ammonia into glutamate, is catalyzed by glutamine synthetase. This central process yields the readily metabolizable glutamine, which in turn is at the basis of all subsequent biosynthesis of nitrogenous compounds. The essential role performed by glutamine synthetase makes it a prime target for herbicidal compounds, but also a suitable intervention point for the improvement of crop yields. Although the majority of crop plants are dicotyledonous, little is known about the structural organization of glutamine synthetase in these organisms and about the functional differences between the different isoforms. Here, the structural characterization of two glutamine synthetase isoforms from the model legume Medicago truncatula is reported: the crystallographic structure of cytoplasmic GSII-1a and an electron cryomicroscopy reconstruction of plastid-located GSII-2a. Together, these structural models unveil a decameric organization of dicotyledonous glutamine synthetase, with two pentameric rings weakly connected by inter-ring loops. Moreover, rearrangement of these dynamic loops changes the relative orientation of the rings, suggesting a zipper-like mechanism for their assembly into a decameric enzyme. Finally, the atomic structure of M. truncatula GSII-1a provides important insights into the structural determinants of herbicide resistance in this family of enzymes, opening new avenues for the development of herbicide-resistant plants.

  5. Local and distal effects of arbuscular mycorrhizal colonization on direct pathway Pi uptake and root growth in Medicago truncatula

    PubMed Central

    Watts-Williams, Stephanie J.; Jakobsen, Iver; Cavagnaro, Timothy R.; Grønlund, Mette

    2015-01-01

    Two pathways exist for plant Pi uptake from soil: via root epidermal cells (direct pathway) or via associations with arbuscular mycorrhizal (AM) fungi, and the two pathways interact in a complex manner. This study investigated distal and local effects of AM colonization on direct root Pi uptake and root growth, at different soil P levels. Medicago truncatula was grown at three soil P levels in split-pots with or without AM fungal inoculation and where one root half grew into soil labelled with 33P. Plant genotypes included the A17 wild type and the mtpt4 mutant. The mtpt4 mutant, colonized by AM fungi, but with no functional mycorrhizal pathway for Pi uptake, was included to better understand effects of AM colonization per se. Colonization by AM fungi decreased expression of direct Pi transporter genes locally, but not distally in the wild type. In mtpt4 mutant plants, direct Pi transporter genes and the Pi starvation-induced gene Mt4 were more highly expressed than in wild-type roots. In wild-type plants, less Pi was taken up via the direct pathway by non-colonized roots when the other root half was colonized by AM fungi, compared with non-mycorrhizal plants. Colonization by AM fungi strongly influenced root growth locally and distally, and direct root Pi uptake activity locally, but had only a weak influence on distal direct pathway activity. The responses to AM colonization in the mtpt4 mutant suggested that in the wild type, the increased P concentration of colonized roots was a major factor driving the effects of AM colonization on direct root Pi uptake. PMID:25944927

  6. Ectopic Expression of GsPPCK3 and SCMRP in Medicago sativa Enhances Plant Alkaline Stress Tolerance and Methionine Content

    PubMed Central

    Zhao, Yang; Zhao, Chaoyue; DuanMu, Huizi; Yu, Yang; Ji, Wei; Zhu, Yanming

    2014-01-01

    So far, it has been suggested that phosphoenolpyruvate carboxylases (PEPCs) and PEPC kinases (PPCKs) fulfill several important non-photosynthetic functions. However, the biological functions of soybean PPCKs, especially in alkali stress response, are not yet well known. In previous studies, we constructed a Glycine soja transcriptional profile, and identified three PPCK genes (GsPPCK1, GsPPCK2 and GsPPCK3) as potential alkali stress responsive genes. In this study, we confirmed the induced expression of GsPPCK3 under alkali stress and investigated its tissue expression specificity by using quantitative real-time PCR analysis. Then we ectopically expressed GsPPCK3 in Medicago sativa and found that GsPPCK3 overexpression improved plant alkali tolerance, as evidenced by lower levels of relative ion leakage and MDA content and higher levels of chlorophyll content and root activity. In this respect, we further co-transformed the GsPPCK3 and SCMRP genes into alfalfa, and demonstrated the increased alkali tolerance of GsPPCK3-SCMRP transgenic lines. Further investigation revealed that GsPPCK3-SCMRP co-overexpression promoted the PEPC activity, net photosynthetic rate and citric acid content of transgenic alfalfa under alkali stress. Moreover, we also observed the up-regulated expression of PEPC, CS (citrate synthase), H+-ATPase and NADP-ME genes in GsPPCK3-SCMRP transgenic alfalfa under alkali stress. As expected, we demonstrated that GsPPCK3-SCMRP transgenic lines displayed higher methionine content than wild type alfalfa. Taken together, results presented in this study supported the positive role of GsPPCK3 in plant response to alkali stress, and provided an effective way to simultaneously improve plant alkaline tolerance and methionine content, at least in legume crops. PMID:24586886

  7. [MEDICAGO SATIVA L: IMPROVEMENT AND NEW APPROACHES OF ITS NUTRITIONAL AND FUNCTIONAL VALUE BY BACTERIAL CO-INOCULATION].

    PubMed

    Martínez, Rosario; Nebot, Elena; Porres, Jesús María; Kapravelou, Garyfallia; Del Moral, Ana; Talbi, Chouhra; Bedmar, Eulogio Jose; López-Jurado, María

    2015-12-01

    Objetivo: estudiar el efecto de la inoculación con Ensifer meliloti y Halomonas maura sobre el crecimiento y el valor nutricional y funcional de la leguminosa Medicago sativa L., cultivada bajo condiciones de salinidad. Método: las plantas de M. sativa se cultivaron con una solución de mezcla de sales CaSO4, MgCl, NaCl and Na- HCO3 y se coinocularon con su rizobio específico y la bacteria H. maura. Se determinaron los parámetros fisiológicos de las plantas, así como el contenido en nitrógeno y minerales, y se llevó a cabo un proceso de digestibilidad in vitro. Resultados: la salinidad ejerció un efecto negativo sobre las plantas; sin embargo, la coinoculación de las mismas incrementó su productividad, el contenido en nitrógeno, minerales totales, Ca y Mg. Además, los parámetros fisiológicos de potencial hídrico y concentración de leghemoglobina se incrementaron. Tanto la salinidad como la coinoculación de las plantas aumentaron la capacidad antioxidante de la leguminosa en los dializados y retenidos obtenidos tras someter a la planta a un proceso de digestibilidad in vitro. Conclusión: la coinoculación con E. meliloti y H. maura podría mejorar el cultivo de la alfalfa bajo condiciones específicas de salinidad, aumentando su composición nutricional y funcional, pudiendo considerarse en la formulación de suplementos nutricionales para el consumo humano.

  8. The bHLH Transcription Factors TSAR1 and TSAR2 Regulate Triterpene Saponin Biosynthesis in Medicago truncatula1[OPEN

    PubMed Central

    2016-01-01

    Plants respond to stresses by producing a broad spectrum of bioactive specialized metabolites. Hormonal elicitors, such as jasmonates, trigger a complex signaling circuit leading to the concerted activation of specific metabolic pathways. However, for many specialized metabolic pathways, the transcription factors involved remain unknown. Here, we report on two homologous jasmonate-inducible transcription factors of the basic helix-loop-helix family, TRITERPENE SAPONIN BIOSYNTHESIS ACTIVATING REGULATOR1 (TSAR1) and TSAR2, which direct triterpene saponin biosynthesis in Medicago truncatula. TSAR1 and TSAR2 are coregulated with and transactivate the genes encoding 3-HYDROXY-3-METHYLGLUTARYL-COENZYME A REDUCTASE1 (HMGR1) and MAKIBISHI1, the rate-limiting enzyme for triterpene biosynthesis and an E3 ubiquitin ligase that controls HMGR1 levels, respectively. Transactivation is mediated by direct binding of TSARs to the N-box in the promoter of HMGR1. In transient expression assays in tobacco (Nicotiana tabacum) protoplasts, TSAR1 and TSAR2 exhibit different patterns of transactivation of downstream triterpene saponin biosynthetic genes, hinting at distinct functionalities within the regulation of the pathway. Correspondingly, overexpression of TSAR1 or TSAR2 in M. truncatula hairy roots resulted in elevated transcript levels of known triterpene saponin biosynthetic genes and strongly increased the accumulation of triterpene saponins. TSAR2 overexpression specifically boosted hemolytic saponin biosynthesis, whereas TSAR1 overexpression primarily stimulated nonhemolytic soyasaponin biosynthesis. Both TSARs also activated all genes of the precursor mevalonate pathway but did not affect sterol biosynthetic genes, pointing to their specific role as regulators of specialized triterpene metabolism in M. truncatula. PMID:26589673

  9. Responses of lucerne (Medicago sativa L.) and rhizobia to copper-based fungicide application in two contrasting soils

    NASA Astrophysics Data System (ADS)

    Schneider, Martin; Dober, Melanie; Jöchlinger, Lisa; Keiblinger, Katharina; Soja, Gerhard; Mentler, Axel; Zechmeister-Boltenstern, Sophie; Bruckner, Alexander; Golestani Fard, Alireza; Wenzel, Walter; Zehetner, Franz

    2016-04-01

    For more than 120 years, salts of copper (Cu) have been used in viticulture to prevent damages by fungal diseases. Due to restrictions in the use of synthetic fungicides and mineral fertilizers, organic viticulture depends on Cu as well as on biological nitrogen fixation. Here, we conducted an eco-toxicological pot experiment with an acidic, sandy soil and a calcareous, loamy soil and incrementally increasing fungicide application rates from 0 to 5000 mg Cu kg-1 soil. Lucerne (Medicago sativa L. cultivar. Plato) was grown in the pots for 3 months under greenhouse conditions. Acetylene reduction assays performed with harvested nodules showed no response to elevated soil Cu concentrations indicating that the nitrogen fixing capacity of rhizobia was not compromised by Cu in our experiment. Nevertheless, the nodule biomass was very sensitive to Cu and strongly decreased due to reduced amounts of fine roots and less energy supply by the plant. Legumes are known to be Cu-sensitive, and our contribution also showed a decrease in harvest by 50 % (EC50) at 21 mg kg-1 plant Cu tissue concentration in the acidic soil and at 30 mg kg-1 in the calcareous soil. This corresponded to diffusional fluxes measured by diffusive gradients in thin films (DGT) of 202 and 368 fmol cm-2 s-1, respectively. DGT measurements showed that in the acidic soil, Cu was 2 to 10 times more available for plants, depending on the concentration applied, than in the calcareous soil. A modeling approach for estimating the effective concentration (EC) by including the DGT-estimated plant Cu content and the pH produced more accurate values (NRMSE of 21.9 to 20.1 %) than EC directly estimated from DGT.

  10. Insight into the role of anthocyanin biosynthesis-related genes in Medicago truncatula mutants impaired in pigmentation in leaves.

    PubMed

    Carletti, Giorgia; Lucini, Luigi; Busconi, Matteo; Marocco, Adriano; Bernardi, Jamila

    2013-09-01

    Flavonoids are the most common antioxidant compounds produced in plants. In this study, two wild types and two independent mutants of Medicago truncatula with altered anthocyanin content in leaves were characterized at the phenotype, metabolite profile, gene structure and transcript levels. Flavonoid profiles showed conserved levels of dihydroflavonols, leucoanthocyanidins and flavonols, while anthocyanidin, anthocyanin and isoflavone levels were lower in the mutants (up to 90% less) compared with the wild types. Genes encoding key enzymes of the anthocyanin pathway and transcriptional factors were analyzed by RT-PCR. Genes involved in the later steps of the anthocyanin pathway (dihydrokaempferol reductase 2, UDP-glucose:anthocyanin 3-O-glucosyltransferase and glutathione S-transferase) were found under-expressed in both mutants. Dihydrokaempferol reductase 1 was downregulated two-fold in the anthocyanin-less mutant while the UDP-glucose:anthocyanin 5-O-glucosyltransferase was strongly repressed only in the mutant with low pigmentation, suggesting a different regulation in the two genotypes. The common feature was that the first enzymes of the flavonoid biosynthesis pathway were not altered in rate of expression. A very high reduction in transcript accumulation was also found for two homologous R2R3 MYB genes, namely MtMYBA and AN2, suggesting that these genes have a role in anthocyanin accumulation in leaves. More evidence was found on analyzing their nucleotide sequence: several SNPs, insertions and deletions in the coding and non-coding regions of both MYB genes were found between mutants and wild types that could influence anthocyanin biosynthesis. Moreover, a subfamily of eight MYB genes with a high homology to MtMYBA was discovered in tandem on chromosome 5 of M. truncatula.

  11. Identification of a group of XTHs genes responding to heavy metal mercury, salinity and drought stresses in Medicago truncatula.

    PubMed

    Xuan, Yun; Zhou, Zhao Sheng; Li, Hai Bo; Yang, Zhi Min

    2016-10-01

    Xyloglucan endotransglucosylase/hydrolases (XTH) are one of the key enzymes regulating cell wall construction, extension and metabolism. In the study, 44 XTH protein genes from Medicago truncatula genome were identified using bioinformatics, microarray and RT-PCR. Each XTH was showed to possess a highly conserved domain ((D/N)-E-(I/L/F/V)-D-(F/I/L)-E-(F/L)-L-G), and most of XTHs possess four Cys in the C terminal region, which suggests the potential for generating disulfide bonds. Based on the XTH protein sequences, these XTHscan be classified into three major families and each family can be subdivided into more groups. Examination of the genomic location of XTH genes on M. truncatula chromosomes showed that the evolutional expansion of the genes was possibly attributed to localized gene duplications. To investigate the possible involvement of the XTHs responding to heavy metals and other abiotic stresses, the XTH genes were exposed to heavy metal (Hg or Cu), salt and drought stresses. There were 28, 21 and 21 MtXTH genes found to respond to HgCl2, salt and drought stresses, respectively, but their expression were different under the stresses. Some of the XTH genes were well confirmed by quantitative RT-PCR (qRT-PCR). We further specified expression of a XTH gene Medtr4g128580 (MtXTH3) under different environmental stresses, and showed that MtXTH3 was induced by Hg exposure. These results indicated that a group of MtXTHs could be differentially expressed under the environmental stresses.

  12. LOOSE FLOWER, a WUSCHEL-like Homeobox gene, is required for lateral fusion of floral organs in Medicago truncatula.

    PubMed

    Niu, Lifang; Lin, Hao; Zhang, Fei; Watira, Tezera W; Li, Guifen; Tang, Yuhong; Wen, Jiangqi; Ratet, Pascal; Mysore, Kirankumar S; Tadege, Million

    2015-02-01

    The Medicago truncatula WOX gene, STENOFOLIA (STF), and its orthologs in Petunia, pea, and Nicotiana sylvestris are required for leaf blade outgrowth and floral organ development as demonstrated by severe phenotypes in single mutants. But the Arabidopsis wox1 mutant displays a narrow leaf phenotype only when combined with the prs/wox3 mutant. In maize and rice, WOX3 homologs are major regulators of leaf blade development. Here we investigated the role of WOX3 in M. truncatula development by isolating the lfl/wox3 loss-of-function mutant and performing genetic crosses with the stf mutant. Lack of WOX3 function in M. truncatula leads to a loose-flower (lfl) phenotype, where defects are observed in sepal and petal development, but leaf blades are apparently normal. The stf lfl double mutant analysis revealed that STF and LFL act mainly independently with minor redundant functions in flower development, but LFL has no obvious role in leaf blade outgrowth in M. truncatula on its own or in combination with STF. Interestingly, LFL acts as a transcriptional repressor by recruiting TOPLESS in the same manner as STF does, and can substitute for STF function in leaf blade and flower development if expressed under the STF promoter. STF also complements the lfl mutant phenotype in the flower if expressed under the LFL promoter. Our data suggest that the STF/WOX1 and LFL/WOX3 genes of M. truncatula employ a similar mechanism of action in organizing cell proliferation for lateral outgrowth but may have evolved different cis elements to acquire distinct functions. PMID:25492397

  13. Response of nitrogen fixation in relation to nodule carbohydrate metabolism in Medicago ciliaris lines subjected to salt stress.

    PubMed

    Ben Salah, Imène; Albacete, Alfonso; Martínez Andújar, Cristina; Haouala, Rabiaa; Labidi, Nehla; Zribi, Fethia; Martinez, Vicente; Pérez-Alfocea, Francisco; Abdelly, Chedly

    2009-03-15

    The effect of salt stress on nitrogen fixation, in relation to sucrose transport towards nodules and other sink organs and the potential of sucrose breakdown by nodules, was investigated in two lines of Medicago ciliaris. Under salt stress conditions, the two lines showed a decrease of total biomass production, but TNC 1.8 was less affected by salt than TNC 11.9. The chlorophyll content was not changed in TNC 1.8, in contrast to TNC 11.9. Shoot, root, and nodule biomass were also affected in the two lines, but TNC 1.8 exhibited the higher potentialities of biomass production of these organs. Nitrogen fixation also decreased in the two lines, and was more sensitive to salt than growth parameters. TNC 1.8 consistently exhibited the higher values of nitrogen fixation. Unlike nodules, leaves of both lines were well supplied in nutrients with some exceptions. Specifically, the calcium content decreased in the sensitive line leaves, and the nodule magnesium content was not changed in either line. The tolerant line accumulated more sodium in its leaves. The two lines did not show any differences in the nodule sodium content. Sucrose allocation towards nodules was affected by salt in the two lines, but this constraint did not seem to affect the repartition of sucrose between sink organs. Salt stress induced perturbations in nodule sucrolytic activities in the two lines. It inhibited sucrose synthase, but the inhibition was more marked in TNC 11.9; alkaline/neutral activity was not altered in TNC 1.8, whereas it decreased more than half in TNC 11.9. Thus, the relative tolerance of TNC 1.8 to salt stress could be attributed to a better use of these photoassimilates by nodules and a better supply of bacteroids in malate. The hypothesis of a competition for sucrose between nodules and other sink organs under salt stress could not be verified. PMID:18804311

  14. [Development of Arbuscular Mycorrhiza in Highly Responsive and Mycotrophic Host Plant-Black Medick (Medicago lupulina L.)].

    PubMed

    Yurkova, A P; Jacobi, L M; Gapeeva, N E; Stepanova, G V; Shishova, M F

    2015-01-01

    The main phases of arbuscular mycorrhiza (AM) development were analyzed in black medick (Medicago lupulina) with Glomus intraradices. Methods of light and transmission electron microscopy were used to investigate AM. The first mycorrhization was identified on the seventh day after sowing. M. lupulina with AM-fungus Glomus intraradices formed Arum type of AM. Roots of black medick at fruiting stage (on the 88th day) were characterized by the development of forceful mycelium. The thickness of mycelium was comparable with the vascular system of root central cylinder. The development of vesicules into intraradical spores was shown. Micelium, arbuscules, and vesicules developed in close vicinity to the division zone of root tip. This might be evidence of an active symbiotic interaction between partners. All stages of fungal development and breeding, including intraradical spores (in inter-cellular matrix of root cortex), were identified in the roots of black medick, which indicated an active utilization of host plant nutrient substrates by the mycosymbiont. Plant cell cytoplasm extension was identified around young arbuscular branches but not for intracellular hyphae. The presence of active symbiosis was confirmed by increased accumulation of phosphorus in M. lupulina root tissues under conditions of G. intraradices inoculation and low phosphorus level in the soil. Thus, black medick cultivar-population can be characterized as an ecologically obligate mycotrophic plant under conditions of low level of available phosphorus in the soil. Specific features of AM development in intensively mycotrophic black medick, starting from the stage of the first true leaf until host plant fruiting, were evaluated. The obtained plant-microbe system is a perspective model object for further ultracytological and molecular genetic studies of the mechanisms controlling arbuscular mycorrhiza symbiotic efficiency, including selection and investigation of new symbiotic plant mutants. PMID

  15. Sinorhizobium meliloti strains TII7 and A5 by Multilocus Sequence Typing (MLST) have chromsomes identical with Rm1021 and form an effective and ineffective symbiosis with Medicago truncatula line Jemalong A17, respectively

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The strains TII7 and A5 formed an effective and ineffective symbiosis with Medicago truncatula Jemalong A17, respectively. Both were shown to have identical chromsomes with strains Rm1021 and RCR2011 using a Multilocus Sequence Typing method. The 2260 bp segments of DNA stretching from the 3’ end ...

  16. Arbuscular mycorrhizal symbiosis elicits shoot proteome changes that are modified during cadmium stress alleviation in Medicago truncatula

    PubMed Central

    2011-01-01

    Background Arbuscular mycorrhizal (AM) fungi, which engage a mutualistic symbiosis with the roots of most plant species, have received much attention for their ability to alleviate heavy metal stress in plants, including cadmium (Cd). While the molecular bases of Cd tolerance displayed by mycorrhizal plants have been extensively analysed in roots, very little is known regarding the mechanisms by which legume aboveground organs can escape metal toxicity upon AM symbiosis. As a model system to address this question, we used Glomus irregulare-colonised Medicago truncatula plants, which were previously shown to accumulate and tolerate heavy metal in their shoots when grown in a substrate spiked with 2 mg Cd kg-1. Results The measurement of three indicators for metal phytoextraction showed that shoots of mycorrhizal M. truncatula plants have a capacity for extracting Cd that is not related to an increase in root-to-shoot translocation rate, but to a high level of allocation plasticity. When analysing the photosynthetic performance in metal-treated mycorrhizal plants relative to those only Cd-supplied, it turned out that the presence of G. irregulare partially alleviated the negative effects of Cd on photosynthesis. To test the mechanisms by which shoots of Cd-treated mycorrhizal plants avoid metal toxicity, we performed a 2-DE/MALDI/TOF-based comparative proteomic analysis of the M. truncatula shoot responses upon mycorrhization and Cd exposure. Whereas the metal-responsive shoot proteins currently identified in non-mycorrhizal M. truncatula indicated that Cd impaired CO2 assimilation, the mycorrhiza-responsive shoot proteome was characterised by an increase in photosynthesis-related proteins coupled to a reduction in glugoneogenesis/glycolysis and antioxidant processes. By contrast, Cd was found to trigger the opposite response coupled the up-accumulation of molecular chaperones in shoot of mycorrhizal plants relative to those metal-free. Conclusion Besides drawing a

  17. The arbuscular mycorrhizal fungus Glomus mosseae can enhance arsenic tolerance in Medicago truncatula by increasing plant phosphorus status and restricting arsenate uptake.

    PubMed

    Xu, Pengliang; Christie, Peter; Liu, Yu; Zhang, Junling; Li, Xiaolin

    2008-11-01

    A pot experiment examined the biomass and As uptake of Medicago truncatula colonized by the arbuscular mycorrhizal (AM) fungus Glomus mosseae in low-P soil experimentally contaminated with different levels of arsenate. The biomass of G. mosseae external mycelium was unaffected by the highest addition level of As studied (200 mg kg(-1)) but shoot and root biomass declined in both mycorrhizal and non-mycorrhizal plants, indicating that the AM fungus was more tolerant than M. truncatula to arsenate. Mycorrhizal inoculation increased shoot and root dry weights by enhancing host plant P nutrition and lowering shoot and root As concentrations compared with uninoculated plants. The AM fungus may have been highly tolerant to As and conferred enhanced tolerance to arsenate on the host plant by enhancing P nutrition and restricting root As uptake.

  18. Isolation and characterization of genetic variability in bacteria with β-hemolytic and antifungal activity isolated from the rhizosphere of Medicago truncatula plants.

    PubMed

    Hernández-Salmerón, J E; Prieto-Barajas, C M; Valencia-Cantero, E; Moreno-Hagelsieb, G; Santoyo, G

    2014-07-04

    In the present study, we analyzed the frequency of hemolytic and antifungal activities in bacterial isolates from the rhizosphere of Medicago truncatula plants. Of the 2000 bacterial colonies, 96 showed β-hemolytic activities (frequency, 4.8 x 10(-2)). Hemolytic isolates were analyzed for their genetic diversity by using random amplification of polymorphic DNA, yielding 88 haplotypes. The similarity coefficient of Nei and Li showed a polymorphic diversity ranging from 0.3 to 1. Additionally, 8 of the hemolytic isolates showed antifungal activity toward plant pathogens, Diaporthe phaseolorum, Colletotrichum acutatum, Rhizoctonia solani, and Fusarium oxysporum. The 16S ribosomal sequencing analysis showed that antagonistic bacterial isolates corresponded to Bacillus subtilis (UM15, UM33, UM42, UM49, UM52, and UM91), Bacillus pumilus (UM24), and Bacillus licheniformis (UM88). The present results revealed a higher genetic diversity among hemolytic isolates compared to that of isolates with antifungal action.

  19. Knockdown of CELL DIVISION CYCLE16 Reveals an Inverse Relationship between Lateral Root and Nodule Numbers and a Link to Auxin in Medicago truncatula1[W][OA

    PubMed Central

    Kuppusamy, Kavitha T.; Ivashuta, Sergey; Bucciarelli, Bruna; Vance, Carroll P.; Gantt, J. Stephen; VandenBosch, Kathryn A.

    2009-01-01

    The postembryonic development of lateral roots and nodules is a highly regulated process. Recent studies suggest the existence of cross talk and interdependency in the growth of these two organs. Although plant hormones, including auxin and cytokinin, appear to be key players in coordinating this cross talk, very few genes that cross-regulate root and nodule development have been uncovered so far. This study reports that a homolog of CELL DIVISION CYCLE16 (CDC16), a core component of the Anaphase Promoting Complex, is one of the key mediators in controlling the overall number of lateral roots and nodules. A partial suppression of this gene in Medicago truncatula leads to a decrease in number of lateral roots and a 4-fold increase in number of nodules. The roots showing lowered expression of MtCDC16 also show reduced sensitivity to phytohormone auxin, thus providing a potential function of CDC16 in auxin signaling. PMID:19789288

  20. LYR3, a high-affinity LCO-binding protein of Medicago truncatula, interacts with LYK3, a key symbiotic receptor.

    PubMed

    Fliegmann, Judith; Jauneau, Alain; Pichereaux, Carole; Rosenberg, Charles; Gasciolli, Virginie; Timmers, Antonius C J; Burlet-Schiltz, Odile; Cullimore, Julie; Bono, Jean-Jacques

    2016-05-01

    LYR3, LYK3, and NFP are lysin motif-containing receptor-like kinases (LysM-RLKs) from Medicago truncatula, involved in perception of symbiotic lipo-chitooligosaccharide (LCO) signals. Here, we show that LYR3, a high-affinity LCO-binding protein, physically interacts with LYK3, a key player regulating symbiotic interactions. In vitro, LYR3 is phosphorylated by the active kinase domain of LYK3. Fluorescence lifetime imaging/Förster resonance energy transfer (FLIM/FRET) experiments in tobacco protoplasts show that the interaction between LYR3 and LYK3 at the plasma membrane is disrupted or inhibited by addition of LCOs. Moreover, LYR3 attenuates the cell death response, provoked by coexpression of NFP and LYK3 in tobacco leaves. PMID:27129432

  1. Seed shape in model legumes: approximation by a cardioid reveals differences in ethylene insensitive mutants of Lotus japonicus and Medicago truncatula.

    PubMed

    Cervantes, Emilio; Martín, José Javier; Chan, Pick Kuen; Gresshoff, Peter M; Tocino, Ángel

    2012-09-15

    Seed shape in the model legumes Lotus japonicus and Medicago truncatula is described. Based in previous work with Arabidopsis, the outline of the longitudinal sections of seeds is compared with a cardioid curve. L. japonicus seeds adjust well to an unmodified cardioid, whereas accurate adjustment in M. truncatula is obtained by the simple transformation of scaling the vertical axis by a factor equal to the Golden Ratio. Adjustments of seed shape measurements with simple geometrical forms are essential tools for the statistical analysis of variations in seed shape under different conditions or in mutants. The efficiency of the adjustment to a cardioid in the model plants suggests that seed morphology may be related to genome complexity. Seeds of ethylene insensitive mutants present differences in size and shape as well as altered responses to imbibition. The biological implication and meaning of these relationships are discussed.

  2. Influence of atmospheric [CO2] on growth, carbon allocation and cost of plant tissues on leaf nitrogen concentration maintenance in nodulated Medicago sativa

    NASA Astrophysics Data System (ADS)

    Pereyra, Gabriela; Hartmann, Henrik; Ziegler, Waldemar; Michalzik, Beate; Gonzalez-Meler, Miquel; Trumbore, Susan

    2015-04-01

    Plant carbon (C) allocation and plant metabolic processes (i.e. photosynthesis and respiration) can be affected by changes in C availability, for example from changing atmospheric [CO2]. In nodulated plants, C availability may also influence nitrogen (N) fixation by bacteriods. But C allocation and N fixation are often studied independently and hence do not allow elucidating interactive effects. We investigated how different atmospheric [CO2] (Pleistocene: 170 ppm, ambient: 400 ppm and projected future: 700 ppm) influence plant growth, allocation to nodules, and the ratio of photosynthesis-to-respiration (R:A) as an indicator of C cost in Medicago sativa inoculated with Ensifer meliloti. M. sativa grew c. 38% more nodules at 400 ppm and 700 ppm than at 170 ppm. However, ratios of above- and belowground plant biomass to nodule biomass were constant over time and independent of atmospheric [CO2]. Total non-structural carbohydrate concentrations were not significantly different between plants grown at 400 and 700 ppm, but were four to five-fold higher than in 170 ppm plants. Leaf level N concentration was similar across treatments, but N-based photosynthetic rates were 82% and 93% higher in leaves of plants grown at 400 and 700 ppm, respectively, than plants grown at 170 ppm. In addition, leaf R:A was greater (48% or 55%) in plants grown at 170 ppm than plants grown at 400 and 700 ppm. Similarly, the greatest proportion of assimilated CO2 released by root respiration occurred in rhizobial plants growing at 170 ppm. Our results suggest that C limitation in nodulated Medicago sativa plants did not influence C allocation to nodule biomass but caused a proportionally greater allocation of C to belowground respiration, most likely to bacteriods. This suggests that N tissue concentration was maintained at low [CO2] by revving up bacteriod metabolism and at the expense of non-structural carbohydrate reserves.

  3. Genome-Wide Association Study Identifies Loci for Salt Tolerance during Germination in Autotetraploid Alfalfa (Medicago sativa L.) Using Genotyping-by-Sequencing.

    PubMed

    Yu, Long-Xi; Liu, Xinchun; Boge, William; Liu, Xiang-Ping

    2016-01-01

    Salinity is one of major abiotic stresses limiting alfalfa (Medicago sativa L.) production in the arid and semi-arid regions in US and other counties. In this study, we used a diverse panel of alfalfa accessions previously described by Zhang et al. (2015) to identify molecular markers associated with salt tolerance during germination using genome-wide association study (GWAS) and genotyping-by-sequencing (GBS). Phenotyping was done by germinating alfalfa seeds under different levels of salt stress. Phenotypic data of adjusted germination rates and SNP markers generated by GBS were used for marker-trait association. Thirty six markers were significantly associated with salt tolerance in at least one level of salt treatments. Alignment of sequence tags to the Medicago truncatula genome revealed genetic locations of the markers on all chromosomes except chromosome 3. Most significant markers were found on chromosomes 1, 2, and 4. BLAST search using the flanking sequences of significant markers identified 14 putative candidate genes linked to 23 significant markers. Most of them were repeatedly identified in two or three salt treatments. Several loci identified in the present study had similar genetic locations to the reported QTL associated with salt tolerance in M. truncatula. A locus identified on chromosome 6 by this study overlapped with that by drought in our previous study. To our knowledge, this is the first report on mapping loci associated with salt tolerance during germination in autotetraploid alfalfa. Further investigation on these loci and their linked genes would provide insight into understanding molecular mechanisms by which salt and drought stresses affect alfalfa growth. Functional markers closely linked to the resistance loci would be useful for MAS to improve alfalfa cultivars with enhanced resistance to drought and salt stresses. PMID:27446182

  4. Evaluation of three endemic Mediterranean plant species Atriplex halimus, Medicago lupulina and Portulaca oleracea for Phytoremediation of Ni, Pb and Zn

    NASA Astrophysics Data System (ADS)

    Chami, Ziad Al; Amer, Nasser; Bitar, Lina Al; Mondelli, Donato; Dumontet, Stefano

    2013-04-01

    The success of phytoremediation depends upon the identification of suitable plants species that hyperaccumulate/tolerate heavy metals and produce large amounts of biomass. In this study, three endemic Mediterranean plant species Atriplex halimus, Medicago lupulina and Portulaca oleracea, were grown hydroponically to assess their potential use in phytoremediation of Ni, Pb and Zn and biomass production. The objective of this research is to improve phytoremediation procedures by searching for a new endemic Mediterranean plant species which can be used for phytoremediation of low/moderate contamination in the Mediterranean arid and semiarid conditions and bioenergy production. The hydroponics experiment was carried out in a growth chamber using half strength Hoagland's solution as control (CTR) and 5 concentrations for Pb and Zn (5, 10, 25, 50 and 100 mg L-1) and 3 concentrations for Ni (1, 2, and 5 mg L-1). Complete randomized design with five replications was adopted. Main growth parameters (shoot and root dry weight, shoot and root length and chlorophyll content) were determined. Shoots and roots were analyzed for their metals contents. Some interesting contributions of this research are: (i) plant metal uptake efficiency ranked as follows: A. halimus > M. lupulina > P. oleracea, whereas heavy metal toxicity ranked as follows: Ni > Zn > Pb, (ii) none of the plant species was identified as hyperaccumulator, (iii) Atriplex halimus and Medicago lupulina can accumulate Ni, Pb and Zn in their roots, (iv) translocate small fraction to their above ground biomass, and (v) indicate moderate pollution levels of the environment. In addition, as they are a good biomass producer, they can be used in phytostabilisation of marginal lands and their above ground biomass can be used for livestock feeding as well for bioenergy production.

  5. Ensifer, Phyllobacterium and Rhizobium species occupy nodules of Medicago sativa (alfalfa) and Melilotus alba (sweet clover) grown at a Canadian site without a history of cultivation.

    PubMed

    Bromfield, E S P; Tambong, J T; Cloutier, S; Prévost, D; Laguerre, G; van Berkum, P; Thi, T V Tran; Assabgui, R; Barran, L R

    2010-02-01

    Phage-resistant and -susceptible bacteria from nodules of alfalfa and sweet clover, grown at a site without a known history of cultivation, were identified as diverse genotypes of Ensifer, Rhizobium and Phyllobacterium species based on sequence analysis of ribosomal (16S and 23S rRNA) and protein-encoding (atpD and recA) genes, Southern hybridization/RFLP and a range of phenotypic characteristics. Among phage-resistant bacteria, one genotype of Rhizobium sp. predominated on alfalfa (frequency approximately 68 %) but was recovered infrequently ( approximately 1 %) from sweet clover. A second genotype was isolated infrequently only from alfalfa. These genotypes fixed nitrogen poorly in association with sweet clover and Phaseolus vulgaris, but were moderately effective with alfalfa. They produced a near-neutral reaction on mineral salts agar containing mannitol, which is atypical of the genus Rhizobium. A single isolate of Ensifer sp. and two of Phyllobacterium sp. were recovered only from sweet clover. All were highly resistant to multiple antibiotics. Phylogenetic analysis indicated that Ensifer sp. strain T173 is closely related to, but separate from, the non-symbiotic species 'Sinorhizobium morelense'. Strain T173 is unique in that it possesses a 175 kb symbiotic plasmid and elicits ineffective nodules on alfalfa, sweet clover, Medicago lupulina and Macroptilium atropurpureum. The two Phyllobacterium spp. were non-symbiotic and probably represent bacterial opportunists. Three genotypes of E. meliloti that were symbiotically effective with alfalfa and sweet clover were encountered infrequently. Among phage-susceptible isolates, two genotypes of E. medicae were encountered infrequently and were highly effective with alfalfa, sweet clover and Medicago polymorpha. The ecological and practical implications of the findings are discussed.

  6. Genome-Wide Association Study Identifies Loci for Salt Tolerance during Germination in Autotetraploid Alfalfa (Medicago sativa L.) Using Genotyping-by-Sequencing

    PubMed Central

    Yu, Long-Xi; Liu, Xinchun; Boge, William; Liu, Xiang-Ping

    2016-01-01

    Salinity is one of major abiotic stresses limiting alfalfa (Medicago sativa L.) production in the arid and semi-arid regions in US and other counties. In this study, we used a diverse panel of alfalfa accessions previously described by Zhang et al. (2015) to identify molecular markers associated with salt tolerance during germination using genome-wide association study (GWAS) and genotyping-by-sequencing (GBS). Phenotyping was done by germinating alfalfa seeds under different levels of salt stress. Phenotypic data of adjusted germination rates and SNP markers generated by GBS were used for marker-trait association. Thirty six markers were significantly associated with salt tolerance in at least one level of salt treatments. Alignment of sequence tags to the Medicago truncatula genome revealed genetic locations of the markers on all chromosomes except chromosome 3. Most significant markers were found on chromosomes 1, 2, and 4. BLAST search using the flanking sequences of significant markers identified 14 putative candidate genes linked to 23 significant markers. Most of them were repeatedly identified in two or three salt treatments. Several loci identified in the present study had similar genetic locations to the reported QTL associated with salt tolerance in M. truncatula. A locus identified on chromosome 6 by this study overlapped with that by drought in our previous study. To our knowledge, this is the first report on mapping loci associated with salt tolerance during germination in autotetraploid alfalfa. Further investigation on these loci and their linked genes would provide insight into understanding molecular mechanisms by which salt and drought stresses affect alfalfa growth. Functional markers closely linked to the resistance loci would be useful for MAS to improve alfalfa cultivars with enhanced resistance to drought and salt stresses. PMID:27446182

  7. The molecular genetic linkage map of the model legume Medicago truncatula: an essential tool for comparative legume genomics and the isolation of agronomically important genes

    PubMed Central

    Thoquet, Philippe; Ghérardi, Michele; Journet, Etienne-Pascal; Kereszt, Attila; Ané, Jean-Michel; Prosperi, Jean-Marie; Huguet, Thierry

    2002-01-01

    Background The legume Medicago truncatula has emerged as a model plant for the molecular and genetic dissection of various plant processes involved in rhizobial, mycorrhizal and pathogenic plant-microbe interactions. Aiming to develop essential tools for such genetic approaches, we have established the first genetic map of this species. Two parental homozygous lines were selected from the cultivar Jemalong and from the Algerian natural population (DZA315) on the basis of their molecular and phenotypic polymorphism. Results An F2 segregating population of 124 individuals between these two lines was obtained using an efficient manual crossing technique established for M. truncatula and was used to construct a genetic map. This map spans 1225 cM (average 470 kb/cM) and comprises 289 markers including RAPD, AFLP, known genes and isoenzymes arranged in 8 linkage groups (2n = 16). Markers are uniformly distributed throughout the map and segregation distortion is limited to only 3 linkage groups. By mapping a number of common markers, the eight linkage groups are shown to be homologous to those of diploid alfalfa (M. sativa), implying a good level of macrosynteny between the two genomes. Using this M. truncatula map and the derived F3 populations, we were able to map the Mtsym6 symbiotic gene on linkage group 8 and the SPC gene, responsible for the direction of pod coiling, on linkage group 7. Conclusions These results demonstrate that Medicago truncatula is amenable to diploid genetic analysis and they open the way to map-based cloning of symbiotic or other agronomically-important genes using this model plant. PMID:11825338

  8. A Medicago truncatula Cystathionine-β-Synthase-like Domain-Containing Protein Is Required for Rhizobial Infection and Symbiotic Nitrogen Fixation1[OPEN

    PubMed Central

    Breakspear, Andrew; Guan, Dian; Nakashima, Jin; Zhang, Shulan; Torres-Jerez, Ivone; Oldroyd, Giles; Murray, Jeremy D.; Udvardi, Michael K.

    2016-01-01

    The symbiosis between leguminous plants and soil rhizobia culminates in the formation of nitrogen-fixing organs called nodules that support plant growth. Two Medicago truncatula Tnt1-insertion mutants were identified that produced small nodules, which were unable to fix nitrogen effectively due to ineffective rhizobial colonization. The gene underlying this phenotype was found to encode a protein containing a putative membrane-localized domain of unknown function (DUF21) and a cystathionine-β-synthase domain. The cbs1 mutants had defective infection threads that were sometimes devoid of rhizobia and formed small nodules with greatly reduced numbers of symbiosomes. We studied the expression of the gene, designated M. truncatula Cystathionine-β-Synthase-like1 (MtCBS1), using a promoter-β-glucuronidase gene fusion, which revealed expression in infected root hair cells, developing nodules, and in the invasion zone of mature nodules. An MtCBS1-GFP fusion protein localized itself to the infection thread and symbiosomes. Nodulation factor-induced Ca2+ responses were observed in the cbs1 mutant, indicating that MtCBS1 acts downstream of nodulation factor signaling. MtCBS1 expression occurred exclusively during Medicago-rhizobium symbiosis. Induction of MtCBS1 expression during symbiosis was found to be dependent on Nodule Inception (NIN), a key transcription factor that controls both rhizobial infection and nodule organogenesis. Interestingly, the closest homolog of MtCBS1, MtCBS2, was specifically induced in mycorrhizal roots, suggesting common infection mechanisms in nodulation and mycorrhization. Related proteins in Arabidopsis have been implicated in cell wall maturation, suggesting a potential role for CBS1 in the formation of the infection thread wall. PMID:26884486

  9. Decomposition of olive mill waste compost, goat manure and Medicago sativa in Lebanese soils using the litterbag technique

    NASA Astrophysics Data System (ADS)

    Atallah, Therese

    2014-05-01

    Organic amendments, green manure and plant residues incorporation are the main sources of nutrients in organic farming, their decomposition rate is crucial for the accumulation and long-term storage of organic matter in soils. In this study the decomposition of compost from olive mill waste (N: 29.3 g kg-1; total dissolved nitrogen or TDN: 3.82 g kg-1), goat manure (N: 31.5 g kg-1; TDN: 0.94 g kg-1), the shoots (N: 33.6 g kg-1; TDN: 17.57 g kg-1) and roots (N: 22.12 g kg-1; TDN: 8.87 g kg-1) of Medicago sativa was followed in three Lebanese soils. The nitrogen, phosphorus and potassium released were followed over one year, starting in early winter (December-January). The mild sub-humid Mediterranean conditions allowed a rapid mass loss in alfalfa shoots 30 days after incorporation. Manure and compost were more persistent. Between 80 and 90% of TDN were released, after 30 days of in-situ incubation for compost, the release was over 90% for alfalfa shoots. The movement of P was slower, as the compost (6.99 g kg-1 of P) and manure (9.81 g kg-1 of P) lost 33% and 22%, respectively, during 30 days of incubation. After one year, 15 to 35% of P remained in the soils. The manure was the richest in potassium (19.66 g kg-1) followed by the alfalfa shoots (15.56 g kg-1), the compost (8.19 g kg-1) and the roots (5.96 g kg-1). The loss of potassium was important, as over 88% had disappeared over the year. All decomposition curves followed an exponential model. The calculated coefficients of decomposition for total nitrogen (lnfinal - lninitial/days) were significantly higher for alfalfa shoots (0.00547 day-1) and similar for the compost (0.00184 day-1) and the manure (0.00175 day-1). The ANOVA test showed a difference between two of the sites (Site A: 521 g kg-1 of clay and 42 g kg-1 of calcium carbonate; Site S: 260 g kg-1 of clay and 269 g kg-1 of CaCO3) and the third one (Site L: 315 g kg-1 of clay and 591 g kg-1 of CaCO3). The relationships between the soil calcium

  10. Two alternative recessive quantitative trait loci influence resistance to spring black stem and leaf spot in Medicago truncatula

    PubMed Central

    Kamphuis, Lars G; Lichtenzveig, Judith; Oliver, Richard P; Ellwood, Simon R

    2008-01-01

    Background Knowledge of the genetic basis of plant resistance to necrotrophic pathogens is incomplete and has been characterised in relatively few pathosystems. In this study, the cytology and genetics of resistance to spring black stem and leaf spot caused by Phoma medicaginis, an economically important necrotrophic pathogen of Medicago spp., was examined in the model legume M. truncatula. Results Macroscopically, the resistant response of accession SA27063 was characterised by small, hypersensitive-like spots following inoculation while the susceptible interaction with accessions A17 and SA3054 showed necrotic lesions and spreading chlorosis. No unique cytological differences were observed during early infection (<48 h) between the resistant and susceptible genotypes, except pathogen growth was restricted to one or a few host cells in SA27063. In both interactions reactive oxygen intermediates and phenolic compounds were produced, and cell death occurred. Two F2 populations segregating for resistance to spring black stem and leaf spot were established between SA27063 and the two susceptible accessions, A17 and SA3054. The cross between SA27063 and A17 represented a wider cross than between SA27063 and SA3054, as evidenced by higher genetic polymorphism, reduced fertility and aberrant phenotypes of F2 progeny. In the SA27063 × A17 F2 population a highly significant quantitative trait locus (QTL, LOD = 7.37; P < 0.00001) named resistance to the necrotroph Phoma medicaginis one (rnpm1) genetically mapped to the top arm of linkage group 4 (LG4). rnpm1 explained 33.6% of the phenotypic variance in the population's response to infection depicted on a 1–5 scale and was tightly linked to marker AW256637. A second highly significant QTL (LOD = 6.77; P < 0.00001), rnpm2, was located on the lower arm of LG8 in the SA27063 × SA3054 map. rnpm2 explained 29.6% of the phenotypic variance and was fine mapped to a 0.8 cM interval between markers h2_16a6a and h2_21h11d. rnpm1

  11. Nodulation by Sinorhizobium meliloti originated from a mining soil alleviates Cd toxicity and increases Cd-phytoextraction in Medicago sativa L.

    PubMed Central

    Ghnaya, Tahar; Mnassri, Majda; Ghabriche, Rim; Wali, Mariem; Poschenrieder, Charlotte; Lutts, Stanley; Abdelly, Chedly

    2015-01-01

    Besides their role in nitrogen supply to the host plants as a result of symbiotic N fixation, the association between legumes and Rhizobium could be useful for the rehabilitation of metal-contaminated soils by phytoextraction. A major limitation presents the metal-sensitivity of the bacterial strains. The aim of this work was to explore the usefulness of Sinorhizobium meliloti originated from a mining site for Cd phytoextraction by Medicago sativa. Inoculated and non-inoculated plants were cultivated for 60 d on soils containing 50 and/or 100 mg Cd kg−1 soil. The inoculation hindered the occurrence of Cd- induced toxicity symptoms that appeared in the shoots of non-inoculated plants. This positive effect of S. meliloti colonization was accompanied by an increase in biomass production and improved nutrient acquisition comparatively to non-inoculated plants. Nodulation enhanced Cd absorption by the roots and Cd translocation to the shoots. The increase of plant biomass concomitantly with the increase of Cd shoot concentration in inoculated plants led to higher potential of Cd-phytoextraction in these plants. In the presence of 50 mg Cd kg−1 in the soil, the amounts of Cd extracted in the shoots were 58 and 178 μg plant−1 in non-inoculated and inoculated plants, respectively. This study demonstrates that this association M. sativa-S. meliloti may be an efficient biological system to extract Cd from contaminated soils. PMID:26528320

  12. Co-downregulation of the hydroxycinnamoyl-CoA:shikimate hydroxycinnamoyl transferase and coumarate 3-hydroxylase significantly increases cellulose content in transgenic alfalfa (Medicago sativa L.).

    PubMed

    Tong, Zongyong; Li, Heng; Zhang, Rongxue; Ma, Lei; Dong, Jiangli; Wang, Tao

    2015-10-01

    Lignin is a component of the cell wall that is essential for growth, development, structure and pathogen resistance in plants, but high lignin is an obstacle to the conversion of cellulose to ethanol for biofuel. Genetically modifying lignin and cellulose contents can be a good approach to overcoming that obstacle. Alfalfa (Medicago sativa L.) is rich in lignocellulose biomass and used as a model plant for the genetic modification of lignin in this study. Two key enzymes in the lignin biosynthesis pathway-hydroxycinnamoyl -CoA:shikimate hydroxycinnamoyl transferase (HCT) and coumarate 3-hydroxylase (C3H)-were co-downregulated. Compared to wild-type plants, the lignin content in the modified strain was reduced by 38%, cellulose was increased by 86.1%, enzyme saccharification efficiency was increased by 10.9%, and cell wall digestibility was increased by 13.0%. The modified alfalfa exhibited a dwarf phenotype, but normal above ground biomass. This approach provides a new strategy for reducing lignin and increasing cellulose contents and creates a new genetically modified crop with enhanced value for biofuel.

  13. Overexpression of Medicago sativa TMT elevates the α-tocopherol content in Arabidopsis seeds, alfalfa leaves, and delays dark-induced leaf senescence.

    PubMed

    Jiang, Jishan; Jia, Huili; Feng, Guangyan; Wang, Zan; Li, Jun; Gao, Hongwen; Wang, Xuemin

    2016-08-01

    Alfalfa (Medicago sativa L.) is a major forage legume for livestock and a target for improving their dietary quality. Vitamin E is an essential vitamin that animals must obtain from their diet for proper growth and development. γ-tocopherol methyltransferase (γ-TMT), which catalyzes the conversion of δ- and γ-tocopherols (or tocotrienols) to β- and α-tocopherols (or tocotrienols), respectively, is the final enzyme involved in the vitamin E biosynthetic pathway. The overexpression of M. sativa L.'s γ-TMT (MsTMT) increased the α-tocopherol content 10-15 fold above that of wild type Arabidopsis seeds without altering the total content of vitamin E. Additionally, in response to osmotic stress, the biomass and the expression levels of several osmotic marker genes were significantly higher in the transgenic lines compared with wild type. Overexpression of MsTMT in alfalfa led to a modest, albeit significant, increase in α-tocopherol in leaves and was also responsible for a delayed leaf senescence phenotype. Additionally, the crude protein content was increased, while the acid and neutral detergent fiber contents were unchanged in these transgenic lines. Thus, increased α-tocopherol content occurred in transgenic alfalfa without compromising the nutritional qualities. The targeted metabolic engineering of vitamin E biosynthesis through MsTMT overexpression provides a promising approach to improve the α-tocopherol content of forage crops.

  14. The structure of Medicago truncatula δ1-pyrroline-5-carboxylate reductase provides new insights into regulation of proline biosynthesis in plants

    DOE PAGES

    Ruszkowski, Milosz; Nocek, Boguslaw; Forlani, Giuseppe; Dauter, Zbigniew

    2015-10-30

    The two pathways for proline biosynthesis in higher plants share the last step, the conversion of δ1-pyrroline-5-carboxylate (P5C) to L-proline, which is catalyzed by P5C reductase (P5CR, EC 1.5.1.2) with the use of NAD(P)H as a coenzyme. There is increasing amount of evidence to suggest a complex regulation of P5CR activity at the post-translational level, yet the molecular basis of these mechanisms is unknown. Here we report the three-dimensional structure of the P5CR enzyme from the model legume Medicago truncatula (Mt). The crystal structures of unliganded MtP5CR decamer, and its complexes with the products NAD+, NADP+, and L-proline were refinedmore » using x-ray diffraction data (at 1.7, 1.85, 1.95, and 2.1 Å resolution, respectively). Based on the presented structural data, the coenzyme preference for NADPH over NADH was explained, and NADPH is suggested to be the only coenzyme used by MtP5CR in vivo. Moreover, the insensitivity of MtP5CR to feed-back inhibition by proline, revealed by enzymatic analysis, was correlated with structural features. Additionally, a mechanism for the modulation of enzyme activity by chloride anions is discussed, as well as the rationale for the possible development of effective enzyme inhibitors.« less

  15. CRISPR/Cas mutagenesis of soybean and Medicago truncatula using a new web-tool and a modified Cas9 enzyme.

    PubMed

    Michno, Jean-Michel; Wang, Xiaobo; Liu, Junqi; Curtin, Shaun J; Kono, Thomas Jy; Stupar, Robert M

    2015-01-01

    The CRISPR/Cas9 system is rapidly becoming the reagent of choice for targeted mutagenesis and gene editing in crop species. There are currently intense research efforts in the crop sciences to identify efficient CRISPR/Cas9 platforms to carry out targeted mutagenesis and gene editing projects. These efforts typically result in the incremental tweaking of various platform components including the identification of crop-specific promoters and terminators for optimal expression of the Cas9 enzyme and identification of promoters for expression of the CRISPR guide RNA. In this report, we demonstrate the development of an online web tool for fast identification of CRISPR/Cas9 target loci within soybean gene models, and generic DNA sequences. The web-tool described in this work can quickly identify a high number of potential CRISPR/Cas9 target sites, including restriction enzyme sites that can facilitate the detection of new mutations. In conjunction with the web tool, a soybean codon-optimized CRISPR/Cas9 platform was designed to direct double-stranded breaks to the targeted loci in hairy root transformed cells. The modified Cas9 enzyme was shown to successfully mutate target genes in somatic cells of 2 legume species, soybean and Medicago truncatula. These new tools may help facilitate targeted mutagenesis in legume and other plant species. PMID:26479970

  16. Identification of genes that regulate phosphate acquisition and plant performance during arbuscular my corrhizal symbiosis in medicago truncatula and brachypodium distachyon

    SciTech Connect

    Harrison, Maria J; Hudson, Matthew E

    2015-11-24

    Most vascular flowering plants have the ability to form symbiotic associations with arbuscular mycorrhizal (AM) fungi. The symbiosis develops in the roots and can have a profound effect on plant productivity, largely through improvements in plant mineral nutrition. Within the root cortical cells, the plant and fungus create novel interfaces specialized for nutrient transfer, while the fungus also develops a network of hyphae in the rhizosphere. Through this hyphal network, the fungus acquires and delivers phosphate and nitrogen to the root. In return, the plant provides the fungus with carbon. In addition, to enhancing plant mineral nutrition, the AM symbiosis has an important role in the carbon cycle, and positive effects on soil health. Here we identified and characterized plant genes involved in the regulation and functioning of the AM symbiosis in Medicago truncatula and Brachypodium distachyon. This included the identification and and characterization of a M. truncatula transcription factors that are required for symbiosis. Additionally, we investigated the molecular basis of functional diversity among AM symbioses in B. distachyon and analysed the transcriptome of Brachypodium distachyon during symbiosis.

  17. Co-transforming bar and CsLEA enhanced tolerance to drought and salt stress in transgenic alfalfa (Medicago sativa L.).

    PubMed

    Zhang, Jiyu; Duan, Zhen; Zhang, Daiyu; Zhang, Jianquan; Di, Hongyan; Wu, Fan; Wang, Yanrong

    2016-03-25

    Drought and high salinity are two major abiotic factors that restrict alfalfa productivity. A dehydrin protein, CsLEA, from the desert grass Cleistogenes songorica was transformed into alfalfa (Medicago sativa L.) via Agrobacterium-mediated transformation using the bar gene as a selectable marker, and the drought and salt stress tolerances of the transgenic plants were assessed. Thirty-nine of 119 transformants were positive, as screened by Basta, and further molecularly authenticated using PCR and RT-PCR. Phenotype observations revealed that the transgenic plants grew better than the wild-type (WT) plants after 15d of drought stress and 10d of salt stress: the leaves of WT alfalfa turned yellow, whereas the transgenic alfalfa leaves only wilted; after rewatering, the transgenic plants returned to a normal state, though the WT plants could not be restored. Evaluation of physiologic and biochemical indices during drought and salt stresses showed a relatively lower Na(+) content in the leaves of the transgenic plants, which would reduce toxic ion effects. In addition, the transgenic plants were able to maintain a higher relative water content (RWC), higher shoot biomass, fewer photosystem changes, decreased membrane injury, and a lower level of osmotic stress injury. These results demonstrate that overexpression of the CsLEA gene can enhance the drought and salt tolerance of transgenic alfalfa; in addition, carrying the bar gene in the genome may increase herbicide resistance.

  18. Do jasmonates play a role in arbuscular mycorrhiza-induced local bioprotection of Medicago truncatula against root rot disease caused by Aphanomyces euteiches?

    PubMed

    Hilou, Adama; Zhang, Haoqiang; Franken, Philipp; Hause, Bettina

    2014-01-01

    Bioprotective effects of mycorrhization with two different arbuscular mycorrhizal (AM) fungi, Funneliformis mosseae and Rhizophagus irregularis, against Aphanomyces euteiches, the causal agent of root rot in legumes, were studied in Medicago truncatula using phenotypic and molecular markers. Previous inoculation with an AM-fungus reduced disease symptoms as well as the amount of pathogen within roots, as determined by the levels of A. euteiches rRNA or transcripts of the gene sterol C24 reductase. Inoculation with R. irregularis was as efficient as that with F. mosseae. To study whether jasmonates play a regulatory role in bioprotection of M. truncatula by the AM fungi, composite plants harboring transgenic roots were used to modulate the expression level of the gene encoding M. truncatula allene oxide cyclase 1, a key enzyme in jasmonic acid biosynthesis. Neither an increase nor a reduction in allene oxide cyclase levels resulted in altered bioprotection by the AM fungi against root infection by A. euteiches. These data suggest that jasmonates do not play a major role in the local bioprotective effect of AM fungi against the pathogen A. euteiches in M. truncatula roots.

  19. Palmate-like pentafoliata1 encodes a novel Cys(2)His(2) zinc finger transcription factor essential for compound leaf morphogenesis in Medicago truncatula

    PubMed Central

    2010-01-01

    As the primary site for photosynthetic carbon fixation and the interface between plants and the environment, plant leaves play a key role in plant growth, biomass production and survival, and global carbon and oxygen cycles. Leaves can be simple with a single blade or compound with multiple units of blades known as leaflets. In a palmate-type compound leaf, leaflets are clustered at the tip of the leaf. In a pinnate-type compound leaf, on the other hand, leaflets are placed on a rachis in distance from each other. Higher orders of complexities such as bipinnate compound leaves of the “sensitive” plant, Mimosa pudica, also occur in nature. However, how different leaf morphologies are determined is still poorly understood. Medicago truncatula is a model legume closely related to alfalfa and soybean with trifoliate compound leaves. Recently, we have shown that Palmate-like Pentafoliata1 (PALM1) encodes a putative Cys(2) His(2) zinc finger transcription factor essential for compound leaf morphogenesis in M. truncatula. Here, we present our phylogenetic relationship analysis of PALM1 homologs from different species and demonstrate that PALM1 has transcriptional activity in the transactivation assay in yeast. PMID:20724826

  20. Effect of Medicago sativa L. and compost on organic and inorganic pollutant removal from a mixed contaminated soil and risk assessment using ecotoxicological tests.

    PubMed

    Marchand, Charlotte; Hogland, William; Kaczala, Fabio; Jani, Yahya; Marchand, Lilian; Augustsson, Anna; Hijri, Mohamed

    2016-11-01

    Several Gentle Remediation Options (GRO), e.g., plant-based options (phytoremediation), singly and combined with soil amendments, can be simultaneously efficient for degrading organic pollutants and either stabilizing or extracting trace elements (TEs). Here, a 5-month greenhouse trial was performed to test the efficiency of Medicago sativa L., singly and combined with a compost addition (30% w/w), to treat soils contaminated by petroleum hydrocarbons (PHC), Co and Pb collected at an auto scrap yard. After 5 months, total soil Pb significantly decreased in the compost-amended soil planted with M. sativa, but not total soil Co. Compost incorporation into the soil promoted PHC degradation, M. sativa growth and survival, and shoot Pb concentrations [3.8 mg kg(-1) dry weight (DW)]. Residual risk assessment after the phytoremediation trial showed a positive effect of compost amendment on plant growth and earthworm development. The O2 uptake by soil microorganisms was lower in the compost-amended soil, suggesting a decrease in microbial activity. This study underlined the benefits of the phytoremediation option based on M. sativa cultivation and compost amendment for remediating PHC- and Pb-contaminated soils.

  1. Loss of function of cinnamyl alcohol dehydrogenase 1 leads to unconventional lignin and a temperature-sensitive growth defect in Medicago truncatula

    PubMed Central

    Zhao, Qiao; Tobimatsu, Yuki; Zhou, Rui; Pattathil, Sivakumar; Gallego-Giraldo, Lina; Fu, Chunxiang; Jackson, Lisa A.; Hahn, Michael G.; Kim, Hoon; Chen, Fang; Ralph, John; Dixon, Richard A.

    2013-01-01

    There is considerable debate over the capacity of the cell wall polymer lignin to incorporate unnatural monomer units. We have identified Tnt1 retrotransposon insertion mutants of barrel medic (Medicago truncatula) that show reduced lignin autofluorescence under UV microscopy and red coloration in interfascicular fibers. The phenotype is caused by insertion of retrotransposons into a gene annotated as encoding cinnamyl alcohol dehydrogenase, here designated M. truncatula CAD1. NMR analysis indicated that the lignin is derived almost exclusively from coniferaldehyde and sinapaldehyde and is therefore strikingly different from classical lignins, which are derived mainly from coniferyl and sinapyl alcohols. Despite such a major alteration in lignin structure, the plants appear normal under standard conditions in the greenhouse or growth chamber. However, the plants are dwarfed when grown at 30 °C. Glycome profiling revealed an increased extractability of some xylan and pectin epitopes from the cell walls of the cad1-1 mutant but decreased extractability of others, suggesting that aldehyde-dominant lignin significantly alters cell wall structure. PMID:23901113

  2. The Transcriptional Repressor MYB2 Regulates Both Spatial and Temporal Patterns of Proanthocyandin and Anthocyanin Pigmentation in Medicago truncatula[OPEN

    PubMed Central

    2015-01-01

    Accumulation of anthocyanins and proanthocyanidins (PAs) is limited to specific cell types and developmental stages, but little is known about how antagonistically acting transcriptional regulators work together to determine temporal and spatial patterning of pigmentation at the cellular level, especially for PAs. Here, we characterize MYB2, a transcriptional repressor regulating both anthocyanin and PA biosynthesis in the model legume Medicago truncatula. MYB2 was strongly upregulated by MYB5, a major regulator of PA biosynthesis in M. truncatula and a component of MYB-basic helix loop helix-WD40 (MBW) activator complexes. Overexpression of MYB2 abolished anthocyanin and PA accumulation in M. truncatula hairy roots and Arabidopsis thaliana seeds, respectively. Anthocyanin deposition was expanded in myb2 mutant seedlings and flowers accompanied by increased anthocyanin content. PA mainly accumulated in the epidermal layer derived from the outer integument in the M. truncatula seed coat, starting from the hilum area. The area of PA accumulation and ANTHOCYANIDIN REDUCTASE expression was expanded into the seed body at the early stage of seed development in the myb2 mutant. Genetic, biochemical, and cell biological evidence suggests that MYB2 functions as part of a multidimensional regulatory network to define the temporal and spatial pattern of anthocyanin and PA accumulation linked to developmental processes. PMID:26410301

  3. A vapBC-type toxin-antitoxin module of Sinorhizobium meliloti influences symbiotic efficiency and nodule senescence of Medicago sativa.

    PubMed

    Lipuma, Justine; Cinege, Gyöngyi; Bodogai, Monica; Oláh, Boglárka; Kiers, Aurélie; Endre, Gabriella; Dupont, Laurence; Dusha, Ilona

    2014-12-01

    The symbiotic nitrogen-fixing soil bacterium Sinorhizobium meliloti carries a large number of toxin-antitoxin (TA) modules both on the chromosome and megaplasmids. One of them, the vapBC-5 module that belongs to the type II systems was characterized here. It encodes an active toxin vapC-5, and was shown to be controlled negatively by the complex of its own proteins. Different mutants of the vapBC-5 genes exhibited diverse effects on symbiotic efficiency during interaction with the host plant Medicago sativa. The absence of the entire vapBC-5 region had no influence on nodule formation and nitrogen fixation properties. The strain carrying an insertion in the antitoxin gene showed a reduced nitrogen fixation capacity resulting in a lower plant yield. In contrast, when the toxin gene was mutated, the strain developed more efficient symbiosis with the host plant. The nitrogen fixing root nodules had a delayed senescent phenotype and contained elevated level of plant-derived molecules characteristic of later steps of nodule development. The longer bacteroid viability and abundance of active nitrogen fixing zone resulted in increased production of plant material. These data indicate that modification of the toxin/antitoxin production may influence bacteroid metabolism and may have an impact on the adaptation to changing environmental conditions. PMID:25156344

  4. Unexpectedly low nitrogen acquisition and absence of root architecture adaptation to nitrate supply in a Medicago truncatula highly branched root mutant

    PubMed Central

    Bourion, Virginie

    2014-01-01

    To complement N2 fixation through symbiosis, legumes can efficiently acquire soil mineral N through adapted root architecture. However, root architecture adaptation to mineral N availability has been little studied in legumes. Therefore, this study investigated the effect of nitrate availability on root architecture in Medicago truncatula and assessed the N-uptake potential of a new highly branched root mutant, TR185. The effects of varying nitrate supply on both root architecture and N uptake were characterized in the mutant and in the wild type. Surprisingly, the root architecture of the mutant was not modified by variation in nitrate supply. Moreover, despite its highly branched root architecture, TR185 had a permanently N-starved phenotype. A transcriptome analysis was performed to identify genes differentially expressed between the two genotypes. This analysis revealed differential responses related to the nitrate acquisition pathway and confirmed that N starvation occurred in TR185. Changes in amino acid content and expression of genes involved in the phenylpropanoid pathway were associated with differences in root architecture between the mutant and the wild type. PMID:24706718

  5. Nod factor-treated Medicago truncatula roots and seeds show an increased number of nodules when inoculated with a limiting population of Sinorhizobium meliloti.

    PubMed

    Macchiavelli, Raúl E; Brelles-Mariño, Graciela

    2004-12-01

    Medicago truncatula is a model legume plant that interacts symbiotically with Sinorhizobium meliloti, the alfalfa symbiont. This process involves a molecular dialogue between the bacterium and the plant. Legume roots exude flavonoids that induce the expression of a set of rhizobial genes, the nod genes, which are essential for nodulation and determination of the host range. In turn, nod genes control the synthesis of lipo-chito-oligosaccharides (LCOs), Nod factors, which are bacteria-to-plant signal molecules mediating recognition and nodule organogenesis. M. truncatula roots or seeds have been treated with Nod factors and hydroponically growing seedlings have been inoculated with a limiting population of S. meliloti. It has been shown that submicromolar concentrations of Nod factors increase the number of nodules per plant on M. truncatula. Compared with roots, this increase is more noticeable when seeds are treated. M. truncatula seeds are receptive to submicromolar concentrations of Nod factors, suggesting the possibility of a high affinity LCO perception system in seeds or embryos as well. PMID:15361530

  6. Assessing quality of Medicago sativa silage by monitoring bacterial composition with single molecule, real-time sequencing technology and various physiological parameters.

    PubMed

    Bao, Weichen; Mi, Zhihui; Xu, Haiyan; Zheng, Yi; Kwok, Lai Yu; Zhang, Heping; Zhang, Wenyi

    2016-01-01

    The present study applied the PacBio single molecule, real-time sequencing technology (SMRT) in evaluating the quality of silage production. Specifically, we produced four types of Medicago sativa silages by using four different lactic acid bacteria-based additives (AD-I, AD-II, AD-III and AD-IV). We monitored the changes in pH, organic acids (including butyric acid, the ratio of acetic acid/lactic acid, γ-aminobutyric acid, 4-hyroxy benzoic acid and phenyl lactic acid), mycotoxins, and bacterial microbiota during silage fermentation. Our results showed that the use of the additives was beneficial to the silage fermentation by enhancing a general pH and mycotoxin reduction, while increasing the organic acids content. By SMRT analysis of the microbial composition in eight silage samples, we found that the bacterial species number and relative abundances shifted apparently after fermentation. Such changes were specific to the LAB species in the additives. Particularly, Bacillus megaterium was the initial dominant species in the raw materials; and after the fermentation process, Pediococcus acidilactici and Lactobacillus plantarum became the most prevalent species, both of which were intrinsically present in the LAB additives. Our data have demonstrated that the SMRT sequencing platform is applicable in assessing the quality of silage. PMID:27340760

  7. The photosynthetic and stomatal response of Medicago sativa cv. saranac to free-air CO{sub 2} enrichment (F.A.C.E.) and nitrogen

    SciTech Connect

    Bridson, N.P.

    1996-08-01

    Plots of Medicago sativa cv. saranac were grown in the field at ambient (355 {mu}mol CO{sub 2} mol{sup -1} air) or elevated (600{mu}mol CO{sub 2} mol{sup -1} air) CO{sub 2} concentrations. High (200kg yr{sup -1}) or low (20kg yr{sup -1}) nitrogen levels were applied to two isogeneic lines, one able and one unable to use nitrogen fixing bacteria. Plants were in the second year of field growth. Exposure to elevated CO{sub 2} was via a Free-Air CO{sub 2} Enrichment System (FACE). Elevated CO{sub 2} increased diurnal assimilation by between 12% and 92%. Analysis of A/C{sub i} responses showed that effective nitrogen fertilisation was more important to rubisCO and RuBP activity than elevated CO{sub 2}. No acclimation was consistently observed. Leaves lower down the canopy were found to have lower Vc{sub max} and J{sub max} values, though age may be the cause of the latter effect. FACE conditions have only a small effect on these responses. There was some evidence found for the down-regulation of photosynthesis in the late afternoon. The FACE conditions had no affect on stomatal density but did increase epidermal cell density.

  8. Kresoxim-methyl primes Medicago truncatula plants against abiotic stress factors via altered reactive oxygen and nitrogen species signalling leading to downstream transcriptional and metabolic readjustment.

    PubMed

    Filippou, Panagiota; Antoniou, Chrystalla; Obata, Toshihiro; Van Der Kelen, Katrien; Harokopos, Vaggelis; Kanetis, Loukas; Aidinis, Vassilis; Van Breusegem, Frank; Fernie, Alisdair R; Fotopoulos, Vasileios

    2016-03-01

    Biotic and abiotic stresses, such as fungal infection and drought, cause major yield losses in modern agriculture. Kresoxim-methyl (KM) belongs to the strobilurins, one of the most important classes of agricultural fungicides displaying a direct effect on several plant physiological and developmental processes. However, the impact of KM treatment on salt and drought stress tolerance is unknown. In this study we demonstrate that KM pre-treatment of Medicago truncatula plants results in increased protection to drought and salt stress. Foliar application with KM prior to stress imposition resulted in improvement of physiological parameters compared with stressed-only plants. This protective effect was further supported by increased proline biosynthesis, modified reactive oxygen and nitrogen species signalling, and attenuation of cellular damage. In addition, comprehensive transcriptome analysis identified a number of transcripts that are differentially accumulating in drought- and salinity-stressed plants (646 and 57, respectively) after KM pre-treatment compared with stressed plants with no KM pre-treatment. Metabolomic analysis suggests that the priming role of KM in drought- and to a lesser extent in salinity-stressed plants can be attributed to the regulation of key metabolites (including sugars and amino acids) resulting in protection against abiotic stress factors. Overall, the present study highlights the potential use of this commonly used fungicide as a priming agent against key abiotic stress conditions. PMID:26712823

  9. Assessing quality of Medicago sativa silage by monitoring bacterial composition with single molecule, real-time sequencing technology and various physiological parameters.

    PubMed

    Bao, Weichen; Mi, Zhihui; Xu, Haiyan; Zheng, Yi; Kwok, Lai Yu; Zhang, Heping; Zhang, Wenyi

    2016-06-24

    The present study applied the PacBio single molecule, real-time sequencing technology (SMRT) in evaluating the quality of silage production. Specifically, we produced four types of Medicago sativa silages by using four different lactic acid bacteria-based additives (AD-I, AD-II, AD-III and AD-IV). We monitored the changes in pH, organic acids (including butyric acid, the ratio of acetic acid/lactic acid, γ-aminobutyric acid, 4-hyroxy benzoic acid and phenyl lactic acid), mycotoxins, and bacterial microbiota during silage fermentation. Our results showed that the use of the additives was beneficial to the silage fermentation by enhancing a general pH and mycotoxin reduction, while increasing the organic acids content. By SMRT analysis of the microbial composition in eight silage samples, we found that the bacterial species number and relative abundances shifted apparently after fermentation. Such changes were specific to the LAB species in the additives. Particularly, Bacillus megaterium was the initial dominant species in the raw materials; and after the fermentation process, Pediococcus acidilactici and Lactobacillus plantarum became the most prevalent species, both of which were intrinsically present in the LAB additives. Our data have demonstrated that the SMRT sequencing platform is applicable in assessing the quality of silage.

  10. An unusual abscisic acid and gibberellic acid synergism increases somatic embryogenesis, facilitates its genetic analysis and improves transformation in Medicago truncatula.

    PubMed

    Nolan, Kim E; Song, Youhong; Liao, Siyang; Saeed, Nasir A; Zhang, Xiyi; Rose, Ray J

    2014-01-01

    Somatic embryogenesis (SE) can be readily induced in leaf explants of the Jemalong 2HA genotype of the model legume Medicago truncatula by auxin and cytokinin, but rarely in wild-type Jemalong. Gibberellic acid (GA), a hormone not included in the medium, appears to act in Arabidopsis as a repressor of the embryonic state such that low ABA (abscisic acid): GA ratios will inhibit SE. It was important to evaluate the GA effect in M. truncatula in order to formulate generic SE mechanisms, given the Arabidopsis information. It was surprising to find that low ABA:GA ratios in M. truncatula acted synergistically to stimulate SE. The unusual synergism between GA and ABA in inducing SE has utility in improving SE for regeneration and transformation in M. truncatula. Expression of genes previously shown to be important in M. truncatula SE was not increased. In investigating genes previously studied in GA investigations of Arabidopsis SE, there was increased expression of GA2ox and decreased expression of PICKLE, a negative regulator of SE in Arabidopsis. We suggest that in M. truncatula there are different ABA:GA ratios required for down-regulating the PICKLE gene, a repressor of the embryonic state. In M. truncatula it is a low ABA:GA ratio while in Arabidopsis it is a high ABA:GA ratio. In different species the expression of key genes is probably related to differences in how the hormone networks optimise their expression. PMID:24937316

  11. Genome-Wide Identification, Evolutionary Analysis and Expression Profiles of LATERAL ORGAN BOUNDARIES DOMAIN Gene Family in Lotus japonicus and Medicago truncatula

    PubMed Central

    Yang, Tianquan; Fang, Genwang yue; He, Hua

    2016-01-01

    The LATERAL ORGAN BOUNDARIES DOMAIN (LBD) gene family has been well-studied in Arabidopsis and play crucial roles in the diverse growth and development processes including establishment and maintenance of boundary of developmental lateral organs. In this study we identified and characterized 38 LBD genes in Lotus japonicus (LjLBD) and 57 LBD genes in Medicago truncatula (MtLBD), both of which are model legume plants that have some specific development features absent in Arabidopsis. The phylogenetic relationships, their locations in the genome, genes structure and conserved motifs were examined. The results revealed that all LjLBD and MtLBD genes could be distinctly divided into two classes: Class I and II. The evolutionary analysis showed that Type I functional divergence with some significantly site-specific shifts may be the main force for the divergence between Class I and Class II. In addition, the expression patterns of LjLBD genes uncovered the diverse functions in plant development. Interestingly, we found that two LjLBD proteins that were highly expressed during compound leaf and pulvinus development, can interact via yeast two-hybrid assays. Taken together, our findings provide an evolutionary and genetic foundation in further understanding the molecular basis of LBD gene family in general, specifically in L. japonicus and M. truncatula. PMID:27560982

  12. Elevated CO2 decreases the response of the ethylene signaling pathway in Medicago truncatula and increases the abundance of the pea aphid.

    PubMed

    Guo, Huijuan; Sun, Yucheng; Li, Yuefei; Liu, Xianghui; Zhang, Wenhao; Ge, Feng

    2014-01-01

    The performance of herbivorous insects is greatly affected by plant nutritional quality and resistance, which are likely to be altered by rising concentrations of atmospheric CO2 . We previously reported that elevated CO2 enhanced biological nitrogen (N) fixation of Medicago truncatula, which could result in an increased supply of amino acids to the pea aphid (Acyrthosiphon pisum). The current study examined the N nutritional quality and aphid resistance of sickle, an ethylene-insensitive mutant of M. truncatula with supernodulation, and its wild-type control A17 under elevated CO2 in open-top field chambers. Regardless of CO2 concentration, growth and amino acid content were greater and aphid resistance was lower in sickle than in A17. Elevated CO2 up-regulated N assimilation and transamination-related enzymes activities and increased phloem amino acids in both genotypes. Furthermore, elevated CO2 down-regulated expression of 1-amino-cyclopropane-carboxylic acid (ACC), sickle gene (SKL) and ethylene response transcription factors (ERF) genes in the ethylene signaling pathway of A17 when infested by aphids and decreased resistance against aphids in terms of lower activities of superoxide dismutase (SOD), peroxidase (POD), and polyphenol oxidase (PPO). Our results suggest that elevated CO2 suppresses the ethylene signaling pathway in M. truncatula, which results in an increase in plant nutritional quality for aphids and a decrease in plant resistance against aphids.

  13. Kresoxim-methyl primes Medicago truncatula plants against abiotic stress factors via altered reactive oxygen and nitrogen species signalling leading to downstream transcriptional and metabolic readjustment

    PubMed Central

    Filippou, Panagiota; Antoniou, Chrystalla; Obata, Toshihiro; Van Der Kelen, Katrien; Harokopos, Vaggelis; Kanetis, Loukas; Aidinis, Vassilis; Van Breusegem, Frank; Fernie, Alisdair R; Fotopoulos, Vasileios

    2016-01-01

    Biotic and abiotic stresses, such as fungal infection and drought, cause major yield losses in modern agriculture. Kresoxim-methyl (KM) belongs to the strobilurins, one of the most important classes of agricultural fungicides displaying a direct effect on several plant physiological and developmental processes. However, the impact of KM treatment on salt and drought stress tolerance is unknown. In this study we demonstrate that KM pre-treatment of Medicago truncatula plants results in increased protection to drought and salt stress. Foliar application with KM prior to stress imposition resulted in improvement of physiological parameters compared with stressed-only plants. This protective effect was further supported by increased proline biosynthesis, modified reactive oxygen and nitrogen species signalling, and attenuation of cellular damage. In addition, comprehensive transcriptome analysis identified a number of transcripts that are differentially accumulating in drought- and salinity-stressed plants (646 and 57, respectively) after KM pre-treatment compared with stressed plants with no KM pre-treatment. Metabolomic analysis suggests that the priming role of KM in drought- and to a lesser extent in salinity-stressed plants can be attributed to the regulation of key metabolites (including sugars and amino acids) resulting in protection against abiotic stress factors. Overall, the present study highlights the potential use of this commonly used fungicide as a priming agent against key abiotic stress conditions. PMID:26712823

  14. A Novel Plant Leucine-Rich Repeat Receptor Kinase Regulates the Response of Medicago truncatula Roots to Salt Stress[W

    PubMed Central

    de Lorenzo, Laura; Merchan, Francisco; Laporte, Philippe; Thompson, Richard; Clarke, Jonathan; Sousa, Carolina; Crespi, Martín

    2009-01-01

    In plants, a diverse group of cell surface receptor-like protein kinases (RLKs) plays a fundamental role in sensing external signals to regulate gene expression. Roots explore the soil environment to optimize their growth via complex signaling cascades, mainly analyzed in Arabidopsis thaliana. However, legume roots have significant physiological differences, notably their capacity to establish symbiotic interactions. These major agricultural crops are affected by environmental stresses such as salinity. Here, we report the identification of a leucine-rich repeat RLK gene, Srlk, from the legume Medicago truncatula. Srlk is rapidly induced by salt stress in roots, and RNA interference (RNAi) assays specifically targeting Srlk yielded transgenic roots whose growth was less inhibited by the presence of salt in the medium. Promoter-β-glucuronidase fusions indicate that this gene is expressed in epidermal root tissues in response to salt stress. Two Srlk-TILLING mutants also failed to limit root growth in response to salt stress and accumulated fewer sodium ions than controls. Furthermore, early salt-regulated genes are downregulated in Srlk-RNAi roots and in the TILLING mutant lines when submitted to salt stress. We propose a role for Srlk in the regulation of the adaptation of M. truncatula roots to salt stress. PMID:19244136

  15. Analysis of Large Seeds from Three Different Medicago truncatula Ecotypes Reveals a Potential Role of Hormonal Balance in Final Size Determination of Legume Grains.

    PubMed

    Bandyopadhyay, Kaustav; Uluçay, Orhan; Şakiroğlu, Muhammet; Udvardi, Michael K; Verdier, Jerome

    2016-01-01

    Legume seeds are important as protein and oil source for human diet. Understanding how their final seed size is determined is crucial to improve crop yield. In this study, we analyzed seed development of three accessions of the model legume, Medicago truncatula, displaying contrasted seed size. By comparing two large seed accessions to the reference accession A17, we described mechanisms associated with large seed size determination and potential factors modulating the final seed size. We observed that early events during embryogenesis had a major impact on final seed size and a delayed heart stage embryo development resulted to large seeds. We also observed that the difference in seed growth rate was mainly due to a difference in embryo cell number, implicating a role of cell division rate. Large seed accessions could be explained by an extended period of cell division due to a longer embryogenesis phase. According to our observations and recent reports, we observed that auxin (IAA) and abscisic acid (ABA) ratio could be a key determinant of cell division regulation at the end of embryogenesis. Overall, our study highlights that timing of events occurring during early seed development play decisive role for final seed size determination. PMID:27618017

  16. Expression of an alfalfa (Medicago sativa L.) peroxidase gene in transgenic Arabidopsis thaliana enhances resistance to NaCl and H2O2.

    PubMed

    Teng, K; Xiao, G Z; Guo, W E; Yuan, J B; Li, J; Chao, Y H; Han, L B

    2016-01-01

    Peroxidases (PODs) are enzymes that play important roles in catalyzing the reduction of H2O2 and the oxidation of various substrates. They function in many different and important biological processes, such as defense mechanisms, immune responses, and pathogeny. The POD genes have been cloned and identified in many plants, but their function in alfalfa (Medicago sativa L.) is not known, to date. Based on the POD gene sequence (GenBank accession No. L36157.1), we cloned the POD gene in alfalfa, which was named MsPOD. MsPOD expression increased with increasing H2O2. The gene was expressed in all of the tissues, including the roots, stems, leaves, and flowers, particularly in stems and leaves under light/dark conditions. A subcellular analysis showed that MsPOD was localized outside the cells. Transgenic Arabidopsis with MsPOD exhibited increased resistance to H2O2 and NaCl. Moreover, POD activity in the transgenic plants was significantly higher than that in wild-type Arabidopsis. These results show that MsPOD plays an important role in resistance to H2O2 and NaCl. PMID:27323080

  17. Analysis of Large Seeds from Three Different Medicago truncatula Ecotypes Reveals a Potential Role of Hormonal Balance in Final Size Determination of Legume Grains

    PubMed Central

    Bandyopadhyay, Kaustav; Uluçay, Orhan; Şakiroğlu, Muhammet; Udvardi, Michael K.; Verdier, Jerome

    2016-01-01

    Legume seeds are important as protein and oil source for human diet. Understanding how their final seed size is determined is crucial to improve crop yield. In this study, we analyzed seed development of three accessions of the model legume, Medicago truncatula, displaying contrasted seed size. By comparing two large seed accessions to the reference accession A17, we described mechanisms associated with large seed size determination and potential factors modulating the final seed size. We observed that early events during embryogenesis had a major impact on final seed size and a delayed heart stage embryo development resulted to large seeds. We also observed that the difference in seed growth rate was mainly due to a difference in embryo cell number, implicating a role of cell division rate. Large seed accessions could be explained by an extended period of cell division due to a longer embryogenesis phase. According to our observations and recent reports, we observed that auxin (IAA) and abscisic acid (ABA) ratio could be a key determinant of cell division regulation at the end of embryogenesis. Overall, our study highlights that timing of events occurring during early seed development play decisive role for final seed size determination. PMID:27618017

  18. Overexpression of Medicago sativa TMT elevates the α-tocopherol content in Arabidopsis seeds, alfalfa leaves, and delays dark-induced leaf senescence.

    PubMed

    Jiang, Jishan; Jia, Huili; Feng, Guangyan; Wang, Zan; Li, Jun; Gao, Hongwen; Wang, Xuemin

    2016-08-01

    Alfalfa (Medicago sativa L.) is a major forage legume for livestock and a target for improving their dietary quality. Vitamin E is an essential vitamin that animals must obtain from their diet for proper growth and development. γ-tocopherol methyltransferase (γ-TMT), which catalyzes the conversion of δ- and γ-tocopherols (or tocotrienols) to β- and α-tocopherols (or tocotrienols), respectively, is the final enzyme involved in the vitamin E biosynthetic pathway. The overexpression of M. sativa L.'s γ-TMT (MsTMT) increased the α-tocopherol content 10-15 fold above that of wild type Arabidopsis seeds without altering the total content of vitamin E. Additionally, in response to osmotic stress, the biomass and the expression levels of several osmotic marker genes were significantly higher in the transgenic lines compared with wild type. Overexpression of MsTMT in alfalfa led to a modest, albeit significant, increase in α-tocopherol in leaves and was also responsible for a delayed leaf senescence phenotype. Additionally, the crude protein content was increased, while the acid and neutral detergent fiber contents were unchanged in these transgenic lines. Thus, increased α-tocopherol content occurred in transgenic alfalfa without compromising the nutritional qualities. The targeted metabolic engineering of vitamin E biosynthesis through MsTMT overexpression provides a promising approach to improve the α-tocopherol content of forage crops. PMID:27297993

  19. Both Free Indole-3-Acetic Acid and Photosynthetic Performance are Important Players in the Response of Medicago truncatula to Urea and Ammonium Nutrition Under Axenic Conditions

    PubMed Central

    Esteban, Raquel; Royo, Beatriz; Urarte, Estibaliz; Zamarreño, Ángel M.; Garcia-Mina, José M.; Moran, Jose F.

    2016-01-01

    We aimed to identify the early stress response and plant performance of Medicago truncatula growing in axenic medium with ammonium or urea as the sole source of nitrogen, with respect to nitrate-based nutrition. Biomass measurements, auxin content analyses, root system architecture (RSA) response analyses, and physiological parameters were determined. Both ammonium and ureic nutrition severely affected the RSA, resulting in changes in the main elongation rate, lateral root development, and insert position from the root base. The auxin content decreased in both urea- and ammonium-treated roots; however, only the ammonium-treated plants were affected at the shoot level. The analysis of chlorophyll a fluorescence transients showed that ammonium affected photosystem II, but urea did not impair photosynthetic activity. Superoxide dismutase isoenzymes in the plastids were moderately affected by urea and ammonium in the roots. Overall, our results showed that low N doses from different sources had no remarkable effects on M. truncatula, with the exception of the differential phenotypic root response. High doses of both ammonium and urea caused great changes in plant length, auxin contents and physiological measurements. Interesting correlations were found between the shoot auxin pool and both plant length and the “performance index” parameter, which is obtained from measurements of the kinetics of chlorophyll a fluorescence. Taken together, these data demonstrate that both the indole-3-acetic acid pool and performance index are important components of the response of M. truncatula under ammonium or urea as the sole N source. PMID:26909089

  20. Protocols for Obtaining Zygotic and Somatic Embryos for Studying the Regulation of Early Embryo Development in the Model Legume Medicago truncatula.

    PubMed

    Kurdyukov, Sergey; Song, Youhong; Tiew, Terence W-Y; Wang, Xin-Ding; Nolan, Kim E; Rose, Ray J

    2015-01-01

    Early embryogenesis starting from a single cell zygote goes through rapid cell division and morphogenesis, and is morphologically characterized by pre-globular, globular, heart, torpedo and cotyledon stages. This progressive development is under the tight regulation of a complex molecular network. Harvesting sufficient early embryos at a similar stage of development is essential for investigating the cellular and molecular regulation of early embryogenesis. This is not straightforward since early embryogenesis undergoes rapid morphogenesis in a short while e.g. 8 days for Medicago truncatula to reach the early cotyledon stage. Here, we address the issue by two approaches. The first one establishes a linkage between embryo development and pod morphology in helping indicate the stage of the zygotic embryo. This is particularly based on the number of pod spirals and development of the spines. An alternative way to complement the in vivo studies is via culturing leaf explants to produce somatic embryos. The medium includes an unusual hormone combination - an auxin (1-naphthaleneacetic acid), a cytokinin (6-benzylaminopurine), abscisic acid and gibberellic acid. The different stages can be discerned growing out of the callus without dissection. PMID:26131626

  1. Genome-Wide Identification, Evolutionary Analysis and Expression Profiles of LATERAL ORGAN BOUNDARIES DOMAIN Gene Family in Lotus japonicus and Medicago truncatula.

    PubMed

    Yang, Tianquan; Fang, Genwang Yue; He, Hua; Chen, Jianghua

    2016-01-01

    The LATERAL ORGAN BOUNDARIES DOMAIN (LBD) gene family has been well-studied in Arabidopsis and play crucial roles in the diverse growth and development processes including establishment and maintenance of boundary of developmental lateral organs. In this study we identified and characterized 38 LBD genes in Lotus japonicus (LjLBD) and 57 LBD genes in Medicago truncatula (MtLBD), both of which are model legume plants that have some specific development features absent in Arabidopsis. The phylogenetic relationships, their locations in the genome, genes structure and conserved motifs were examined. The results revealed that all LjLBD and MtLBD genes could be distinctly divided into two classes: Class I and II. The evolutionary analysis showed that Type I functional divergence with some significantly site-specific shifts may be the main force for the divergence between Class I and Class II. In addition, the expression patterns of LjLBD genes uncovered the diverse functions in plant development. Interestingly, we found that two LjLBD proteins that were highly expressed during compound leaf and pulvinus development, can interact via yeast two-hybrid assays. Taken together, our findings provide an evolutionary and genetic foundation in further understanding the molecular basis of LBD gene family in general, specifically in L. japonicus and M. truncatula. PMID:27560982

  2. Elevated CO2 decreases the response of the ethylene signaling pathway in Medicago truncatula and increases the abundance of the pea aphid.

    PubMed

    Guo, Huijuan; Sun, Yucheng; Li, Yuefei; Liu, Xianghui; Zhang, Wenhao; Ge, Feng

    2014-01-01

    The performance of herbivorous insects is greatly affected by plant nutritional quality and resistance, which are likely to be altered by rising concentrations of atmospheric CO2 . We previously reported that elevated CO2 enhanced biological nitrogen (N) fixation of Medicago truncatula, which could result in an increased supply of amino acids to the pea aphid (Acyrthosiphon pisum). The current study examined the N nutritional quality and aphid resistance of sickle, an ethylene-insensitive mutant of M. truncatula with supernodulation, and its wild-type control A17 under elevated CO2 in open-top field chambers. Regardless of CO2 concentration, growth and amino acid content were greater and aphid resistance was lower in sickle than in A17. Elevated CO2 up-regulated N assimilation and transamination-related enzymes activities and increased phloem amino acids in both genotypes. Furthermore, elevated CO2 down-regulated expression of 1-amino-cyclopropane-carboxylic acid (ACC), sickle gene (SKL) and ethylene response transcription factors (ERF) genes in the ethylene signaling pathway of A17 when infested by aphids and decreased resistance against aphids in terms of lower activities of superoxide dismutase (SOD), peroxidase (POD), and polyphenol oxidase (PPO). Our results suggest that elevated CO2 suppresses the ethylene signaling pathway in M. truncatula, which results in an increase in plant nutritional quality for aphids and a decrease in plant resistance against aphids. PMID:24015892

  3. Fixed-precision sequential sampling plans for estimating alfalfa caterpillar, Colias lesbia, egg density in alfalfa, Medicago sativa, fields in Córdoba, Argentina.

    PubMed

    Serra, Gerardo V; Porta, Norma C La; Avalos, Susana; Mazzuferi, Vilma

    2013-01-01

    The alfalfa caterpillar, Colias lesbia (Fabricius) (Lepidoptera: Pieridae), is a major pest of alfalfa, Medicago sativa L. (Fabales: Fabaceae), crops in Argentina. Its management is based mainly on chemical control of larvae whenever the larvae exceed the action threshold. To develop and validate fixed-precision sequential sampling plans, an intensive sampling programme for C. lesbia eggs was carried out in two alfalfa plots located in the Province of Córdoba, Argentina, from 1999 to 2002. Using Resampling for Validation of Sampling Plans software, 12 additional independent data sets were used to validate the sequential sampling plan with precision levels of 0.10 and 0.25 (SE/mean), respectively. For a range of mean densities of 0.10 to 8.35 eggs/sample, an average sample size of only 27 and 26 sample units was required to achieve a desired precision level of 0.25 for the sampling plans of Green and Kuno, respectively. As the precision level was increased to 0.10, average sample size increased to 161 and 157 sample units for the sampling plans of Green and Kuno, respectively. We recommend using Green's sequential sampling plan because it is less sensitive to changes in egg density. These sampling plans are a valuable tool for researchers to study population dynamics and to evaluate integrated pest management strategies.

  4. Effect of Medicago sativa L. and compost on organic and inorganic pollutant removal from a mixed contaminated soil and risk assessment using ecotoxicological tests.

    PubMed

    Marchand, Charlotte; Hogland, William; Kaczala, Fabio; Jani, Yahya; Marchand, Lilian; Augustsson, Anna; Hijri, Mohamed

    2016-11-01

    Several Gentle Remediation Options (GRO), e.g., plant-based options (phytoremediation), singly and combined with soil amendments, can be simultaneously efficient for degrading organic pollutants and either stabilizing or extracting trace elements (TEs). Here, a 5-month greenhouse trial was performed to test the efficiency of Medicago sativa L., singly and combined with a compost addition (30% w/w), to treat soils contaminated by petroleum hydrocarbons (PHC), Co and Pb collected at an auto scrap yard. After 5 months, total soil Pb significantly decreased in the compost-amended soil planted with M. sativa, but not total soil Co. Compost incorporation into the soil promoted PHC degradation, M. sativa growth and survival, and shoot Pb concentrations [3.8 mg kg(-1) dry weight (DW)]. Residual risk assessment after the phytoremediation trial showed a positive effect of compost amendment on plant growth and earthworm development. The O2 uptake by soil microorganisms was lower in the compost-amended soil, suggesting a decrease in microbial activity. This study underlined the benefits of the phytoremediation option based on M. sativa cultivation and compost amendment for remediating PHC- and Pb-contaminated soils. PMID:27216854

  5. Medicago truncatula Natural Resistance-Associated Macrophage Protein1 Is Required for Iron Uptake by Rhizobia-Infected Nodule Cells1[OPEN

    PubMed Central

    Tejada-Jiménez, Manuel; Castro-Rodríguez, Rosario; Kryvoruchko, Igor; Lucas, M. Mercedes; Udvardi, Michael; Imperial, Juan; González-Guerrero, Manuel

    2015-01-01

    Iron is critical for symbiotic nitrogen fixation (SNF) as a key component of multiple ferroproteins involved in this biological process. In the model legume Medicago truncatula, iron is delivered by the vasculature to the infection/maturation zone (zone II) of the nodule, where it is released to the apoplast. From there, plasma membrane iron transporters move it into rhizobia-containing cells, where iron is used as the cofactor of multiple plant and rhizobial proteins (e.g. plant leghemoglobin and bacterial nitrogenase). MtNramp1 (Medtr3g088460) is the M. truncatula Natural Resistance-Associated Macrophage Protein family member, with the highest expression levels in roots and nodules. Immunolocalization studies indicate that MtNramp1 is mainly targeted to the plasma membrane. A loss-of-function nramp1 mutant exhibited reduced growth compared with the wild type under symbiotic conditions, but not when fertilized with mineral nitrogen. Nitrogenase activity was low in the mutant, whereas exogenous iron and expression of wild-type MtNramp1 in mutant nodules increased nitrogen fixation to normal levels. These data are consistent with a model in which MtNramp1 is the main transporter responsible for apoplastic iron uptake by rhizobia-infected cells in zone II. PMID:25818701

  6. Loss of Abaxial Leaf Epicuticular Wax in Medicago truncatula irg1/palm1 Mutants Results in Reduced Spore Differentiation of Anthracnose and Nonhost Rust Pathogens[W

    PubMed Central

    Uppalapati, Srinivasa Rao; Ishiga, Yasuhiro; Doraiswamy, Vanthana; Bedair, Mohamed; Mittal, Shipra; Chen, Jianghua; Nakashima, Jin; Tang, Yuhong; Tadege, Million; Ratet, Pascal; Chen, Rujin; Schultheiss, Holger; Mysore, Kirankumar S.

    2012-01-01

    To identify genes that confer nonhost resistance to biotrophic fungal pathogens, we did a forward-genetics screen using Medicago truncatula Tnt1 retrotransposon insertion lines. From this screen, we identified an inhibitor of rust germ tube differentation1 (irg1) mutant that failed to promote preinfection structure differentiation of two rust pathogens, Phakopsora pachyrhizi and Puccinia emaculata, and one anthracnose pathogen, Colletotrichum trifolii, on the abaxial leaf surface. Cytological and chemical analyses revealed that the inhibition of rust preinfection structures in irg1 mutants is due to complete loss of the abaxial epicuticular wax crystals and reduced surface hydrophobicity. The composition of waxes on abaxial leaf surface of irg1 mutants had >90% reduction of C30 primary alcohols and a preferential increase of C29 and C31 alkanes compared with the wild type. IRG1 encodes a Cys(2)His(2) zinc finger transcription factor, PALM1, which also controls dissected leaf morphology in M. truncatula. Transcriptome analysis of irg1/palm1 mutants revealed downregulation of eceriferum4, an enzyme implicated in primary alcohol biosynthesis, and MYB96, a major transcription factor that regulates wax biosynthesis. Our results demonstrate that PALM1 plays a role in regulating epicuticular wax metabolism and transport and that epicuticular wax influences spore differentiation of host and nonhost fungal pathogens. PMID:22294617

  7. Dissection of Bacterial Wilt on Medicago truncatula Revealed Two Type III Secretion System Effectors Acting on Root Infection Process and Disease Development[C][W][OA

    PubMed Central

    Turner, Marie; Jauneau, Alain; Genin, Stéphane; Tavella, Marie-José; Vailleau, Fabienne; Gentzbittel, Laurent; Jardinaud, Marie-Françoise

    2009-01-01

    Ralstonia solanacearum is the causal agent of the devastating bacterial wilt disease, which colonizes susceptible Medicago truncatula via the intact root tip. Infection involves four steps: appearance of root tip symptoms, root tip cortical cell invasion, vessel colonization, and foliar wilting. We examined this pathosystem by in vitro inoculation of intact roots of susceptible or resistant M. truncatula with the pathogenic strain GMI1000. The infection process was type III secretion system dependent and required two type III effectors, Gala7 and AvrA, which were shown to be involved at different stages of infection. Both effectors were involved in development of root tip symptoms, and Gala7 was the main determinant for bacterial invasion of cortical cells. Vessel invasion depended on the host genetic background and was never observed in the resistant line. The invasion of the root tip vasculature in the susceptible line caused foliar wilting. The avrA mutant showed reduced aggressiveness in all steps of the infection process, suggesting a global role in R. solanacearum pathogenicity. The roles of these two effectors in subsequent stages were studied using an assay that bypassed the penetration step; with this assay, the avrA mutant showed no effect compared with the GMI1000 strain, indicating that AvrA is important in early stages of infection. However, later disease symptoms were reduced in the gala7 mutant, indicating a key role in later stages of infection. PMID:19493968

  8. Co-transforming bar and CsLEA enhanced tolerance to drought and salt stress in transgenic alfalfa (Medicago sativa L.).

    PubMed

    Zhang, Jiyu; Duan, Zhen; Zhang, Daiyu; Zhang, Jianquan; Di, Hongyan; Wu, Fan; Wang, Yanrong

    2016-03-25

    Drought and high salinity are two major abiotic factors that restrict alfalfa productivity. A dehydrin protein, CsLEA, from the desert grass Cleistogenes songorica was transformed into alfalfa (Medicago sativa L.) via Agrobacterium-mediated transformation using the bar gene as a selectable marker, and the drought and salt stress tolerances of the transgenic plants were assessed. Thirty-nine of 119 transformants were positive, as screened by Basta, and further molecularly authenticated using PCR and RT-PCR. Phenotype observations revealed that the transgenic plants grew better than the wild-type (WT) plants after 15d of drought stress and 10d of salt stress: the leaves of WT alfalfa turned yellow, whereas the transgenic alfalfa leaves only wilted; after rewatering, the transgenic plants returned to a normal state, though the WT plants could not be restored. Evaluation of physiologic and biochemical indices during drought and salt stresses showed a relatively lower Na(+) content in the leaves of the transgenic plants, which would reduce toxic ion effects. In addition, the transgenic plants were able to maintain a higher relative water content (RWC), higher shoot biomass, fewer photosystem changes, decreased membrane injury, and a lower level of osmotic stress injury. These results demonstrate that overexpression of the CsLEA gene can enhance the drought and salt tolerance of transgenic alfalfa; in addition, carrying the bar gene in the genome may increase herbicide resistance. PMID:26906624

  9. Abscisic acid-induced nitric oxide and proline accumulation in independent pathways under water-deficit stress during seedling establishment in Medicago truncatula.

    PubMed

    Planchet, Elisabeth; Verdu, Isabelle; Delahaie, Julien; Cukier, Caroline; Girard, Clément; Morère-Le Paven, Marie-Christine; Limami, Anis M

    2014-05-01

    Nitric oxide (NO) production and amino acid metabolism modulation, in particular abscisic acid (ABA)-dependent proline accumulation, are stimulated in planta by most abiotic stresses. However, the relationship between NO production and proline accumulation under abiotic stress is still poorly understood, especially in the early phases of plant development. To unravel this question, this work investigated the tight relationship between NO production and proline metabolism under water-deficit stress during seedling establishment. Endogenous nitrate reductase-dependent NO production in Medicago truncatula seedlings increased in a time-dependent manner after short-term water-deficit stress. This water-deficit-induced endogenous NO accumulation was mediated through a ABA-dependent pathway and accompanied by an inhibition of seed germination, a loss of water content, and a decrease in elongation of embryo axes. Interestingly, a treatment with a specific NO scavenger (cPTIO) alleviated these water-deficit detrimental effects. However, the content of total amino acids, in particular glutamate and proline, as well as the expression of genes encoding enzymes of synthesis and degradation of proline were not affected by cPTIO treatment under water-deficit stress. Under normal conditions, exogenous NO donor stimulated neither the expression of P5CS2 nor the proline content, as observed after PEG treatment. These results strongly suggest that the modulation of proline metabolism is independent of NO production under short-term water-deficit stress during seedling establishment.

  10. Suppression of Arbuscule Degeneration in Medicago truncatula phosphate transporter4 Mutants is Dependent on the Ammonium Transporter 2 Family Protein AMT2;3.

    PubMed

    Breuillin-Sessoms, Florence; Floss, Daniela S; Gomez, S Karen; Pumplin, Nathan; Ding, Yi; Levesque-Tremblay, Veronique; Noar, Roslyn D; Daniels, Dierdra A; Bravo, Armando; Eaglesham, James B; Benedito, Vagner A; Udvardi, Michael K; Harrison, Maria J

    2015-04-01

    During arbuscular mycorrhizal (AM) symbiosis, the plant gains access to phosphate (Pi) and nitrogen delivered by its fungal symbiont. Transfer of mineral nutrients occurs at the interface between branched hyphae called arbuscules and root cortical cells. In Medicago truncatula, a Pi transporter, PT4, is required for symbiotic Pi transport, and in pt4, symbiotic Pi transport fails, arbuscules degenerate prematurely, and the symbiosis is not maintained. Premature arbuscule degeneration (PAD) is suppressed when pt4 mutants are nitrogen-deprived, possibly the result of compensation by PT8, a second AM-induced Pi transporter. However, PAD is also suppressed in nitrogen-starved pt4 pt8 double mutants, negating this hypothesis and furthermore indicating that in this condition, neither of these symbiotic Pi transporters is required for symbiosis. In M. truncatula, three AMT2 family ammonium transporters are induced during AM symbiosis. To test the hypothesis that suppression of PAD involves AMT2 transporters, we analyzed double and triple Pi and ammonium transporter mutants. ATM2;3 but not AMT2;4 was required for suppression of PAD in pt4, while AMT2;4, but not AMT2;3, complemented growth of a yeast ammonium transporter mutant. In summary, arbuscule life span is influenced by PT4 and ATM2;3, and their relative importance varies with the nitrogen status of the plant.

  11. Fixed-Precision Sequential Sampling Plans for Estimating Alfalfa Caterpillar, Colias lesbia, Egg Density in Alfalfa, Medicago sativa, Fields in Córdoba, Argentina

    PubMed Central

    Serra, Gerardo V.; Porta, Norma C. La; Avalos, Susana; Mazzuferi, Vilma

    2013-01-01

    The alfalfa caterpillar, Colias lesbia (Fabricius) (Lepidoptera: Pieridae), is a major pest of alfalfa, Medicago sativa L. (Fabales: Fabaceae), crops in Argentina. Its management is based mainly on chemical control of larvae whenever the larvae exceed the action threshold. To develop and validate fixed-precision sequential sampling plans, an intensive sampling programme for C. lesbia eggs was carried out in two alfalfa plots located in the Province of Córdoba, Argentina, from 1999 to 2002. Using Resampling for Validation of Sampling Plans software, 12 additional independent data sets were used to validate the sequential sampling plan with precision levels of 0.10 and 0.25 (SE/mean), respectively. For a range of mean densities of 0.10 to 8.35 eggs/sample, an average sample size of only 27 and 26 sample units was required to achieve a desired precision level of 0.25 for the sampling plans of Green and Kuno, respectively. As the precision level was increased to 0.10, average sample size increased to 161 and 157 sample units for the sampling plans of Green and Kuno, respectively. We recommend using Green's sequential sampling plan because it is less sensitive to changes in egg density. These sampling plans are a valuable tool for researchers to study population dynamics and to evaluate integrated pest management strategies. PMID:23909840

  12. The symbiotic transcription factor MtEFD and cytokinins are positively acting in the Medicago truncatula and Ralstonia solanacearum pathogenic interaction.

    PubMed

    Moreau, Sandra; Fromentin, Justine; Vailleau, Fabienne; Vernié, Tatiana; Huguet, Stéphanie; Balzergue, Sandrine; Frugier, Florian; Gamas, Pascal; Jardinaud, Marie-Françoise

    2014-03-01

    • A plant-microbe dual biological system was set up involving the model legume Medicago truncatula and two bacteria, the soil-borne root pathogen Ralstonia solanacearum and the beneficial symbiont Sinorhizobium meliloti. • Comparison of transcriptomes under symbiotic and pathogenic conditions highlighted the transcription factor MtEFD (Ethylene response Factor required for nodule Differentiation) as being upregulated in both interactions, together with a set of cytokinin-related transcripts involved in metabolism, signaling and response. MtRR4 (Response Regulator), a cytokinin primary response gene negatively regulating cytokinin signaling and known as a target of MtEFD in nodulation processes, was retrieved in this set of transcripts. • Refined studies of MtEFD and MtRR4 expression during M. truncatula and R. solanacearum interaction indicated differential kinetics of induction and requirement of central regulators of bacterial pathogenicity, HrpG and HrpB. Similar to MtRR4, MtEFD upregulation during the pathogenic interaction was dependent on cytokinin perception mediated by the MtCRE1 (Cytokinin REsponse 1) receptor. • The use of M. truncatula efd-1 and cre1-1 mutants evidenced MtEFD and cytokinin perception as positive factors for bacterial wilt development. These factors therefore play an important role in both root nodulation and root disease development.

  13. Loss of the nodule-specific cysteine rich peptide, NCR169, abolishes symbiotic nitrogen fixation in the Medicago truncatula dnf7 mutant

    PubMed Central

    Horváth, Beatrix; Domonkos, Ágota; Szűcs, Attila; Ábrahám, Edit; Ayaydin, Ferhan; Bóka, Károly; Chen, Yuhui; Chen, Rujin; Murray, Jeremy D.; Udvardi, Michael K.; Kondorosi, Éva; Kaló, Péter

    2015-01-01

    Host compatible rhizobia induce the formation of legume root nodules, symbiotic organs within which intracellular bacteria are present in plant-derived membrane compartments termed symbiosomes. In Medicago truncatula nodules, the Sinorhizobium microsymbionts undergo an irreversible differentiation process leading to the development of elongated polyploid noncultivable nitrogen fixing bacteroids that convert atmospheric dinitrogen into ammonia. This terminal differentiation is directed by the host plant and involves hundreds of nodule specific cysteine-rich peptides (NCRs). Except for certain in vitro activities of cationic peptides, the functional roles of individual NCR peptides in planta are not known. In this study, we demonstrate that the inability of M. truncatula dnf7 mutants to fix nitrogen is due to inactivation of a single NCR peptide, NCR169. In the absence of NCR169, bacterial differentiation was impaired and was associated with early senescence of the symbiotic cells. Introduction of the NCR169 gene into the dnf7-2/NCR169 deletion mutant restored symbiotic nitrogen fixation. Replacement of any of the cysteine residues in the NCR169 peptide with serine rendered it incapable of complementation, demonstrating an absolute requirement for all cysteines in planta. NCR169 was induced in the cell layers in which bacteroid elongation was most pronounced, and high expression persisted throughout the nitrogen-fixing nodule zone. Our results provide evidence for an essential role of NCR169 in the differentiation and persistence of nitrogen fixing bacteroids in M. truncatula. PMID:26401023

  14. A Change in SHATTERPROOF Protein Lies at the Origin of a Fruit Morphological Novelty and a New Strategy for Seed Dispersal in Medicago Genus1[C][W

    PubMed Central

    Fourquin, Chloé; del Cerro, Carolina; Victoria, Filipe C.; Vialette-Guiraud, Aurélie; de Oliveira, Antonio C.; Ferrándiz, Cristina

    2013-01-01

    Angiosperms are the most diverse and numerous group of plants, and it is generally accepted that this evolutionary success owes in part to the diversity found in fruits, key for protecting the developing seeds and ensuring seed dispersal. Although studies on the molecular basis of morphological innovations are few, they all illustrate the central role played by transcription factors acting as developmental regulators. Here, we show that a small change in the protein sequence of a MADS-box transcription factor correlates with the origin of a highly modified fruit morphology and the change in seed dispersal strategies that occurred in Medicago, a genus belonging to the large legume family. This protein sequence modification alters the functional properties of the protein, affecting the affinities for other protein partners involved in high-order complexes. Our work illustrates that variation in coding regions can generate evolutionary novelties not based on gene duplication/subfunctionalization but by interactions in complex networks, contributing also to the current debate on the relative importance of changes in regulatory or coding regions of master regulators in generating morphological novelties. PMID:23640757

  15. Loss of the nodule-specific cysteine rich peptide, NCR169, abolishes symbiotic nitrogen fixation in the Medicago truncatula dnf7 mutant.

    PubMed

    Horváth, Beatrix; Domonkos, Ágota; Kereszt, Attila; Szűcs, Attila; Ábrahám, Edit; Ayaydin, Ferhan; Bóka, Károly; Chen, Yuhui; Chen, Rujin; Murray, Jeremy D; Udvardi, Michael K; Kondorosi, Éva; Kaló, Péter

    2015-12-01

    Host compatible rhizobia induce the formation of legume root nodules, symbiotic organs within which intracellular bacteria are present in plant-derived membrane compartments termed symbiosomes. In Medicago truncatula nodules, the Sinorhizobium microsymbionts undergo an irreversible differentiation process leading to the development of elongated polyploid noncultivable nitrogen fixing bacteroids that convert atmospheric dinitrogen into ammonia. This terminal differentiation is directed by the host plant and involves hundreds of nodule specific cysteine-rich peptides (NCRs). Except for certain in vitro activities of cationic peptides, the functional roles of individual NCR peptides in planta are not known. In this study, we demonstrate that the inability of M. truncatula dnf7 mutants to fix nitrogen is due to inactivation of a single NCR peptide, NCR169. In the absence of NCR169, bacterial differentiation was impaired and was associated with early senescence of the symbiotic cells. Introduction of the NCR169 gene into the dnf7-2/NCR169 deletion mutant restored symbiotic nitrogen fixation. Replacement of any of the cysteine residues in the NCR169 peptide with serine rendered it incapable of complementation, demonstrating an absolute requirement for all cysteines in planta. NCR169 was induced in the cell layers in which bacteroid elongation was most pronounced, and high expression persisted throughout the nitrogen-fixing nodule zone. Our results provide evidence for an essential role of NCR169 in the differentiation and persistence of nitrogen fixing bacteroids in M. truncatula.

  16. A two-dimensional electrophoresis proteomic reference map and systematic identification of 1367 proteins from a cell suspension culture of the model legume Medicago truncatula.

    PubMed

    Lei, Zhentian; Elmer, Aaron M; Watson, Bonnie S; Dixon, Richard A; Mendes, Pedro J; Sumner, Lloyd W

    2005-11-01

    The proteome of a Medicago truncatula cell suspension culture was analyzed using two-dimensional electrophoresis and nanoscale HPLC coupled to a tandem Q-TOF mass spectrometer (QSTAR Pulsar i) to yield an extensive protein reference map. Coomassie Brilliant Blue R-250 was used to visualize more than 1661 proteins, which were excised, subjected to in-gel trypsin digestion, and analyzed using nanoscale HPLC/MS/MS. The resulting spectral data were queried against a custom legume protein database using the MASCOT search engine. A total of 1367 of the 1661 proteins were identified with high rigor, yielding an identification success rate of 83% and 907 unique protein accession numbers. Functional annotation of the M. truncatula suspension cell proteins revealed a complete tricarboxylic acid cycle, a nearly complete glycolytic pathway, a significant portion of the ubiquitin pathway with the associated proteolytic and regulatory complexes, and many enzymes involved in secondary metabolism such as flavonoid/isoflavonoid, chalcone, and lignin biosynthesis. Proteins were also identified from most other functional classes including primary metabolism, energy production, disease/defense, protein destination/storage, protein synthesis, transcription, cell growth/division, and signal transduction. This work represents the most extensive proteomic description of M. truncatula suspension cells to date and provides a reference map for future comparative proteomic and functional genomic studies of the response of these cells to biotic and abiotic stress.

  17. Identification of Sources of Resistance to Phoma medicaginis Isolates in Medicago truncatula SARDI Core Collection Accessions, and Multigene Differentiation of Isolates.

    PubMed

    Ellwood, Simon R; Kamphuis, Lars G; Oliver, Richard P

    2006-12-01

    ABSTRACT Phoma medicaginis is a necrotrophic fungal pathogen, commonly found infecting the annual medic Medicago truncatula. To differentiate eight P. medicaginis isolates, five gene regions were examined: actin, beta-tubulin, calmodulin, translation elongation factor 1-alpha (EF-1alpha), and the internal transcribed spacer ribosomal DNA. Sequence comparisons showed that specimens isolated from M. truncatula in Western Australia formed a group that was consistently different from, but allied to, a P. medicaginis var. medicaginis type specimen. EF-1alpha contained a hyper-variable 55-bp repeat unit, which forms the basis of a rapid polymerase chain reaction-based method of reliably distinguishing isolates. Characterization of three isolates showed that all exhibited a narrow host range, causing disease only in M. sativa and M. truncatula among eight commonly cultivated legume species sampled. Infection of 86 M. truncatula single-seeded accessions showed a continuous distribution in disease phenotypes, with the majority of accessions susceptible. On a 1-to-5 disease reaction scale increasing in severity, individual fungal isolates showed means of 2.6 to 3.2, and scores ranged from 1 to 4.8 among accessions. The results presented here suggest that M. truncatula harbors specific and diverse sources of resistance to individual P. medicaginis genotypes. PMID:18943665

  18. Screening of saponins and sapogenins from Medicago species as potential PPARγ agonists and X-ray structure of the complex PPARγ/caulophyllogenin

    PubMed Central

    Montanari, Roberta; Capelli, Davide; Tava, Aldo; Galli, Andrea; Laghezza, Antonio; Tortorella, Paolo; Loiodice, Fulvio; Pochetti, Giorgio

    2016-01-01

    A series of saponins and sapogenins from Medicago species were tested for their ability to bind and activate the nuclear receptor PPARγ by SPR experiments and transactivation assay, respectively. The SPR analysis proved to be a very powerful and fast technique for screening a large number of compounds for their affinity to PPARγ and selecting the better candidates for further studies. Based on the obtained results, the sapogenin caulophyllogenin was proved to be a partial agonist towards PPARγ and the X-ray structure of its complex with PPARγ was also solved, in order to investigate the binding mode in the ligand binding domain of the nuclear receptor. This is the first known crystal structure of a sapogenin directly interacting with PPARγ. Another compound of the series, the echinocistic acid, showed antagonist activity towards PPARγ, a property that could be useful to inhibit the adipocyte differentiation which is a typical adverse effect of PPARγ agonists. This study confirms the interest on saponins and sapogenins as a valuable natural resource exploitable in the medical and food industry for ameliorating the metabolic syndrome. PMID:27283034

  19. The structure of Medicago truncatula δ1-pyrroline-5-carboxylate reductase provides new insights into regulation of proline biosynthesis in plants

    PubMed Central

    Ruszkowski, Milosz; Nocek, Boguslaw; Forlani, Giuseppe; Dauter, Zbigniew

    2015-01-01

    The two pathways for proline biosynthesis in higher plants share the last step, the conversion of δ1-pyrroline-5-carboxylate (P5C) to L-proline, which is catalyzed by P5C reductase (P5CR, EC 1.5.1.2) with the use of NAD(P)H as a coenzyme. There is increasing amount of evidence to suggest a complex regulation of P5CR activity at the post-translational level, yet the molecular basis of these mechanisms is unknown. Here we report the three-dimensional structure of the P5CR enzyme from the model legume Medicago truncatula (Mt). The crystal structures of unliganded MtP5CR decamer, and its complexes with the products NAD+, NADP+, and L-proline were refined using x-ray diffraction data (at 1.7, 1.85, 1.95, and 2.1 Å resolution, respectively). Based on the presented structural data, the coenzyme preference for NADPH over NADH was explained, and NADPH is suggested to be the only coenzyme used by MtP5CR in vivo. Furthermore, the insensitivity of MtP5CR to feed-back inhibition by proline, revealed by enzymatic analysis, was correlated with structural features. Additionally, a mechanism for the modulation of enzyme activity by chloride anions is discussed, as well as the rationale for the possible development of effective enzyme inhibitors. PMID:26579138

  20. Abscisic Acid and LATERAL ROOT ORGAN DEFECTIVE/NUMEROUS INFECTIONS AND POLYPHENOLICS Modulate Root Elongation via Reactive Oxygen Species in Medicago truncatula1[W][OPEN

    PubMed Central

    Zhang, Chang; Bousquet, Amanda; Harris, Jeanne M.

    2014-01-01

    Abscisic acid (ABA) modulates root growth in plants grown under normal and stress conditions and can rescue the root growth defects of the Medicago truncatula lateral root-organ defective (latd) mutant. Here, we demonstrate that reactive oxygen species (ROS) function downstream of ABA in the regulation of root growth by controlling cell elongation. We also show that the MtLATD/NUMEROUS INFECTIONS AND POLYPHENOLICS (NIP) nitrate transporter is required for ROS homeostasis and cell elongation in roots and that this balance is perturbed in latd mutants, leading to an excess of superoxide and hydrogen peroxide and a corresponding decrease in cell elongation. We found that expression of the superoxide-generating NADPH oxidase genes, MtRbohA and MtRbohC (for respiratory burst oxidase homologs), is increased in latd roots and that inhibition of NADPH oxidase activity pharmacologically can both reduce latd root ROS levels and increase cell length, implicating NADPH oxidase function in latd root growth defects. Finally, we demonstrate that ABA treatment alleviates ectopic ROS accumulation in latd roots, restores MtRbohC expression to wild-type levels, and promotes an increase in cell length. Reducing the expression of MtRbohC using RNA interference leads to increased root elongation in both wild-type and latd roots. These results reveal a mechanism by which the MtLATD/NIP nitrate transporter and ABA modulate root elongation via superoxide generation by the MtRbohC NADPH oxidase. PMID:25192698

  1. Pseudomonas fluorescens and Glomus mosseae trigger DMI3-dependent activation of genes related to a signal transduction pathway in roots of Medicago truncatula.

    PubMed

    Sanchez, Lisa; Weidmann, Stéphanie; Arnould, Christine; Bernard, Anne Rose; Gianinazzi, Silvio; Gianinazzi-Pearson, Vivienne

    2005-10-01

    Plant genes induced during early root colonization of Medicago truncatula Gaertn. J5 by a growth-promoting strain of Pseudomonas fluorescens (C7R12) have been identified by suppressive subtractive hybridization. Ten M. truncatula genes, coding proteins associated with a putative signal transduction pathway, showed an early and transient activation during initial interactions between M. truncatula and P. fluorescens, up to 8 d after root inoculation. Gene expression was not significantly enhanced, except for one gene, in P. fluorescens-inoculated roots of a Myc(-)Nod(-) genotype (TRV25) of M. truncatula mutated for the DMI3 (syn. MtSYM13) gene. This gene codes a Ca(2+) and calmodulin-dependent protein kinase, indicating a possible role of calcium in the cellular interactions between M. truncatula and P. fluorescens. When expression of the 10 plant genes was compared in early stages of root colonization by mycorrhizal and rhizobial microsymbionts, Glomus mosseae activated all 10 genes, whereas Sinorhizobium meliloti only activated one and inhibited four others. None of the genes responded to inoculation by either microsymbiont in roots of the TRV25 mutant. The similar response of the M. truncatula genes to P. fluorescens and G. mosseae points to common molecular pathways in the perception of the microbial signals by plant roots.

  2. Investigation of the Demographic and Selective Forces Shaping the Nucleotide Diversity of Genes Involved in Nod Factor Signaling in Medicago truncatula

    PubMed Central

    De Mita, Stéphane; Ronfort, Joëlle; McKhann, Heather I.; Poncet, Charles; El Malki, Redouane; Bataillon, Thomas

    2007-01-01

    Symbiotic nitrogen-fixing rhizobia are able to trigger root deformation in their Fabaceae host plants, allowing their intracellular accommodation. They do so by delivering molecules called Nod factors. We analyzed the patterns of nucleotide polymorphism of five genes controlling early Nod factor perception and signaling in the Fabaceae Medicago truncatula to understand the selective forces shaping the evolution of these genes. We used 30 M. truncatula genotypes sampled in a genetically homogeneous region of the species distribution range. We first sequenced 24 independent loci and detected a genomewide departure from the hypothesis of neutrality and demographic equilibrium that suggests a population expansion. These data were used to estimate parameters of a simple demographic model incorporating population expansion. The selective neutrality of genes controlling Nod factor perception was then examined using a combination of two complementary neutrality tests, Tajima's D and Fay and Wu's standardized H. The joint distribution of D and H expected under neutrality was obtained under the fitted population expansion model. Only the gene DMI1, which is expected to regulate the downstream signal, shows a pattern consistent with a putative selective event. In contrast, the receptor-encoding genes NFP and NORK show no significant signatures of selection. Among the genes that we analyzed, only DMI1 should be viewed as a candidate for adaptation in the recent history of M. truncatula. PMID:18073426

  3. A Medicago truncatula H+-pyrophosphatase gene, MtVP1, improves sucrose accumulation and anthocyanin biosynthesis in potato (Solanum tuberosum L.).

    PubMed

    Wang, J W; Wang, H Q; Xiang, W W; Chai, T Y

    2014-05-09

    We recently cloned MtVP1, a type I vacuolar-type H(+)-translocating inorganic pyrophosphatase from Medicago truncatula. In the present study, we investigated the cellular location and the function of this H(+)-PPase in Arabidopsis and potato (Solanum tuberosum L.). An MtVP1::enhanced green fluorescent protein fusion was constructed, which localized to the plasma membrane of onion epidermal cells. Transgenic Arabidopsis thaliana overexpressing MtVP1 had more robust root systems and redder shoots than wild-type (WT) plants under conditions of cold stress. Furthermore, overexpression of MtVP1 in potato accelerated the formation and growth of vegetative organs. The tuber buds and stem base of transgenic potatoes became redder than those of WT plants, but flowering was delayed by approximately half a month. Interestingly, anthocyanin biosynthesis was promoted in transgenic Arabidopsis seedlings and potato tuber buds. The sucrose concentration of transgenic potato tubers and tuber buds was enhanced compared with that of WT plants. Furthermore, sucrose concentration in tubers was higher than that in tuber buds. Although there was no direct evidence to support Fuglsang's hypothetical model regarding the effects of H(+)-PPase on sucrose phloem loading, we speculated that sucrose concentration was increased in tuber buds owing to the increased concentration in tubers. Therefore, overexpressed MtVP1 enhanced sucrose accumulation of source organs, which might enhance sucrose transport to sink organs, thus affecting anthocyanin biosynthesis.

  4. Co-expression of bacterial aspartate kinase and adenylylsulfate reductase genes substantially increases sulfur amino acid levels in transgenic alfalfa (Medicago sativa L.).

    PubMed

    Tong, Zongyong; Xie, Can; Ma, Lei; Liu, Liping; Jin, Yongsheng; Dong, Jiangli; Wang, Tao

    2014-01-01

    Alfalfa (Medicago sativa L.) is one of the most important forage crops used to feed livestock, such as cattle and sheep, and the sulfur amino acid (SAA) content of alfalfa is used as an index of its nutritional value. Aspartate kinase (AK) catalyzes the phosphorylation of aspartate to Asp-phosphate, the first step in the aspartate family biosynthesis pathway, and adenylylsulfate reductase (APR) catalyzes the conversion of activated sulfate to sulfite, providing reduced sulfur for the synthesis of cysteine, methionine, and other essential metabolites and secondary compounds. To reduce the feedback inhibition of other metabolites, we cloned bacterial AK and APR genes, modified AK, and introduced them into alfalfa. Compared to the wild-type alfalfa, the content of cysteine increased by 30% and that of methionine increased substantially by 60%. In addition, a substantial increase in the abundance of essential amino acids (EAAs), such as aspartate and lysine, was found. The results also indicated a close connection between amino acid metabolism and the tricarboxylic acid (TCA) cycle. The total amino acid content and the forage biomass tested showed no significant changes in the transgenic plants. This approach provides a new method for increasing SAAs and allows for the development of new genetically modified crops with enhanced nutritional value. PMID:24520364

  5. Involvement of peroxidase activity in developing somatic embryos of Medicago arborea L. Identification of an isozyme peroxidase as biochemical marker of somatic embryogenesis.

    PubMed

    Gallego, Piedad; Martin, Luisa; Blazquez, Antonio; Guerra, Hilario; Villalobos, Nieves

    2014-01-15

    The legume Medicago arborea L. is very interesting as regards the regeneration of marginal arid soils. The problem is that it does not have a good germinative yield. It was therefore decided to regenerate via somatic embryogenesis and find a marker of embryogenic potential. In this study, peroxidase activity was evaluated in non-embryogenic and embryogenic calli from M. arborea L. A decrease in soluble peroxidase activity is observed in its embryonic calli at the time at which the somatic embryos begin to appear. This activity is always lower in embryonic calli than in non-embryonic ones (unlike what happens in the case of wall-bound peroxidases). These results suggest that peroxidases can be considered to be enzymes involved in somatic embryogenesis in M. arborea. In addition, isozyme analyses were carried out on protein extracts using polyacrylamide gel electrophoresis. The band called P5 was detected only in embryogenic cultures at very early stages of development. This band was digested with trypsin and analyzed using linear ion trap (LTQ) mass spectrometer. In P5 isoform a peroxidase-L-ascorbate peroxidase was identified. It can be used as a marker that allows the identification of embryological potential.

  6. Differential effects of AM fungal isolates on Medicago truncatula growth and metal uptake in a multimetallic (Cd, Zn, Pb) contaminated agricultural soil.

    PubMed

    Redon, Paul-Olivier; Béguiristain, Thierry; Leyval, Corinne

    2009-03-01

    Toxic metal accumulation in soils of agricultural interest is a serious problem needing more attention, and investigations on soil-plant metal transfer must be pursued to better understand the processes involved in metal uptake. Arbuscular mycorrhizal (AM) fungi are known to influence metal transfer in plants by increasing plant biomass and reducing metal toxicity to plants even if diverging results were reported. The effects of five AM fungi isolated from metal contaminated or non-contaminated soils on metal (Cd, Zn) uptake by plant and transfer to leachates was assessed with Medicago truncatula grown in a multimetallic contaminated agricultural soil. Fungi isolated from metal-contaminated soils were more effective to reduce shoot Cd concentration. Metal uptake capacity differed between AM fungi and depended on the origin of the isolate. Not only fungal tolerance and ability to reduce metal concentrations in plant but also interactions with rhizobacteria affected heavy metal transfer and plant growth. Indeed, thanks to association with nodulating rhizobacteria, one Glomus intraradices inoculum increased particularly plant biomass which allowed exporting twofold more Cd and Zn in shoots as compared to non-mycorrhizal treatment. Cd concentrations in leachates were variable among fungal treatments, but can be significantly influenced by AM inoculation. The differential strategies of AM fungal colonisation in metal stress conditions are also discussed. PMID:19169716

  7. Assessing quality of Medicago sativa silage by monitoring bacterial composition with single molecule, real-time sequencing technology and various physiological parameters

    PubMed Central

    Bao, Weichen; Mi, Zhihui; Xu, Haiyan; Zheng, Yi; Kwok, Lai Yu; Zhang, Heping; Zhang, Wenyi

    2016-01-01

    The present study applied the PacBio single molecule, real-time sequencing technology (SMRT) in evaluating the quality of silage production. Specifically, we produced four types of Medicago sativa silages by using four different lactic acid bacteria-based additives (AD-I, AD-II, AD-III and AD-IV). We monitored the changes in pH, organic acids (including butyric acid, the ratio of acetic acid/lactic acid, γ-aminobutyric acid, 4-hyroxy benzoic acid and phenyl lactic acid), mycotoxins, and bacterial microbiota during silage fermentation. Our results showed that the use of the additives was beneficial to the silage fermentation by enhancing a general pH and mycotoxin reduction, while increasing the organic acids content. By SMRT analysis of the microbial composition in eight silage samples, we found that the bacterial species number and relative abundances shifted apparently after fermentation. Such changes were specific to the LAB species in the additives. Particularly, Bacillus megaterium was the initial dominant species in the raw materials; and after the fermentation process, Pediococcus acidilactici and Lactobacillus plantarum became the most prevalent species, both of which were intrinsically present in the LAB additives. Our data have demonstrated that the SMRT sequencing platform is applicable in assessing the quality of silage. PMID:27340760

  8. The Small GTPase ROP10 of Medicago truncatula Is Required for Both Tip Growth of Root Hairs and Nod Factor-Induced Root Hair Deformation

    PubMed Central

    Lei, Ming-Juan; Wang, Qi; Li, Xiaolin; Chen, Aimin; Luo, Li; Xie, Yajun; Li, Guan; Luo, Da; Mysore, Kirankumar S.; Wen, Jiangqi; Xie, Zhi-Ping; Staehelin, Christian; Wang, Yan-Zhang

    2015-01-01

    Rhizobia preferentially enter legume root hairs via infection threads, after which root hairs undergo tip swelling, branching, and curling. However, the mechanisms underlying such root hair deformation are poorly understood. Here, we showed that a type II small GTPase, ROP10, of Medicago truncatula is localized at the plasma membrane (PM) of root hair tips to regulate root hair tip growth. Overexpression of ROP10 and a constitutively active mutant (ROP10CA) generated depolarized growth of root hairs, whereas a dominant negative mutant (ROP10DN) inhibited root hair elongation. Inoculated with Sinorhizobium meliloti, the depolarized swollen and ballooning root hairs exhibited extensive root hair deformation and aberrant infection symptoms. Upon treatment with rhizobia-secreted nodulation factors (NFs), ROP10 was transiently upregulated in root hairs, and ROP10 fused to green fluorescent protein was ectopically localized at the PM of NF-induced outgrowths and curls around rhizobia. ROP10 interacted with the kinase domain of the NF receptor NFP in a GTP-dependent manner. Moreover, NF-induced expression of the early nodulin gene ENOD11 was enhanced by the overexpression of ROP10 and ROP10CA. These data suggest that NFs spatiotemporally regulate ROP10 localization and activity at the PM of root hair tips and that interactions between ROP10 and NF receptors are required for root hair deformation and continuous curling during rhizobial infection. PMID:25794934

  9. A Medicago truncatula H+-pyrophosphatase gene, MtVP1, improves sucrose accumulation and anthocyanin biosynthesis in potato (Solanum tuberosum L.).

    PubMed

    Wang, J W; Wang, H Q; Xiang, W W; Chai, T Y

    2014-01-01

    We recently cloned MtVP1, a type I vacuolar-type H(+)-translocating inorganic pyrophosphatase from Medicago truncatula. In the present study, we investigated the cellular location and the function of this H(+)-PPase in Arabidopsis and potato (Solanum tuberosum L.). An MtVP1::enhanced green fluorescent protein fusion was constructed, which localized to the plasma membrane of onion epidermal cells. Transgenic Arabidopsis thaliana overexpressing MtVP1 had more robust root systems and redder shoots than wild-type (WT) plants under conditions of cold stress. Furthermore, overexpression of MtVP1 in potato accelerated the formation and growth of vegetative organs. The tuber buds and stem base of transgenic potatoes became redder than those of WT plants, but flowering was delayed by approximately half a month. Interestingly, anthocyanin biosynthesis was promoted in transgenic Arabidopsis seedlings and potato tuber buds. The sucrose concentration of transgenic potato tubers and tuber buds was enhanced compared with that of WT plants. Furthermore, sucrose concentration in tubers was higher than that in tuber buds. Although there was no direct evidence to support Fuglsang's hypothetical model regarding the effects of H(+)-PPase on sucrose phloem loading, we speculated that sucrose concentration was increased in tuber buds owing to the increased concentration in tubers. Therefore, overexpressed MtVP1 enhanced sucrose accumulation of source organs, which might enhance sucrose transport to sink organs, thus affecting anthocyanin biosynthesis. PMID:24854441

  10. The 3' untranslated region of the two cytosolic glutamine synthetase (GS(1)) genes in alfalfa (Medicago sativa) regulates transcript stability in response to glutamine.

    PubMed

    Simon, Bindu; Sengupta-Gopalan, Champa

    2010-10-01

    Glutamine synthetase (GS) catalyzes the ATP-dependent condensation of ammonia with glutamate to produce glutamine. The GS enzyme is located either in the chloroplast (GS(2)) or in the cytoplasm (GS(1)). GS(1) is encoded by a small gene family and the members exhibit differential expression pattern mostly attributed to transcriptional regulation. Based on our recent finding that a soybean GS(1) gene, Gmglnβ ( 1 ) is subject to its 3'UTR-mediated post-transcriptional regulation as a transgene in alfalfa (Medicago sativa) we have raised the question of whether the 3'UTR-mediated transcript destabilization is a more universal phenomenon. Gene constructs consisting of the CaMV35S promoter driving the reporter gene, GUS, followed by the 3'UTRs of the two alfalfa GS(1) genes, MsGSa and MsGSb, were introduced into alfalfa and tobacco. The analysis of these transformants suggests that while both the 3'UTRs promote transcript turnover, the MsGSb 3'UTR is more effective than the MsGSa 3'UTR. However, both the 3'UTRs along with Gmglnβ ( 1 ) 3'UTR respond to nitrate as a trigger in transcript turnover. More detailed analysis points to glutamine rather than nitrate as the mediator of transcript turnover. Our data suggests that the 3'UTR-mediated regulation of GS(1) genes at the level of transcript turnover is probably universal and is used for fine-tuning the expression in keeping with the availability of the substrates.

  11. Does phloem-based resistance to aphid feeding affect host-plant acceptance for reproduction? Parturition of the pea aphid, Acyrthosiphon pisum, on two near-isogenic lines of Medicago truncatula.

    PubMed

    Nam, K Jung; Powell, G; Hardie, J

    2013-12-01

    Probing behaviour (prior to parturition) and parturition of two clones (PS01 and N116) of the pea aphid, Acyrthosiphon pisum on two genotypes (near-isogenic lines (NILs)) (Q174_5.13 and Q174_9.10) of Medicago truncatula were investigated using electrical penetration graph (EPG) coupled with simultaneous visual monitoring for parturition. Line Q174_5.13 has been reported to show a phloem-based resistance to feeding in the clone PS01 but to be susceptible to the clone N116, whereas Q174_9.10 has shown to be susceptible to both aphid clones. The time taken to first parturition by clone PS01 was similar on Q174_5.13 and Q174_9.10. Prior to parturition, no aphids on Q174_5.13 contacted phloem, but 5% of the aphids on Q174_9.10 showed phloem salivation (recognized by EPG pattern E1). No phloem contact was observed with aphid clone N116 on either NILs of Medicago before first parturition occurred, and the time taken to first larviposition was similar on Q174_5.13 and Q174_9.10. The results indicate that the initiation of parturition of the clone PS01 and N116 on both NILs does not require the phloem contact and seems unchanged by a phloem-based resistance mechanism to feeding on Medicago. This finding suggests that host recognition and decisions about parturition occur before phloem contact or ingestion, and act independently on R-gene-mediated resistance.

  12. Molecular cloning and characterization of the MsHSP17.7 gene from Medicago sativa L.

    PubMed

    Li, Zhen-Yi; Long, Rui-Cai; Zhang, Tie-Jun; Yang, Qing-Chuan; Kang, Jun-Mei

    2016-08-01

    Heat shock proteins (HSPs) are ubiquitous protective proteins that play crucial roles in plant development and adaptation to stress, and the aim of this study is to characterize the HSP gene in alfalfa. Here we isolated a small heat shock protein gene (MsHSP17.7) from alfalfa by homology-based cloning. MsHSP17.7 contains a 477-bp open reading frame and encodes a protein of 17.70-kDa. The amino acid sequence shares high identity with MtHSP (93.98 %), PsHSP17.1 (83.13 %), GmHSP17.9 (74.10 %) and SlHSP17.6 (79.25 %). Phylogenetic analysis revealed that MsHSP17.7 belongs to the group of cytosolic class II small heat shock proteins (sHSP), and likely localizes to the cytoplasm. Quantitative RT-PCR indicated that MsHSP17.7 was induced by heat shock, high salinity, peroxide and drought stress. Prokaryotic expression indicated that the salt and peroxide tolerance of Escherichia coli was remarkably enhanced. Transgenic Arabidopsis plants overexpressing MsHSP17.7 exhibited increased root length of transgenic Arabidopsis lines under salt stress compared to the wild-type line. The malondialdehyde (MDA) levels in the transgenic lines were significantly lower than in wild-type, although proline levels were similar between transgenic and wild-type lines. MsHSP17.7 was induced by heat shock, high salinity, oxidative stress and drought stress. Overexpression analysis suggests that MsHSP17.7 might play a key role in response to high salinity stress. PMID:27193169

  13. An RNA Sequencing Transcriptome Analysis Reveals Novel Insights into Molecular Aspects of the Nitrate Impact on the Nodule Activity of Medicago truncatula1[W

    PubMed Central

    Cabeza, Ricardo; Koester, Beke; Liese, Rebecca; Lingner, Annika; Baumgarten, Vanessa; Dirks, Jan; Salinas-Riester, Gabriela; Pommerenke, Claudia; Dittert, Klaus; Schulze, Joachim

    2014-01-01

    The mechanism through which nitrate reduces the activity of legume nodules is controversial. The objective of the study was to follow Medicago truncatula nodule activity after nitrate provision continuously and to identify molecular mechanisms, which down-regulate the activity of the nodules. Nodule H2 evolution started to decline after about 4 h of nitrate application. At that point in time, a strong shift in nodule gene expression (RNA sequencing) had occurred (1,120 differentially expressed genes). The most pronounced effect was the down-regulation of 127 genes for nodule-specific cysteine-rich peptides. Various other nodulins were also strongly down-regulated, in particular all the genes for leghemoglobins. In addition, shifts in the expression of genes involved in cellular iron allocation and mitochondrial ATP synthesis were observed. Furthermore, the expression of numerous genes for the formation of proteins and glycoproteins with no obvious function in nodules (e.g. germins, patatin, and thaumatin) was strongly increased. This occurred in conjunction with an up-regulation of genes for proteinase inhibitors, in particular those containing the Kunitz domain. The additionally formed proteins might possibly be involved in reducing nodule oxygen permeability. Between 4 and 28 h of nitrate exposure, a further reduction in nodule activity occurred, and the number of differentially expressed genes almost tripled. In particular, there was a differential expression of genes connected with emerging senescence. It is concluded that nitrate exerts rapid and manifold effects on nitrogenase activity. A certain degree of nitrate tolerance might be achieved when the down-regulatory effect on late nodulins can be alleviated. PMID:24285852

  14. A single amino acid substitution in a chitinase of the legume Medicago truncatula is sufficient to gain Nod-factor hydrolase activity.

    PubMed

    Zhang, Lan-Yue; Cai, Jie; Li, Ru-Jie; Liu, Wei; Wagner, Christian; Wong, Kam-Bo; Xie, Zhi-Ping; Staehelin, Christian

    2016-07-01

    The symbiotic interaction between nitrogen-fixing rhizobia and legumes depends on lipo-chitooligosaccharidic Nod-factors (NFs). The NF hydrolase MtNFH1 of Medicago truncatula is a symbiotic enzyme that hydrolytically inactivates NFs with a C16 : 2 acyl chain produced by the microsymbiont Sinorhizobium meliloti 1021. MtNFH1 is related to class V chitinases (glycoside hydrolase family 18) but lacks chitinase activity. Here, we investigated the substrate specificity of MtNFH1-related proteins. MtCHIT5a and MtCHIT5b of M. truncatula as well as LjCHIT5 of Lotus japonicus showed chitinase activity, suggesting a role in plant defence. The enzymes failed to hydrolyse NFs from S. meliloti. NFs from Rhizobium leguminosarum with a C18 : 4 acyl moiety were neither hydrolysed by these chitinases nor by MtNFH1. Construction of chimeric proteins and further amino acid replacements in MtCHIT5b were performed to identify chitinase variants that gained the ability to hydrolyse NFs. A single serine-to-proline substitution was sufficient to convert MtCHIT5b into an NF-cleaving enzyme. MtNFH1 with the corresponding proline-to-serine substitution failed to hydrolyse NFs. These results are in agreement with a substrate-enzyme model that predicts NF cleavage when the C16 : 2 moiety is placed into a distinct fatty acid-binding cleft. Our findings support the view that MtNFH1 evolved from the ancestral MtCHIT5b by gene duplication and subsequent symbiosis-related neofunctionalization. PMID:27383628

  15. Transport and partitioning of CO/sub 2/ fixed by root nodules of ureide and amide producing legumes. [Vigna angularis; Glycine max; Medicago sativa

    SciTech Connect

    Vance, C.P.; Boylan, K.L.M.; Maxwell, C.A.; Heichel, G.H.; Hardman, L.L.

    1985-01-01

    Nodulated and denodulated roots of adzuki bean (Vigna angularis), soybean (Glycine max), and alfalfa (Medicago sativa) were exposed to /sup 14/CO/sub 2/ to investigate the contribution of nodule CO/sub 2/ fixation to assimilation and transport of fixed nitrogen. The distribution of radioactivity in xylem sap and partitioning of carbon fixed by nodules to the whole plant were measured. Radioactivity in the xylem sap of nodulated soybean and adzuki bean was located primarily (70 to 87%) in the acid fraction while the basic (amino acid) fraction contained 10 to 22%. In contrast radioactivity in the xylem sap of nodulated alfalfa was primarily in amino acids with about 20% in organic acids. Total ureide concentration was 8.1, 4.7, and 0.0 micromoles per milliliter xylem sap for soybean, adzuki bean, and alfalfa, respectively. While the major nitrogen transport products in soybeans and adzuki beans are ureides, this class of metabolites contained less than 20% of the the total radioactivity. When nodules of plants were removed, radioactivity in xylem sap decreased by 90% or more. Pulse-chase experiments indicated that CO/sub 2/ fixed by nodules was rapidly transported to shoots and incorporated into acid stable constituents. The data are consistent with a role for nodule CO/sub 2/ fixation providing carbon for the assimilation and transport of fixed nitrogen in amide-based legumes. In contrast, CO/sub 2/ fixation by nodules of ureide transporting legumes appears to contribute little to assimilation and transport of fixed nitrogen. 19 references, 2 figures, 5 tables.

  16. A Regulatory Network-Based Approach Dissects Late Maturation Processes Related to the Acquisition of Desiccation Tolerance and Longevity of Medicago truncatula Seeds1[C][W][OPEN

    PubMed Central

    Verdier, Jerome; Lalanne, David; Pelletier, Sandra; Torres-Jerez, Ivone; Righetti, Karima; Bandyopadhyay, Kaustav; Leprince, Olivier; Chatelain, Emilie; Vu, Benoit Ly; Gouzy, Jerome; Gamas, Pascal; Udvardi, Michael K.; Buitink, Julia

    2013-01-01

    In seeds, desiccation tolerance (DT) and the ability to survive the dry state for prolonged periods of time (longevity) are two essential traits for seed quality that are consecutively acquired during maturation. Using transcriptomic and metabolomic profiling together with a conditional-dependent network of global transcription interactions, we dissected the maturation events from the end of seed filling to final maturation drying during the last 3 weeks of seed development in Medicago truncatula. The network revealed distinct coexpression modules related to the acquisition of DT, longevity, and pod abscission. The acquisition of DT and dormancy module was associated with abiotic stress response genes, including late embryogenesis abundant (LEA) genes. The longevity module was enriched in genes involved in RNA processing and translation. Concomitantly, LEA polypeptides accumulated, displaying an 18-d delayed accumulation compared with transcripts. During maturation, gulose and stachyose levels increased and correlated with longevity. A seed-specific network identified known and putative transcriptional regulators of DT, including ABSCISIC ACID-INSENSITIVE3 (MtABI3), MtABI4, MtABI5, and APETALA2/ ETHYLENE RESPONSE ELEMENT BINDING PROTEIN (AtAP2/EREBP) transcription factor as major hubs. These transcriptional activators were highly connected to LEA genes. Longevity genes were highly connected to two MtAP2/EREBP and two basic leucine zipper transcription factors. A heat shock factor was found at the transition of DT and longevity modules, connecting to both gene sets. Gain- and loss-of-function approaches of MtABI3 confirmed 80% of its predicted targets, thereby experimentally validating the network. This study captures the coordinated regulation of seed maturation and identifies distinct regulatory networks underlying the preparation for the dry and quiescent states. PMID:23929721

  17. Haem oxygenase-1 is involved in salicylic acid-induced alleviation of oxidative stress due to cadmium stress in Medicago sativa

    PubMed Central

    Shen, Wenbiao

    2012-01-01

    This work examines the involvement of haem oxygenase-1 (HO-1) in salicylic acid (SA)-induced alleviation of oxidative stress as a result of cadmium (Cd) stress in alfalfa (Medicago sativa L.) seedling roots. CdCl2 exposure caused severe growth inhibition and Cd accumulation, which were potentiated by pre-treatment with zinc protoporphyrin (ZnPPIX), a potent HO-1 inhibitor. Pre-treatment of plants with the HO-1 inducer haemin or SA, both of which could induce MsHO1 gene expression, significantly reduced the inhibition of growth and Cd accumulation. The alleviation effects were also evidenced by a decreased content of thiobarbituric acid-reactive substances (TBARS). The antioxidant behaviour was confirmed by histochemical staining for the detection of lipid peroxidation and the loss of plasma membrane integrity. Furthermore, haemin and SA pre-treatment modulated the activities of ascorbate peroxidase (APX), superoxide dismutase (SOD), and guaiacol peroxidase (POD), or their corresponding transcripts. Significant enhancement of the ratios of reduced/oxidized homoglutathione (hGSH), ascorbic acid (ASA)/dehydroascorbate (DHA), and NAD(P)H/NAD(P)+, and expression of their metabolism genes was observed, consistent with a decreased reactive oxygen species (ROS) distribution in the root tips. These effects are specific for HO-1, since ZnPPIX blocked the above actions, and the aggravated effects triggered by SA plus ZnPPIX were differentially reversed when carbon monoxide (CO) or bilirubin (BR), two catalytic by-products of HO-1, was added. Together, the results suggest that HO-1 is involved in the SA-induced alleviation of Cd-triggered oxidative stress by re-establishing redox homeostasis. PMID:22915740

  18. Deep Sequencing of the Medicago truncatula Root Transcriptome Reveals a Massive and Early Interaction between Nodulation Factor and Ethylene Signals1[OPEN

    PubMed Central

    Larrainzar, Estíbaliz; Riely, Brendan K.; Kim, Sang Cheol; Carrasquilla-Garcia, Noelia; Yu, Hee-Ju; Hwang, Hyun-Ju; Oh, Mijin; Kim, Goon Bo; Surendrarao, Anandkumar K.; Chasman, Deborah; Siahpirani, Alireza F.; Penmetsa, Ramachandra V.; Lee, Gang-Seob; Kim, Namshin; Roy, Sushmita; Mun, Jeong-Hwan; Cook, Douglas R.

    2015-01-01

    The legume-rhizobium symbiosis is initiated through the activation of the Nodulation (Nod) factor-signaling cascade, leading to a rapid reprogramming of host cell developmental pathways. In this work, we combine transcriptome sequencing with molecular genetics and network analysis to quantify and categorize the transcriptional changes occurring in roots of Medicago truncatula from minutes to days after inoculation with Sinorhizobium medicae. To identify the nature of the inductive and regulatory cues, we employed mutants with absent or decreased Nod factor sensitivities (i.e. Nodulation factor perception and Lysine motif domain-containing receptor-like kinase3, respectively) and an ethylene (ET)-insensitive, Nod factor-hypersensitive mutant (sickle). This unique data set encompasses nine time points, allowing observation of the symbiotic regulation of diverse biological processes with high temporal resolution. Among the many outputs of the study is the early Nod factor-induced, ET-regulated expression of ET signaling and biosynthesis genes. Coupled with the observation of massive transcriptional derepression in the ET-insensitive background, these results suggest that Nod factor signaling activates ET production to attenuate its own signal. Promoter:β-glucuronidase fusions report ET biosynthesis both in root hairs responding to rhizobium as well as in meristematic tissue during nodule organogenesis and growth, indicating that ET signaling functions at multiple developmental stages during symbiosis. In addition, we identified thousands of novel candidate genes undergoing Nod factor-dependent, ET-regulated expression. We leveraged the power of this large data set to model Nod factor- and ET-regulated signaling networks using MERLIN, a regulatory network inference algorithm. These analyses predict key nodes regulating the biological process impacted by Nod factor perception. We have made these results available to the research community through a searchable online

  19. Single-plant, Sterile Microcosms for Nodulation and Growth of the Legume Plant Medicago truncatula with the Rhizobial Symbiont Sinorhizobium meliloti

    PubMed Central

    Jones, Kathryn M.

    2013-01-01

    Rhizobial bacteria form symbiotic, nitrogen-fixing nodules on the roots of compatible host legume plants. One of the most well-developed model systems for studying these interactions is the plant Medicago truncatula cv. Jemalong A17 and the rhizobial bacterium Sinorhizobium meliloti 1021. Repeated imaging of plant roots and scoring of symbiotic phenotypes requires methods that are non-destructive to either plants or bacteria. The symbiotic phenotypes of some plant and bacterial mutants become apparent after relatively short periods of growth, and do not require long-term observation of the host/symbiont interaction. However, subtle differences in symbiotic efficiency and nodule senescence phenotypes that are not apparent in the early stages of the nodulation process require relatively long growth periods before they can be scored. Several methods have been developed for long-term growth and observation of this host/symbiont pair. However, many of these methods require repeated watering, which increases the possibility of contamination by other microbes. Other methods require a relatively large space for growth of large numbers of plants. The method described here, symbiotic growth of M. truncatula/S. meliloti in sterile, single-plant microcosms, has several advantages. Plants in these microcosms have sufficient moisture and nutrients to ensure that watering is not required for up to 9 weeks, preventing cross-contamination during watering. This allows phenotypes to be quantified that might be missed in short-term growth systems, such as subtle delays in nodule development and early nodule senescence. Also, the roots and nodules in the microcosm are easily viewed through the plate lid, so up-rooting of the plants for observation is not required. PMID:24121837

  20. Overlaps in the Transcriptional Profiles of Medicago truncatula Roots Inoculated with Two Different Glomus Fungi Provide Insights into the Genetic Program Activated during Arbuscular Mycorrhiza1[w

    PubMed Central

    Hohnjec, Natalija; Vieweg, Martin F.; Pühler, Alfred; Becker, Anke; Küster, Helge

    2005-01-01

    Arbuscular mycorrhiza (AM) is a widespread symbiotic association between plants and fungal microsymbionts that supports plant development under nutrient-limiting and various stress conditions. In this study, we focused on the overlapping genetic program activated by two commonly studied microsymbionts in addition to identifying AM-related genes. We thus applied 16,086 probe microarrays to profile the transcriptome of the model legume Medicago truncatula during interactions with Glomus mosseae and Glomus intraradices and specified a total of 201 plant genes as significantly coinduced at least 2-fold, with more than 160 being reported as AM induced for the first time. Several hundred genes were additionally up-regulated during a sole interaction, indicating that the plant genetic program activated in AM to some extent depends on the colonizing microsymbiont. Genes induced during both interactions specified AM-related nitrate, ion, and sugar transporters, enzymes involved in secondary metabolism, proteases, and Kunitz-type protease inhibitors. Furthermore, coinduced genes encoded receptor kinases and other components of signal transduction pathways as well as AM-induced transcriptional regulators, thus reflecting changes in signaling. By the use of reporter gene expression, we demonstrated that one member of the AM-induced gene family encoding blue copper binding proteins (MtBcp1) was both specifically and strongly up-regulated in arbuscule-containing regions of mycorrhizal roots. A comparison of the AM expression profiles to those of nitrogen-fixing root nodules suggested only a limited overlap between the genetic programs orchestrating root endosymbioses. PMID:15778460

  1. A set of fluorescent protein-based markers expressed from constitutive and arbuscular mycorrhiza-inducible promoters to label organelles, membranes and cytoskeletal elements in Medicago truncatula.

    PubMed

    Ivanov, Sergey; Harrison, Maria J

    2014-12-01

    Medicago truncatula is widely used for analyses of arbuscular mycorrhizal (AM) symbiosis and nodulation. To complement the genetic and genomic resources that exist for this species, we generated fluorescent protein fusions that label the nucleus, endoplasmic reticulum, Golgi apparatus, trans-Golgi network, plasma membrane, apoplast, late endosome/multivesicular bodies (MVB), transitory late endosome/ tonoplast, tonoplast, plastids, mitochondria, peroxisomes, autophagosomes, plasmodesmata, actin, microtubules, periarbuscular membrane (PAM) and periarbuscular apoplastic space (PAS) and expressed them from the constitutive AtUBQ10 promoter and the AM symbiosis-specific MtBCP1 promoter. All marker constructs showed the expected expression patterns and sub-cellular locations in M. truncatula root cells. As a demonstration of their utility, we used several markers to investigate AM symbiosis where root cells undergo major cellular alterations to accommodate their fungal endosymbiont. We demonstrate that changes in the position and size of the nuclei occur prior to hyphal entry into the cortical cells and do not require DELLA signaling. Changes in the cytoskeleton, tonoplast and plastids also occur in the colonized cells and in contrast to previous studies, we show that stromulated plastids are abundant in cells with developing and mature arbuscules, while lens-shaped plastids occur in cells with degenerating arbuscules. Arbuscule development and secretion of the PAM creates a periarbuscular apoplastic compartment which has been assumed to be continuous with apoplast of the cell. However, fluorescent markers secreted to the periarbuscular apoplast challenge this assumption. This marker resource will facilitate cell biology studies of AM symbiosis, as well as other aspects of legume biology.

  2. Systemic Signaling of the Plant Nitrogen Status Triggers Specific Transcriptome Responses Depending on the Nitrogen Source in Medicago truncatula1[W

    PubMed Central

    Ruffel, Sandrine; Freixes, Sandra; Balzergue, Sandrine; Tillard, Pascal; Jeudy, Christian; Martin-Magniette, Marie Laure; van der Merwe, Margaretha J.; Kakar, Klementina; Gouzy, Jerôme; Fernie, Alisdair R.; Udvardi, Michael; Salon, Christophe; Gojon, Alain; Lepetit, Marc

    2008-01-01

    Legumes can acquire nitrogen (N) from NO3−, NH4+, and N2 (through symbiosis with Rhizobium bacteria); however, the mechanisms by which uptake and assimilation of these N forms are coordinately regulated to match the N demand of the plant are currently unknown. Here, we find by use of the split-root approach in Medicago truncatula plants that NO3− uptake, NH4+ uptake, and N2 fixation are under general control by systemic signaling of plant N status. Indeed, irrespective of the nature of the N source, N acquisition by one side of the root system is repressed by high N supply to the other side. Transcriptome analysis facilitated the identification of over 3,000 genes that were regulated by systemic signaling of the plant N status. However, detailed scrutiny of the data revealed that the observation of differential gene expression was highly dependent on the N source. Localized N starvation results, in the unstarved roots of the same plant, in a strong compensatory up-regulation of NO3− uptake but not of either NH4+ uptake or N2 fixation. This indicates that the three N acquisition pathways do not always respond similarly to a change in plant N status. When taken together, these data indicate that although systemic signals of N status control root N acquisition, the regulatory gene networks targeted by these signals, as well as the functional response of the N acquisition systems, are predominantly determined by the nature of the N source. PMID:18287487

  3. RNA-seq transcriptome profiling reveals that Medicago truncatula nodules acclimate N2 fixation before emerging P deficiency reaches the nodules

    PubMed Central

    Cabeza, Ricardo A.; Liese, Rebecca; Lingner, Annika; von Stieglitz, Ilsabe; Neumann, Janice; Salinas-Riester, Gabriela; Pommerenke, Claudia; Dittert, Klaus; Schulze, Joachim

    2014-01-01

    Legume nodules are plant tissues with an exceptionally high concentration of phosphorus (P), which, when there is scarcity of P, is preferentially maintained there rather than being allocated to other plant organs. The hypothesis of this study was that nodules are affected before the P concentration in the organ declines during whole-plant P depletion. Nitrogen (N2) fixation and P concentration in various organs were monitored during a whole-plant P-depletion process in Medicago truncatula. Nodule gene expression was profiled through RNA-seq at day 5 of P depletion. Until that point in time P concentration in leaves reached a lower threshold but was maintained in nodules. N2-fixation activity per plant diverged from that of fully nourished plants beginning at day 5 of the P-depletion process, primarily because fewer nodules were being formed, while the activity of the existing nodules was maintained for as long as two weeks into P depletion. RNA-seq revealed nodule acclimation on a molecular level with a total of 1140 differentially expressed genes. Numerous genes for P remobilization from organic structures were increasingly expressed. Various genes involved in nodule malate formation were upregulated, while genes involved in fermentation were downregulated. The fact that nodule formation was strongly repressed with the onset of P deficiency is reflected in the differential expression of various genes involved in nodulation. It is concluded that plants follow a strategy to maintain N2 fixation and viable leaf tissue as long as possible during whole-plant P depletion to maintain their ability to react to emerging new P sources (e.g. through active P acquisition by roots). PMID:25151618

  4. Simulating the impact of genetic diversity of Medicago truncatula on germination and emergence using a crop emergence model for ideotype breeding

    PubMed Central

    Brunel-Muguet, S.; Aubertot, J.-N.; Dürr, C.

    2011-01-01

    Background and Aims Germination and heterotrophic growth are crucial steps for stand establishment. Numerical experiments based on the modelling of these early stages in relation to major environmental factors at sowing were used as a powerful tool to browse the effects of the genetic diversity of Medicago truncatula, one of the model legume species, under a range of agronomic scenarios, and to highlight the most important plant parameters for emergence. To this end, the emergence of several genotypes of M. truncatula was simulated under a range of sowing conditions with a germination and emergence simulation model. Methods After testing the predictive quality of the model by comparing simulations to field observations of several genotypes of M. truncatula, numerical experiments were performed under a wide range of environmental conditions (sowing dates × years × seedbed structure). Germination and emergence was simulated for a set of five genotypes previously parameterized and for two virtual genotypes engineered to maximize the potential effects of genetic diversity. Key Results The simulation results gave an average value of 5–10 % difference in final emergence between genotypes, which was low, but the analysis underlined considerable inter-annual variation. The effects of parameters describing germination and emergence processes were quantified and ranked according to their contribution to the variation in emergence. Seedling non-emergence was mainly related to mechanical obstacles (40–50 %). More generally, plant parameters that accelerated the emergence time course significantly contributed to limiting the risk of soil surface crusting occurring before seedling emergence. Conclusions The model-assisted analysis of the effects of genetic diversity demonstrated its usefulness in helping to identify the parameters which have most influence that could be improved by breeding programmes. These results should also enable a deeper analysis of the genetic

  5. Pea aphid promotes amino acid metabolism both in Medicago truncatula and bacteriocytes to favor aphid population growth under elevated CO2.

    PubMed

    Guo, Huijuan; Sun, Yucheng; Li, Yuefei; Tong, Bin; Harris, Marvin; Zhu-Salzman, Keyan; Ge, Feng

    2013-10-01

    Rising atmospheric CO(2) levels can dilute the nitrogen (N) resource in plant tissue, which is disadvantageous to many herbivorous insects. Aphids appear to be an exception that warrants further study. The effects of elevated CO(2) (750 ppm vs. 390 ppm) were evaluated on N assimilation and transamination by two Medicago truncatula genotypes, a N-fixing-deficient mutant (dnf1) and its wild-type control (Jemalong), with and without pea aphid (Acyrthosiphon pisum) infestation. Elevated CO(2) increased population abundance and feeding efficiency of aphids fed on Jemalong, but reduced those on dnf1. Without aphid infestation, elevated CO(2) increased photosynthetic rate, chlorophyll content, nodule number, biomass, and pod number for Jemalong, but only increased pod number and chlorophyll content for dnf1. Furthermore, aphid infested Jemalong plants had enhanced activities of N assimilation-related enzymes (glutamine synthetase, Glutamate synthase) and transamination-related enzymes (glutamate oxalate transaminase, glutamine phenylpyruvate transaminase), which presumably increased amino acid concentration in leaves and phloem sap under elevated CO(2). In contrast, aphid infested dnf1 plants had decreased activities of N assimilation-related enzymes and transmination-related enzymes and amino acid concentrations under elevated CO(2). Furthermore, elevated CO(2) up-regulated expression of genes relevant to amino acid metabolism in bacteriocytes of aphids associated with Jemalong, but down-regulated those associated with dnf1. Our results suggest that pea aphids actively elicit host responses that promote amino acid metabolism in both the host plant and in its bacteriocytes to favor the population growth of the aphid under elevated CO(2).

  6. Knockdown of the Symbiotic Sucrose Synthase MtSucS1 Affects Arbuscule Maturation and Maintenance in Mycorrhizal Roots of Medicago truncatula1[W

    PubMed Central

    Baier, Markus Christian; Keck, Matthias; Gödde, Victoria; Niehaus, Karsten; Küster, Helge; Hohnjec, Natalija

    2010-01-01

    The relevance of the symbiosis-induced Medicago truncatula sucrose synthase gene MtSucS1 for an efficient arbuscular mycorrhiza (AM) was studied using two independent antisense lines that displayed up to 10-fold reduced SucS1 levels in roots. Mycorrhizal MtSucS1-reduced lines exhibited an overall stunted aboveground growth under inorganic phosphorus limitation. Apart from a reduced plant height, shoot weight, and leaf development, a delayed flowering, resulting in a lower seed yield, was observed. In addition, the root-to-shoot and root weight ratios increased significantly. Gene expression studies demonstrated a major reversion of AM-associated transcription, exhibiting a significant repression of well-known plant AM marker and mycosymbiont genes, together indicating a diminished AM fungus colonization of MtSucS1-antisense lines. Concomitantly, gas chromatography-mass spectrometry-based metabolite profiling revealed that mycorrhizal MtSucS1-reduced lines were affected in important nodes of the carbon, nitrogen, and phosphorus metabolism, accentuating a physiological significance of MtSucS1 for AM. In fact, antisensing MtSucS1 provoked an impaired fungal colonization within the less abundant infected regions, evident from strongly reduced frequencies of internal hyphae, vesicles, and arbuscules. Moreover, arbuscules were early senescing, accompanied with a reduced development of mature arbuscules. This defective mycorrhiza status correlated with reduced phosphorus and nitrogen levels and was proportional to the extent of MtSucS1 knockdown. Together, our results point to an important role for MtSucS1 in the establishment and maintenance of arbuscules in the AM symbiosis. PMID:20007443

  7. Glutamine synthetase is a molecular target of nitric oxide in root nodules of Medicago truncatula and is regulated by tyrosine nitration.

    PubMed

    Melo, Paula M; Silva, Liliana S; Ribeiro, Isa; Seabra, Ana R; Carvalho, Helena G

    2011-11-01

    Nitric oxide (NO) is emerging as an important regulatory player in the Rhizobium-legume symbiosis, but its biological role in nodule functioning is still far from being understood. To unravel the signal transduction cascade and ultimately NO function, it is necessary to identify its molecular targets. This study provides evidence that glutamine synthetase (GS), a key enzyme for root nodule metabolism, is a molecular target of NO in root nodules of Medicago truncatula, being regulated by tyrosine (Tyr) nitration in relation to active nitrogen fixation. In vitro studies, using purified recombinant enzymes produced in Escherichia coli, demonstrated that the M. truncatula nodule GS isoenzyme (MtGS1a) is subjected to NO-mediated inactivation through Tyr nitration and identified Tyr-167 as the regulatory nitration site crucial for enzyme inactivation. Using a sandwich enzyme-linked immunosorbent assay, it is shown that GS is nitrated in planta and that its nitration status changes in relation to active nitrogen fixation. In ineffective nodules and in nodules fed with nitrate, two conditions in which nitrogen fixation is impaired and GS activity is reduced, a significant increase in nodule GS nitration levels was observed. Furthermore, treatment of root nodules with the NO donor sodium nitroprusside resulted in increased in vivo GS nitration accompanied by a reduction in GS activity. Our results support a role of NO in the regulation of nitrogen metabolism in root nodules and places GS as an important player in the process. We propose that the NO-mediated GS posttranslational inactivation is related to metabolite channeling to boost the nodule antioxidant defenses in response to NO.

  8. Risks and benefits of compost-like materials prepared by the thermal treatment of raw scallop hepatopancreas for supplying cadmium and the growth of alfalfa (Medicago sativa L.).

    PubMed

    Ushijima, Kensuke; Fukushima, Masami; Kanno, Shinya; Kanno, Itoko; Ohnishi, Mitsuhiro

    2016-01-01

    Scallop hepatopancreas, fishery waste, contains relatively high levels of Cd and organic nitrogen compounds, the latter of which represent a fertilizer. In this study, raw scallop hepatopancreas tissue was thermally treated with sawdust and red loam in the presence of an iron catalyst to produce compost-like materials (CLMs). Two CLM samples were prepared by varying the content of raw scallop hepatopancreas tissue: 46 wt.% for CLM-1 and 18 wt.% for CLM-2. Mixtures of control soil (CTL) and CLMs (CLM content: 10 and 25 wt.%) were examined for the growth of alfalfa (Medicago sativa L.) to evaluate the risks and benefits of using this material for fertilization. The Cd content in shoots and roots of alfalfa, that were grown in the presence of CLMs, was significantly higher than those for the plants grown in the CTL, indicating that Cd had accumulated in the plants from CLMs. The accumulation of Cd in the alfalfa roots was quite high in the case of the 25% CLM-1 sample. However, alfalfa growth was significantly promoted in the presence of 10% CLM-1. This can be attributed to the higher levels of nitrogen and humic substances, which serve as fertilizer components. Although the fertilization effect in case of CLM-1showed a potential benefit, the accumulation of Cd in alfalfa was clearly increased in the presence of both CLMs. In conclusion, the use of CLMs produced from raw scallop hepatopancreas tissue can be considered to have a desirable benefit from standpoint of its use as fertilizer, but is accompanied by a risk of the accumulation of Cd in alfalfa plants.

  9. Transcriptional profiling of Medicago truncatula under salt stress identified a novel CBF transcription factor MtCBF4 that plays an important role in abiotic stress responses

    PubMed Central

    2011-01-01

    Background Salt stress hinders the growth of plants and reduces crop production worldwide. However, different plant species might possess different adaptive mechanisms to mitigate salt stress. We conducted a detailed pathway analysis of transcriptional dynamics in the roots of Medicago truncatula seedlings under salt stress and selected a transcription factor gene, MtCBF4, for experimental validation. Results A microarray experiment was conducted using root samples collected 6, 24, and 48 h after application of 180 mM NaCl. Analysis of 11 statistically significant expression profiles revealed different behaviors between primary and secondary metabolism pathways in response to external stress. Secondary metabolism that helps to maintain osmotic balance was induced. One of the highly induced transcription factor genes was successfully cloned, and was named MtCBF4. Phylogenetic analysis revealed that MtCBF4, which belongs to the AP2-EREBP transcription factor family, is a novel member of the CBF transcription factor in M. truncatula. MtCBF4 is shown to be a nuclear-localized protein. Expression of MtCBF4 in M. truncatula was induced by most of the abiotic stresses, including salt, drought, cold, and abscisic acid, suggesting crosstalk between these abiotic stresses. Transgenic Arabidopsis over-expressing MtCBF4 enhanced tolerance to drought and salt stress, and activated expression of downstream genes that contain DRE elements. Over-expression of MtCBF4 in M. truncatula also enhanced salt tolerance and induced expression level of corresponding downstream genes. Conclusion Comprehensive transcriptomic analysis revealed complex mechanisms exist in plants in response to salt stress. The novel transcription factor gene MtCBF4 identified here played an important role in response to abiotic stresses, indicating that it might be a good candidate gene for genetic improvement to produce stress-tolerant plants. PMID:21718548

  10. A single amino acid substitution in a chitinase of the legume Medicago truncatula is sufficient to gain Nod-factor hydrolase activity

    PubMed Central

    Zhang, Lan-Yue; Cai, Jie; Li, Ru-Jie; Liu, Wei; Wagner, Christian; Wong, Kam-Bo; Xie, Zhi-Ping; Staehelin, Christian

    2016-01-01

    The symbiotic interaction between nitrogen-fixing rhizobia and legumes depends on lipo-chitooligosaccharidic Nod-factors (NFs). The NF hydrolase MtNFH1 of Medicago truncatula is a symbiotic enzyme that hydrolytically inactivates NFs with a C16 : 2 acyl chain produced by the microsymbiont Sinorhizobium meliloti 1021. MtNFH1 is related to class V chitinases (glycoside hydrolase family 18) but lacks chitinase activity. Here, we investigated the substrate specificity of MtNFH1-related proteins. MtCHIT5a and MtCHIT5b of M. truncatula as well as LjCHIT5 of Lotus japonicus showed chitinase activity, suggesting a role in plant defence. The enzymes failed to hydrolyse NFs from S. meliloti. NFs from Rhizobium leguminosarum with a C18 : 4 acyl moiety were neither hydrolysed by these chitinases nor by MtNFH1. Construction of chimeric proteins and further amino acid replacements in MtCHIT5b were performed to identify chitinase variants that gained the ability to hydrolyse NFs. A single serine-to-proline substitution was sufficient to convert MtCHIT5b into an NF-cleaving enzyme. MtNFH1 with the corresponding proline-to-serine substitution failed to hydrolyse NFs. These results are in agreement with a substrate-enzyme model that predicts NF cleavage when the C16 : 2 moiety is placed into a distinct fatty acid-binding cleft. Our findings support the view that MtNFH1 evolved from the ancestral MtCHIT5b by gene duplication and subsequent symbiosis-related neofunctionalization. PMID:27383628

  11. Effect of elevated CO₂ and temperature on the oxidative stress response to drought in Lolium perenne L. and Medicago sativa L.

    PubMed

    Farfan-Vignolo, Evelyn Roxana; Asard, Han

    2012-10-01

    Studies addressing the combined impact of multiple climate factors on plant abiotic stress responses are still scarce. We investigated physiological and molecular (antioxidant), responses to water deficit, in grassland-model species, Lolium perenne L. and Medicago lupulina L., under future climate conditions, i.e. elevated CO₂ (+CO₂, +375 ppm) and elevated temperature (+T, +3 °C). Elevated CO₂, but not warming, significantly increased biomass (gDW) in L. perenne, but not in M. lupulina. Photosynthesis (A(sat)) and stomatal conductance (g(s)), were differently affected by climate in each species, L. perenne generally being more sensitive. Elevated CO₂ increased lipid peroxidation levels in M. lupulina, but not in L. perenne, and had no effect on protein oxidation and little effect on antioxidant levels. Drought stress caused severe inhibition in biomass and photosynthesis, most severely in L. perenne, and strongly increased oxidative damage. Elevated CO₂ protected against the drought-induced damage. Decreased activities of APX and POX may indicate lower levels of oxidative challenge (relaxation) at the level of H₂O₂ production. Polyphenols, tocopherols and antioxidant capacity, increased under drought stress, in all climate conditions. Elevated CO₂, increased reduced ascorbate (ASC) and reduced glutathione (GSH), and their redox status, in both species, although to different levels. Changes in activities of key ASC/GSH cycle enzymes, under stress and climate treatments, showed weak correlations with ASC and GSH levels, indicating the complexity of this network. Together this work supports the idea that redox changes are involved in responses to climate changes, in the absence and presence of water-deficit stress.

  12. Co-transforming bar and CsALDH Genes Enhanced Resistance to Herbicide and Drought and Salt Stress in Transgenic Alfalfa (Medicago sativa L.).

    PubMed

    Duan, Zhen; Zhang, Daiyu; Zhang, Jianquan; Di, Hongyan; Wu, Fan; Hu, Xiaowen; Meng, Xuanchen; Luo, Kai; Zhang, Jiyu; Wang, Yanrong

    2015-01-01

    Drought and high salinity are two major abiotic factors that restrict the productivity of alfalfa. By application of the Agrobacterium-mediated transformation method, an oxidative responsive gene, CsALDH12A1, from the desert grass Cleistogenes songorica together with the bar gene associated with herbicide resistance, were co-transformed into alfalfa (Medicago sativa L.). From the all 90 transformants, 16 were positive as screened by spraying 1 mL L(-1) 10% Basta solution and molecularly diagnosis using PCR. Real-time PCR analysis indicated that drought and salt stress induced high CsALDH expression in the leaves of the transgenic plants. The CsALDH expression levels under drought (15 d) and salt stress (200 mM NaCl) were 6.11 and 6.87 times higher than in the control plants, respectively. In comparison to the WT plants, no abnormal phenotypes were observed among the transgenic plants, which showed significant enhancement of tolerance to 15 d of drought and 10 d of salinity treatment. Evaluation of the physiological and biochemical indices during drought and salt stress of the transgenic plants revealed relatively lower Na(+) content and higher K(+) content in the leaves relative to the WT plants, a reduction of toxic on effects and maintenance of osmotic adjustment. In addition, the transgenic plants could maintain a higher relative water content level, higher shoot biomass, fewer changes in the photosystem, decreased membrane injury, and a lower level of osmotic stress. These results indicate that the co-expression of the introduced bar and CsALDH genes enhanced the herbicide, drought and salt tolerance of alfalfa and therefore can potentially be used as a novel genetic resource for the future breeding programs to develop new cultivars.

  13. WRKY domain-encoding genes of a crop legume chickpea (Cicer arietinum): comparative analysis with Medicago truncatula WRKY family and characterization of group-III gene(s)

    PubMed Central

    Kumar, Kamal; Srivastava, Vikas; Purayannur, Savithri; Kaladhar, V. Chandra; Cheruvu, Purnima Jaiswal; Verma, Praveen Kumar

    2016-01-01

    The WRKY genes have been identified as important transcriptional modulators predominantly during the environmental stresses, but they also play critical role at various stages of plant life cycle. We report the identification of WRKY domain (WD)-encoding genes from galegoid clade legumes chickpea (Cicer arietinum L.) and barrel medic (Medicago truncatula). In total, 78 and 98 WD-encoding genes were found in chickpea and barrel medic, respectively. Comparative analysis suggests the presence of both conserved and unique WRKYs, and expansion of WRKY family in M. truncatula primarily by tandem duplication. Exclusively found in galegoid legumes, CaWRKY16 and its orthologues encode for a novel protein having a transmembrane and partial Exo70 domains flanking a group-III WD. Genomic region of galegoids, having CaWRKY16, is more dynamic when compared with millettioids. In onion cells, fused CaWRKY16-EYFP showed punctate fluorescent signals in cytoplasm. The chickpea WRKY group-III genes were further characterized for their transcript level modulation during pathogenic stress and treatments of abscisic acid, jasmonic acid, and salicylic acid (SA) by real-time PCR. Differential regulation of genes was observed during Ascochyta rabiei infection and SA treatment. Characterization of A. rabiei and SA inducible gene CaWRKY50 showed that it localizes to plant nucleus, binds to W-box, and have a C-terminal transactivation domain. Overexpression of CaWRKY50 in tobacco plants resulted in early flowering and senescence. The in-depth comparative account presented here for two legume WRKY genes will be of great utility in hastening functional characterization of crop legume WRKYs and will also help in characterization of Exo70Js. PMID:27060167

  14. A Sinorhizobium meliloti-specific N-acyl homoserine lactone quorum-sensing signal increases nodule numbers in Medicago truncatula independent of autoregulation

    PubMed Central

    Veliz-Vallejos, Debora F.; van Noorden, Giel E.; Yuan, Mengqi; Mathesius, Ulrike

    2014-01-01

    N-acyl homoserine lactones (AHLs) act as quorum sensing signals that regulate cell-density dependent behaviors in many gram-negative bacteria, in particular those important for plant-microbe interactions. AHLs can also be recognized by plants, and this may influence their interactions with bacteria. Here we tested whether the exposure to AHLs affects the nodule-forming symbiosis between legume hosts and rhizobia. We treated roots of the model legume, Medicago truncatula, with a range of AHLs either from its specific symbiont, Sinorhizobium meliloti, or from the potential pathogens, Pseudomonas aeruginosa and Agrobacterium vitis. We found increased numbers of nodules formed on root systems treated with the S. meliloti-specific AHL, 3-oxo-C14-homoserine lactone, at a concentration of 1 μM, while the other AHLs did not result in significant changes to nodule numbers. We did not find any evidence for altered nodule invasion by the rhizobia. Quantification of flavonoids that could act as nod gene inducers in S. meliloti did not show any correlation with increased nodule numbers. The effects of AHLs were specific for an increase in nodule numbers, but not lateral root numbers or root length. Increased nodule numbers following 3-oxo-C14-homoserine lactone treatment were under control of autoregulation of nodulation and were still observed in the autoregulation mutant, sunn4 (super numeric nodules4). However, increases in nodule numbers by 3-oxo-C14-homoserine lactone were not found in the ethylene-insensitive sickle mutant. A comparison between M. truncatula with M. sativa (alfalfa) and Trifolium repens (white clover) showed that the observed effects of AHLs on nodule numbers were specific to M. truncatula, despite M. sativa nodulating with the same symbiont. We conclude that plant perception of the S. meliloti-specific 3-oxo-C14-homoserine lactone influences nodule numbers in M. truncatula via an ethylene-dependent, but autoregulation-independent mechanism. PMID

  15. Citric acid- and Tween(®) 80-assisted phytoremediation of a co-contaminated soil: alfalfa (Medicago sativa L.) performance and remediation potential.

    PubMed

    Agnello, A C; Huguenot, D; van Hullebusch, E D; Esposito, G

    2016-05-01

    A pot experiment was designed to assess the phytoremediation potential of alfalfa (Medicago sativa L.) in a co-contaminated (i.e., heavy metals and petroleum hydrocarbons) soil and the influence of citric acid and Tween(®) 80 (polyethylene glycol sorbitan monooleate), applied individually and combined together, for their possible use in chemically assisted phytoremediation. The results showed that alfalfa plants could tolerate and grow in a co-contaminated soil. Over a 90-day experimental time, shoot and root biomass increased and negligible plant mortality occurred. Heavy metals were uptaken by alfalfa to a limited extent, mostly by plant roots, and their concentration in plant tissues were in the following order: Zn > Cu > Pb. Microbial population (alkane-degrading microorganisms) and activity (lipase enzyme) were enhanced in the presence of alfalfa with rhizosphere effects of 9.1 and 1.5, respectively, after 90 days. Soil amendments did not significantly enhance plant metal concentration or total uptake. In contrast, the combination of citric acid and Tween(®) 80 significantly improved alkane-degrading microorganisms (2.4-fold increase) and lipase activity (5.3-fold increase) in the rhizosphere of amended plants, after 30 days of experiment. This evidence supports a favorable response of alfalfa in terms of tolerance to a co-contaminated soil and improvement of rhizosphere microbial number and activity, additionally enhanced by the joint application of citric acid and Tween(®) 80, which could be promising for future phytoremediation applications.

  16. Analysis of gene expression during flowering in apomeiotic mutants of Medicago spp.: cloning of ESTs and candidate genes for 2n eggs.

    PubMed

    Barcaccia, G; Varotto, S; Meneghetti, S; Albertini, E; Porceddu, A; Parrini, P; Lucchin, M

    2001-12-01

    Mutants showing features of apomixis have been documented in alfalfa (Medicago sativa L.), a natural outcrossing sexual species. A differential display of mRNAs that combines cDNA-AFLP markers and bulked segregant analysis was carried out with the aim of selecting expressed sequence tags (ESTs) and cloning candidate genes for apomeiosis in mutants of alfalfa characterized by 2n egg formation at high frequencies. The approach enabled us to select either mutant- or wild type-specific transcript derived-fragments and to detect transcriptional changes potentially related to 2n eggs. Sequence alignments of a subset of 40 polymorphic clones showed significant homologies to genes of known function. An EST with identity to a β-tubulin gene, highly expressed in the wild type and poorly expressed in the apomeiotic mutants, and an EST with identity to a Mob1-like gene, qualitatively polymorphic between pre- and post-meiotic stages, were selected as candidate genes for apomeiosis because of their putative roles in the cell cycle. A number of clone-specific primers were designed for performing both 5' and 3' rapid amplification of cDNA ends to obtain the full-length clones. Southern blot hybridization revealed that both clones belong to a multi-gene family with a minimum of three genomic DNA members each. Northern blot hybridization of total RNA samples and in situ hybridization of whole buds enabled the definition of their temporal and spatial expression patterns in reproductive organs. Experimental achievements towards the elucidation of apomeiotic megasporogenesis in alfalfa are presented and discussed.

  17. Both Plant and Bacterial Nitrate Reductases Contribute to Nitric Oxide Production in Medicago truncatula Nitrogen-Fixing Nodules1[W][OA

    PubMed Central

    Horchani, Faouzi; Prévot, Marianne; Boscari, Alexandre; Evangelisti, Edouard; Meilhoc, Eliane; Bruand, Claude; Raymond, Philippe; Boncompagni, Eric; Aschi-Smiti, Samira; Puppo, Alain; Brouquisse, Renaud

    2011-01-01

    Nitric oxide (NO) is a signaling and defense molecule of major importance in living organisms. In the model legume Medicago truncatula, NO production has been detected in the nitrogen fixation zone of the nodule, but the systems responsible for its synthesis are yet unknown and its role in symbiosis is far from being elucidated. In this work, using pharmacological and genetic approaches, we explored the enzymatic source of NO production in M. truncatula-Sinorhizobium meliloti nodules under normoxic and hypoxic conditions. When transferred from normoxia to hypoxia, nodule NO production was rapidly increased, indicating that NO production capacity is present in functioning nodules and may be promptly up-regulated in response to decreased oxygen availability. Contrary to roots and leaves, nodule NO production was stimulated by nitrate and nitrite and inhibited by tungstate, a nitrate reductase inhibitor. Nodules obtained with either plant nitrate reductase RNA interference double knockdown (MtNR1/2) or bacterial nitrate reductase-deficient (napA) and nitrite reductase-deficient (nirK) mutants, or both, exhibited reduced nitrate or nitrite reductase activities and NO production levels. Moreover, NO production in nodules was found to be inhibited by electron transfer chain inhibitors, and nodule energy state (ATP-ADP ratio) was significantly reduced when nodules were incubated in the presence of tungstate. Our data indicate that both plant and bacterial nitrate reductase and electron transfer chains are involved in NO synthesis. We propose the existence of a nitrate-NO respiration process in nodules that could play a role in the maintenance of the energy status required for nitrogen fixation under oxygen-limiting conditions. PMID:21139086

  18. Structural Studies of Medicago truncatula Histidinol Phosphate Phosphatase from Inositol Monophosphatase Superfamily Reveal Details of Penultimate Step of Histidine Biosynthesis in Plants.

    PubMed

    Ruszkowski, Milosz; Dauter, Zbigniew

    2016-05-01

    The penultimate enzyme in the histidine biosynthetic pathway catalyzes dephosphorylation of l-histidinol 1-phosphate (HOLP) into l-histidinol. The recently discovered in Arabidopsis thaliana plant-type histidinol phosphate phosphatase (HPP) shares no homology with the two other HPP superfamilies known previously in prokaryotes and resembles myo-inositol monophosphatases (IMPases). In this work, identification of an HPP enzyme from a model legume, Medicago truncatula (MtHPP) was based on the highest sequence identity to A. thaliana enzyme. Biochemical assays confirmed that MtHPP was able to cleave inorganic phosphate from HOLP but not from d-myo-inositol-1-phosphate, the main substrate of IMPases. Dimers of MtHPP, determined by size exclusion chromatography, in the presence of CO2 or formaldehyde form mutual, methylene-bridged cross-links between Lys(158) and Cys(245) residues. Four high resolution crystal structures, namely complexes with HOLP (substrate), l-histidinol (product), and PO4 (3-) (by-product) as well as the structure showing the cross-linking between two MtHPP molecules, provide detailed structural information on the enzyme. Based on the crystal structures, the enzymatic reaction mechanism of IMPases is accustomed to fit the data for MtHPP. The enzymatic reaction, which requires Mg(2+) cations, is catalyzed mainly by amino acid residues from the N-terminal domain. The C-terminal domain, sharing little identity with IMPases, is responsible for the substrate specificity (i.e. allows the enzyme to distinguish between HOLP and d-myo-inositol-1-phosphate). Structural features, mainly the presence of a conserved Asp(246), allow MtHPP to bind HOLP specifically. PMID:26994138

  19. A Laser Dissection-RNAseq Analysis Highlights the Activation of Cytokinin Pathways by Nod Factors in the Medicago truncatula Root Epidermis.

    PubMed

    Jardinaud, Marie-Françoise; Boivin, Stéphane; Rodde, Nathalie; Catrice, Olivier; Kisiala, Anna; Lepage, Agnes; Moreau, Sandra; Roux, Brice; Cottret, Ludovic; Sallet, Erika; Brault, Mathias; Emery, R J Neil; Gouzy, Jérôme; Frugier, Florian; Gamas, Pascal

    2016-07-01

    Nod factors (NFs) are lipochitooligosaccharidic signal molecules produced by rhizobia, which play a key role in the rhizobium-legume symbiotic interaction. In this study, we analyzed the gene expression reprogramming induced by purified NF (4 and 24 h of treatment) in the root epidermis of the model legume Medicago truncatula Tissue-specific transcriptome analysis was achieved by laser-capture microdissection coupled to high-depth RNA sequencing. The expression of 17,191 genes was detected in the epidermis, among which 1,070 were found to be regulated by NF addition, including previously characterized NF-induced marker genes. Many genes exhibited strong levels of transcriptional activation, sometimes only transiently at 4 h, indicating highly dynamic regulation. Expression reprogramming affected a variety of cellular processes, including perception, signaling, regulation of gene expression, as well as cell wall, cytoskeleton, transport, metabolism, and defense, with numerous NF-induced genes never identified before. Strikingly, early epidermal activation of cytokinin (CK) pathways was indicated, based on the induction of CK metabolic and signaling genes, including the CRE1 receptor essential to promote nodulation. These transcriptional activations were independently validated using promoter:β-glucuronidase fusions with the MtCRE1 CK receptor gene and a CK response reporter (TWO COMPONENT SIGNALING SENSOR NEW). A CK pretreatment reduced the NF induction of the EARLY NODULIN11 (ENOD11) symbiotic marker, while a CK-degrading enzyme (CYTOKININ OXIDASE/DEHYDROGENASE3) ectopically expressed in the root epidermis led to increased NF induction of ENOD11 and nodulation. Therefore, CK may play both positive and negative roles in M. truncatula nodulation. PMID:27217496

  20. A Laser Dissection-RNAseq Analysis Highlights the Activation of Cytokinin Pathways by Nod Factors in the Medicago truncatula Root Epidermis.

    PubMed

    Jardinaud, Marie-Françoise; Boivin, Stéphane; Rodde, Nathalie; Catrice, Olivier; Kisiala, Anna; Lepage, Agnes; Moreau, Sandra; Roux, Brice; Cottret, Ludovic; Sallet, Erika; Brault, Mathias; Emery, R J Neil; Gouzy, Jérôme; Frugier, Florian; Gamas, Pascal

    2016-07-01

    Nod factors (NFs) are lipochitooligosaccharidic signal molecules produced by rhizobia, which play a key role in the rhizobium-legume symbiotic interaction. In this study, we analyzed the gene expression reprogramming induced by purified NF (4 and 24 h of treatment) in the root epidermis of the model legume Medicago truncatula Tissue-specific transcriptome analysis was achieved by laser-capture microdissection coupled to high-depth RNA sequencing. The expression of 17,191 genes was detected in the epidermis, among which 1,070 were found to be regulated by NF addition, including previously characterized NF-induced marker genes. Many genes exhibited strong levels of transcriptional activation, sometimes only transiently at 4 h, indicating highly dynamic regulation. Expression reprogramming affected a variety of cellular processes, including perception, signaling, regulation of gene expression, as well as cell wall, cytoskeleton, transport, metabolism, and defense, with numerous NF-induced genes never identified before. Strikingly, early epidermal activation of cytokinin (CK) pathways was indicated, based on the induction of CK metabolic and signaling genes, including the CRE1 receptor essential to promote nodulation. These transcriptional activations were independently validated using promoter:β-glucuronidase fusions with the MtCRE1 CK receptor gene and a CK response reporter (TWO COMPONENT SIGNALING SENSOR NEW). A CK pretreatment reduced the NF induction of the EARLY NODULIN11 (ENOD11) symbiotic marker, while a CK-degrading enzyme (CYTOKININ OXIDASE/DEHYDROGENASE3) ectopically expressed in the root epidermis led to increased NF induction of ENOD11 and nodulation. Therefore, CK may play both positive and negative roles in M. truncatula nodulation.

  1. High toxicity and specificity of the saponin 3-GlcA-28-AraRhaxyl-medicagenate, from Medicago truncatula seeds, for Sitophilus oryzae

    PubMed Central

    2012-01-01

    Background Because of the increasingly concern of consumers and public policy about problems for environment and for public health due to chemical pesticides, the search for molecules more safe is currently of great importance. Particularly, plants are able to fight the pathogens as insects, bacteria or fungi; so that plants could represent a valuable source of new molecules. Results It was observed that Medicago truncatula seed flour displayed a strong toxic activity towards the adults of the rice weevil Sitophilus oryzae (Coleoptera), a major pest of stored cereals. The molecule responsible for toxicity was purified, by solvent extraction and HPLC, and identified as a saponin, namely 3-GlcA-28-AraRhaxyl-medicagenate. Saponins are detergents, and the CMC of this molecule was found to be 0.65 mg per mL. Neither the worm Caenorhabditis elegans nor the bacteria E. coli were found to be sensitive to this saponin, but growth of the yeast Saccharomyces cerevisiae was inhibited at concentrations higher than 100 μg per mL. The purified molecule is toxic for the adults of the rice weevils at concentrations down to 100 μg per g of food, but this does not apply to the others insects tested, including the coleopteran Tribolium castaneum and the Sf9 insect cultured cells. Conclusions This specificity for the weevil led us to investigate this saponin potential for pest control and to propose the hypothesis that this saponin has a specific mode of action, rather than acting via its non-specific detergent properties. PMID:22536832

  2. Contribution of NFP LysM Domains to the Recognition of Nod Factors during the Medicago truncatula/Sinorhizobium meliloti Symbiosis

    PubMed Central

    Bensmihen, Sandra; de Billy, Françoise; Gough, Clare

    2011-01-01

    The root nodule nitrogen fixing symbiosis between legume plants and soil bacteria called rhizobia is of great agronomical and ecological interest since it provides the plant with fixed atmospheric nitrogen. The establishment of this symbiosis is mediated by the recognition by the host plant of lipo-chitooligosaccharides called Nod Factors (NFs), produced by the rhizobia. This recognition is highly specific, as precise NF structures are required depending on the host plant. Here, we study the importance of different LysM domains of a LysM-Receptor Like Kinase (LysM-RLK) from Medicago truncatula called Nod factor perception (NFP) in the recognition of different substitutions of NFs produced by its symbiont Sinorhizobium meliloti. These substitutions are a sulphate group at the reducing end, which is essential for host specificity, and a specific acyl chain at the non-reducing end, that is critical for the infection process. The NFP extracellular domain (ECD) contains 3 LysM domains that are predicted to bind NFs. By swapping the whole ECD or individual LysM domains of NFP for those of its orthologous gene from pea, SYM10 (a legume plant that interacts with another strain of rhizobium producing NFs with different substitutions), we showed that NFP is not directly responsible for specific recognition of the sulphate substitution of S. meliloti NFs, but probably interacts with the acyl substitution. Moreover, we have demonstrated the importance of the NFP LysM2 domain for rhizobial infection and we have pinpointed the importance of a single leucine residue of LysM2 in that step of the symbiosis. Together, our data put into new perspective the recognition of NFs in the different steps of symbiosis in M. truncatula, emphasising the probable existence of a missing component for early NF recognition and reinforcing the important role of NFP for NF recognition during rhizobial infection. PMID:22087221

  3. WRKY domain-encoding genes of a crop legume chickpea (Cicer arietinum): comparative analysis with Medicago truncatula WRKY family and characterization of group-III gene(s).

    PubMed

    Kumar, Kamal; Srivastava, Vikas; Purayannur, Savithri; Kaladhar, V Chandra; Cheruvu, Purnima Jaiswal; Verma, Praveen Kumar

    2016-06-01

    The WRKY genes have been identified as important transcriptional modulators predominantly during the environmental stresses, but they also play critical role at various stages of plant life cycle. We report the identification of WRKY domain (WD)-encoding genes from galegoid clade legumes chickpea (Cicer arietinum L.) and barrel medic (Medicago truncatula). In total, 78 and 98 WD-encoding genes were found in chickpea and barrel medic, respectively. Comparative analysis suggests the presence of both conserved and unique WRKYs, and expansion of WRKY family in M. truncatula primarily by tandem duplication. Exclusively found in galegoid legumes, CaWRKY16 and its orthologues encode for a novel protein having a transmembrane and partial Exo70 domains flanking a group-III WD. Genomic region of galegoids, having CaWRKY16, is more dynamic when compared with millettioids. In onion cells, fused CaWRKY16-EYFP showed punctate fluorescent signals in cytoplasm. The chickpea WRKY group-III genes were further characterized for their transcript level modulation during pathogenic stress and treatments of abscisic acid, jasmonic acid, and salicylic acid (SA) by real-time PCR. Differential regulation of genes was observed during Ascochyta rabiei infection and SA treatment. Characterization of A. rabiei and SA inducible gene CaWRKY50 showed that it localizes to plant nucleus, binds to W-box, and have a C-terminal transactivation domain. Overexpression of CaWRKY50 in tobacco plants resulted in early flowering and senescence. The in-depth comparative account presented here for two legume WRKY genes will be of great utility in hastening functional characterization of crop legume WRKYs and will also help in characterization of Exo70Js. PMID:27060167

  4. WRKY domain-encoding genes of a crop legume chickpea (Cicer arietinum): comparative analysis with Medicago truncatula WRKY family and characterization of group-III gene(s).

    PubMed

    Kumar, Kamal; Srivastava, Vikas; Purayannur, Savithri; Kaladhar, V Chandra; Cheruvu, Purnima Jaiswal; Verma, Praveen Kumar

    2016-06-01

    The WRKY genes have been identified as important transcriptional modulators predominantly during the environmental stresses, but they also play critical role at various stages of plant life cycle. We report the identification of WRKY domain (WD)-encoding genes from galegoid clade legumes chickpea (Cicer arietinum L.) and barrel medic (Medicago truncatula). In total, 78 and 98 WD-encoding genes were found in chickpea and barrel medic, respectively. Comparative analysis suggests the presence of both conserved and unique WRKYs, and expansion of WRKY family in M. truncatula primarily by tandem duplication. Exclusively found in galegoid legumes, CaWRKY16 and its orthologues encode for a novel protein having a transmembrane and partial Exo70 domains flanking a group-III WD. Genomic region of galegoids, having CaWRKY16, is more dynamic when compared with millettioids. In onion cells, fused CaWRKY16-EYFP showed punctate fluorescent signals in cytoplasm. The chickpea WRKY group-III genes were further characterized for their transcript level modulation during pathogenic stress and treatments of abscisic acid, jasmonic acid, and salicylic acid (SA) by real-time PCR. Differential regulation of genes was observed during Ascochyta rabiei infection and SA treatment. Characterization of A. rabiei and SA inducible gene CaWRKY50 showed that it localizes to plant nucleus, binds to W-box, and have a C-terminal transactivation domain. Overexpression of CaWRKY50 in tobacco plants resulted in early flowering and senescence. The in-depth comparative account presented here for two legume WRKY genes will be of great utility in hastening functional characterization of crop legume WRKYs and will also help in characterization of Exo70Js.

  5. A Laser Dissection-RNAseq Analysis Highlights the Activation of Cytokinin Pathways by Nod Factors in the Medicago truncatula Root Epidermis1[OPEN

    PubMed Central

    Jardinaud, Marie-Françoise; Lepage, Agnes; Moreau, Sandra; Cottret, Ludovic; Brault, Mathias; Gouzy, Jérôme; Frugier, Florian

    2016-01-01

    Nod factors (NFs) are lipochitooligosaccharidic signal molecules produced by rhizobia, which play a key role in the rhizobium-legume symbiotic interaction. In this study, we analyzed the gene expression reprogramming induced by purified NF (4 and 24 h of treatment) in the root epidermis of the model legume Medicago truncatula. Tissue-specific transcriptome analysis was achieved by laser-capture microdissection coupled to high-depth RNA sequencing. The expression of 17,191 genes was detected in the epidermis, among which 1,070 were found to be regulated by NF addition, including previously characterized NF-induced marker genes. Many genes exhibited strong levels of transcriptional activation, sometimes only transiently at 4 h, indicating highly dynamic regulation. Expression reprogramming affected a variety of cellular processes, including perception, signaling, regulation of gene expression, as well as cell wall, cytoskeleton, transport, metabolism, and defense, with numerous NF-induced genes never identified before. Strikingly, early epidermal activation of cytokinin (CK) pathways was indicated, based on the induction of CK metabolic and signaling genes, including the CRE1 receptor essential to promote nodulation. These transcriptional activations were independently validated using promoter:β-glucuronidase fusions with the MtCRE1 CK receptor gene and a CK response reporter (TWO COMPONENT SIGNALING SENSOR NEW). A CK pretreatment reduced the NF induction of the EARLY NODULIN11 (ENOD11) symbiotic marker, while a CK-degrading enzyme (CYTOKININ OXIDASE/DEHYDROGENASE3) ectopically expressed in the root epidermis led to increased NF induction of ENOD11 and nodulation. Therefore, CK may play both positive and negative roles in M. truncatula nodulation. PMID:27217496

  6. A single gene, AIN, in Medicago truncatula mediates a hypersensitive response to both bluegreen aphid and pea aphid, but confers resistance only to bluegreen aphid.

    PubMed

    Klingler, John P; Nair, Ramakrishnan M; Edwards, Owain R; Singh, Karam B

    2009-01-01

    Biotic stress in plants frequently induces a hypersensitive response (HR). This distinctive reaction has been studied intensively in several pathosystems and has shed light on the biology of defence signalling. Compared with microbial pathogens, relatively little is known about the role of the HR in defence against insects. Reference genotype A17 of Medicago truncatula Gaertn., a model legume, responds to aphids of the genus Acyrthosiphon with necrotic lesions resembling a HR. In this study, the biochemical nature of this response, its mode of inheritance, and its relationship with defence against aphids were investigated. The necrotic lesion phenotype and resistance to the bluegreen aphid (BGA, Acyrthosiphon kondoi Shinji) and the pea aphid (PA, Acyrthosiphon pisum (Harris)) were analysed using reference genotypes A17 and A20, their F(2) progeny and recombinant inbred lines. BGA-induced necrotic lesions co-localized with the production of H(2)O(2), consistent with an oxidative burst widely associated with hypersensitivity. This HR correlated with stronger resistance to BGA in A17 than in A20; these phenotypes cosegregated as a semi-dominant gene, AIN (Acyrthosiphon-induced necrosis). In contrast to BGA, stronger resistance to PA in A17, compared with A20, did not cosegregate with a PA-induced HR. The AIN locus resides in a cluster of sequences predicted to encode the CC-NBS-LRR subfamily of resistance proteins. AIN-mediated resistance presents a novel opportunity to use a model plant and model aphid to study the role of the HR in defence responses to phloem-feeding insects. PMID:19690018

  7. Nitrogen Reserves, Spring Regrowth and Winter Survival of Field-grown Alfalfa (Medicago sativa) Defoliated in the Autumn

    PubMed Central

    DHONT, CATHERINE; CASTONGUAY, YVES; NADEAU, PAUL; BÉLANGER, GILLES; DRAPEAU, RAYNALD; LABERGE, SERGE; AVICE, JEAN-CHRISTOPHE; CHALIFOUR, FRANÇOIS-P.

    2006-01-01

    • Aims The objective of the study was to characterize variations in proline, arginine, histidine, vegetative storage proteins, and cold-inducible gene expression in overwintering roots of field-grown alfalfa, in response to autumn defoliation, and in relation to spring regrowth and winter survival. • Methods Field trials, established in 1996 in eastern Canada, consisted of two alfalfa cultivars (‘AC Caribou’ and ‘WL 225’) defoliated in 1997 and 1998 either only twice during the summer or three times with the third defoliation taken 400, 500 or 600 growing degree days (basis 5 °C) after the second summer defoliation. • Key Results The root accumulation of proline, arginine, histidine and soluble proteins of 32, 19 and 15 kDa, characterized as alfalfa vegetative storage proteins, was reduced the following spring by an early autumn defoliation at 400 or 500 growing degree days in both cultivars; the 600-growing-degree-days defoliation treatment had less or no effect. Transcript levels of the cold-inducible gene msaCIA, encoding a glycine-rich protein, were markedly reduced by autumn defoliation in ‘WL 225’, but remained unaffected in the more winter-hardy cultivar ‘AC Caribou’. The expression of another cold-inducible gene, the dehydrin homologue msaCIG, was not consistently affected by autumn defoliation. Principal component analyses, including components of root organic reserves at the onset of winter, along with yield and plant density in the following spring, revealed that (a) amino acids and soluble proteins are positively related to the vigour of spring regrowth but poorly related to winter survival and (b) winter survival, as indicated by plant density in the spring, is associated with higher concentrations of cryoprotective sugars in alfalfa roots the previous autumn. • Conclusions An untimely autumn defoliation of alfalfa reduces root accumulation of specific N reserves such as proline, arginine, histidine and vegetative storage

  8. Effect of Microgravity on Early Events of Biological Nitrogen Fixation in Medicago Truncatula: Initial Results from the SyNRGE Experiment

    NASA Technical Reports Server (NTRS)

    Stutte, Gary W.; Roberts, Michael S.

    2011-01-01

    SyNRGE (Symbiotic Nodulation in a Reduced Gravity Environment) was a sortie mission on STS-135 in the Biological Research in Canisters (BRIC) hardware to study the effect of microgravity on a plant-microbe symbiosis resulting in biological nitrogen fixation. Medicago truncatula, a model species of the legume family, was inoculated with its bacterial symbiont, Sinorhizobium meliloti, to observe early events associated with infection and nodulation in Petri Dish Fixation Units (PDFUs). Two sets of experiments were conducted in orbit and in 24-hour delayed ground controls. Experiment one was designed to determine if S. meliloti infect M. truncatula and initiate physiological changes associated with nodule formation. Roots of five-day-old M. truncatula cultivar Jemalong A17 (Enodll::gus) were inoculated 24 hr before launch with either S. meliloti strain 1021 or strain ABS7 and integrated into BRIC-PDFU hardware placed in a 4 C Cold Bag for launch on Atlantis. Inoculated plants and uninoculated controls were maintained in the dark at ambient temperature in the middeck of STS-135 for 11 days before fixation in RNAlater(tM) by crew activation of the PDFU. Experiment two was designed to determine if microgravity altered the process of bacterial infection and host plant nodule formation. Seeds of two M. truncatula cultivar Jemalong A17 lines, the Enodll::gus used in experiment 1, and SUNN, a super-nodulating mutant of A17, were germinated on orbit for 11 days in the middeck cabin and returned to Earth alive inside of BRIC-PDFU's at 4 C. S. meliloti strains 1021 and ABS7 were cultivated separately in broth culture on orbit and also returned to Earth alive. After landing, flight- and groundgrown plants and bacteria were transferred from BRIC-PDFU's into Nunc(tm) 4-well plates for reciprocity crosses. Rates of plant growth and nodule development on Buffered Nodulation Medium (lacking nitrogen) were measured for 14 days. Preliminary analysis' of Experiment 1 confirms that

  9. Genes conferring copper resistance in Sinorhizobium meliloti CCNWSX0020 also promote the growth of Medicago lupulina in copper-contaminated soil.

    PubMed

    Li, Zhefei; Ma, Zhanqiang; Hao, Xiuli; Rensing, Christopher; Wei, Gehong

    2014-03-01

    Sinorhizobium meliloti CCNWSX0020, isolated from root nodules of Medicago lupulina growing in gold mine tailings in the northwest of China, displayed both copper resistance and growth promotion of leguminous plants in copper-contaminated soil. Nevertheless, the genetic and biochemical mechanisms responsible for copper resistance in S. meliloti CCNWSX0020 remained uncharacterized. To investigate genes involved in copper resistance, an S. meliloti CCNWSX0020 Tn5 insertion library of 14,000 mutants was created. Five copper-sensitive mutants, named SXa-1, SXa-2, SXc-1, SXc-2, and SXn, were isolated, and the disrupted regions involved were identified by inverse PCR and subsequent sequencing. Both SXa-1 and SXa-2 carried a transposon insertion in lpxXL (SM0020_18047), encoding the LpxXL C-28 acyltransferase; SXc-1 and SXc-2 carried a transposon insertion in merR (SM0020_29390), encoding the regulatory activator; SXn contained a transposon insertion in omp (SM0020_18792), encoding a hypothetical outer membrane protein. The results of reverse transcriptase PCR (RT-PCR) combined with transposon gene disruptions revealed that SM0020_05862, encoding an unusual P-type ATPase, was regulated by the MerR protein. Analysis of the genome sequence showed that this P-type ATPase did not contain an N-terminal metal-binding domain or a CPC motif but rather TPCP compared with CopA from Escherichia coli. Pot experiments were carried out to determine whether growth and copper accumulation of the host plant M. lupulina were affected in the presence of the wild type or the different mutants. Soil samples were subjected to three levels of copper contamination, namely, the uncontaminated control and 47.36 and 142.08 mg/kg, and three replicates were conducted for each treatment. The results showed that the wild-type S. meliloti CCNWSX0020 enabled the host plant to grow better and accumulate copper ions. The plant dry weight and copper content of M. lupulina inoculated with the 5 copper

  10. Impact of Soil Salinity on the Structure of the Bacterial Endophytic Community Identified from the Roots of Caliph Medic (Medicago truncatula).

    PubMed

    Yaish, Mahmoud W; Al-Lawati, Abbas; Jana, Gerry Aplang; Vishwas Patankar, Himanshu; Glick, Bernard R

    2016-01-01

    In addition to being a forage crop, Caliph medic (Medicago truncatula) is also a model legume plant and is used for research focusing on the molecular characterization of the interaction between rhizobia and plants. However, the endophytic microbiome in this plant is poorly defined. Endophytic bacteria play a role in supplying plants with the basic requirements necessary for growth and development. Moreover, these bacteria also play a role in the mechanism of salinity stress adaptation in plants. As a prelude to the isolation and utilization of these bacteria in Caliph medic farming, 41 bacterial OTUs were identified in this project from within the interior of the roots of this plant by pyrosequencing of the small ribosomal subunit gene (16S rDNA) using a cultivation-independent approach. In addition, the differential abundance of these bacteria was studied following exposure of the plants to salinity stress. About 29,064 high-quality reads were obtained from the sequencing of six libraries prepared from control and salinity-treated tissues. Statistical analysis revealed that the abundance of ~70% of the OTUs was significantly (p ≤ 0.05) altered in roots that were exposed to salinity stress. Sequence analysis showed a similarity between some of the identified species and other, known, growth-promoting bacteria, marine and salt-stressed soil-borne bacteria, and nitrogen-fixing bacterial isolates. Determination of the amendments to the bacterial community due to salinity stress in Caliph medic provides a crucial step toward developing an understanding of the association of these endophytes, under salt stress conditions, in this model plant. To provide direct evidence regarding their growth promoting activity, a group of endophytic bacteria were isolated from inside of plant roots using a cultivation-dependent approach. Several of these isolates were able to produce ACC-deaminase, ammonia and IAA; and to solubilize Zn+2 and PO4-3. This data is consistent with the

  11. Genes conferring copper resistance in Sinorhizobium meliloti CCNWSX0020 also promote the growth of Medicago lupulina in copper-contaminated soil.

    PubMed

    Li, Zhefei; Ma, Zhanqiang; Hao, Xiuli; Rensing, Christopher; Wei, Gehong

    2014-03-01

    Sinorhizobium meliloti CCNWSX0020, isolated from root nodules of Medicago lupulina growing in gold mine tailings in the northwest of China, displayed both copper resistance and growth promotion of leguminous plants in copper-contaminated soil. Nevertheless, the genetic and biochemical mechanisms responsible for copper resistance in S. meliloti CCNWSX0020 remained uncharacterized. To investigate genes involved in copper resistance, an S. meliloti CCNWSX0020 Tn5 insertion library of 14,000 mutants was created. Five copper-sensitive mutants, named SXa-1, SXa-2, SXc-1, SXc-2, and SXn, were isolated, and the disrupted regions involved were identified by inverse PCR and subsequent sequencing. Both SXa-1 and SXa-2 carried a transposon insertion in lpxXL (SM0020_18047), encoding the LpxXL C-28 acyltransferase; SXc-1 and SXc-2 carried a transposon insertion in merR (SM0020_29390), encoding the regulatory activator; SXn contained a transposon insertion in omp (SM0020_18792), encoding a hypothetical outer membrane protein. The results of reverse transcriptase PCR (RT-PCR) combined with transposon gene disruptions revealed that SM0020_05862, encoding an unusual P-type ATPase, was regulated by the MerR protein. Analysis of the genome sequence showed that this P-type ATPase did not contain an N-terminal metal-binding domain or a CPC motif but rather TPCP compared with CopA from Escherichia coli. Pot experiments were carried out to determine whether growth and copper accumulation of the host plant M. lupulina were affected in the presence of the wild type or the different mutants. Soil samples were subjected to three levels of copper contamination, namely, the uncontaminated control and 47.36 and 142.08 mg/kg, and three replicates were conducted for each treatment. The results showed that the wild-type S. meliloti CCNWSX0020 enabled the host plant to grow better and accumulate copper ions. The plant dry weight and copper content of M. lupulina inoculated with the 5 copper

  12. Nod Factor Effects on Root Hair-Specific Transcriptome of Medicago truncatula: Focus on Plasma Membrane Transport Systems and Reactive Oxygen Species Networks.

    PubMed

    Damiani, Isabelle; Drain, Alice; Guichard, Marjorie; Balzergue, Sandrine; Boscari, Alexandre; Boyer, Jean-Christophe; Brunaud, Véronique; Cottaz, Sylvain; Rancurel, Corinne; Da Rocha, Martine; Fizames, Cécile; Fort, Sébastien; Gaillard, Isabelle; Maillol, Vincent; Danchin, Etienne G J; Rouached, Hatem; Samain, Eric; Su, Yan-Hua; Thouin, Julien; Touraine, Bruno; Puppo, Alain; Frachisse, Jean-Marie; Pauly, Nicolas; Sentenac, Hervé

    2016-01-01

    Root hairs are involved in water and nutrient uptake, and thereby in plant autotrophy. In legumes, they also play a crucial role in establishment of rhizobial symbiosis. To obtain a holistic view of Medicago truncatula genes expressed in root hairs and of their regulation during the first hours of the engagement in rhizobial symbiotic interaction, a high throughput RNA sequencing on isolated root hairs from roots challenged or not with lipochitooligosaccharides Nod factors (NF) for 4 or 20 h was carried out. This provided a repertoire of genes displaying expression in root hairs, responding or not to NF, and specific or not to legumes. In analyzing the transcriptome dataset, special attention was paid to pumps, transporters, or channels active at the plasma membrane, to other proteins likely to play a role in nutrient ion uptake, NF electrical and calcium signaling, control of the redox status or the dynamic reprogramming of root hair transcriptome induced by NF treatment, and to the identification of papilionoid legume-specific genes expressed in root hairs. About 10% of the root hair expressed genes were significantly up- or down-regulated by NF treatment, suggesting their involvement in remodeling plant functions to allow establishment of the symbiotic relationship. For instance, NF-induced changes in expression of genes encoding plasma membrane transport systems or disease response proteins indicate that root hairs reduce their involvement in nutrient ion absorption and adapt their immune system in order to engage in the symbiotic interaction. It also appears that the redox status of root hair cells is tuned in response to NF perception. In addition, 1176 genes that could be considered as "papilionoid legume-specific" were identified in the M. truncatula root hair transcriptome, from which 141 were found to possess an ortholog in every of the six legume genomes that we considered, suggesting their involvement in essential functions specific to legumes. This

  13. Nod Factor Effects on Root Hair-Specific Transcriptome of Medicago truncatula: Focus on Plasma Membrane Transport Systems and Reactive Oxygen Species Networks

    PubMed Central

    Damiani, Isabelle; Drain, Alice; Guichard, Marjorie; Balzergue, Sandrine; Boscari, Alexandre; Boyer, Jean-Christophe; Brunaud, Véronique; Cottaz, Sylvain; Rancurel, Corinne; Da Rocha, Martine; Fizames, Cécile; Fort, Sébastien; Gaillard, Isabelle; Maillol, Vincent; Danchin, Etienne G. J.; Rouached, Hatem; Samain, Eric; Su, Yan-Hua; Thouin, Julien; Touraine, Bruno; Puppo, Alain; Frachisse, Jean-Marie; Pauly, Nicolas; Sentenac, Hervé

    2016-01-01

    Root hairs are involved in water and nutrient uptake, and thereby in plant autotrophy. In legumes, they also play a crucial role in establishment of rhizobial symbiosis. To obtain a holistic view of Medicago truncatula genes expressed in root hairs and of their regulation during the first hours of the engagement in rhizobial symbiotic interaction, a high throughput RNA sequencing on isolated root hairs from roots challenged or not with lipochitooligosaccharides Nod factors (NF) for 4 or 20 h was carried out. This provided a repertoire of genes displaying expression in root hairs, responding or not to NF, and specific or not to legumes. In analyzing the transcriptome dataset, special attention was paid to pumps, transporters, or channels active at the plasma membrane, to other proteins likely to play a role in nutrient ion uptake, NF electrical and calcium signaling, control of the redox status or the dynamic reprogramming of root hair transcriptome induced by NF treatment, and to the identification of papilionoid legume-specific genes expressed in root hairs. About 10% of the root hair expressed genes were significantly up- or down-regulated by NF treatment, suggesting their involvement in remodeling plant functions to allow establishment of the symbiotic relationship. For instance, NF-induced changes in expression of genes encoding plasma membrane transport systems or disease response proteins indicate that root hairs reduce their involvement in nutrient ion absorption and adapt their immune system in order to engage in the symbiotic interaction. It also appears that the redox status of root hair cells is tuned in response to NF perception. In addition, 1176 genes that could be considered as “papilionoid legume-specific” were identified in the M. truncatula root hair transcriptome, from which 141 were found to possess an ortholog in every of the six legume genomes that we considered, suggesting their involvement in essential functions specific to legumes. This

  14. Impact of Soil Salinity on the Structure of the Bacterial Endophytic Community Identified from the Roots of Caliph Medic (Medicago truncatula)

    PubMed Central

    Al-Lawati, Abbas; Jana, Gerry Aplang; Vishwas Patankar, Himanshu; Glick, Bernard R.

    2016-01-01

    In addition to being a forage crop, Caliph medic (Medicago truncatula) is also a model legume plant and is used for research focusing on the molecular characterization of the interaction between rhizobia and plants. However, the endophytic microbiome in this plant is poorly defined. Endophytic bacteria play a role in supplying plants with the basic requirements necessary for growth and development. Moreover, these bacteria also play a role in the mechanism of salinity stress adaptation in plants. As a prelude to the isolation and utilization of these bacteria in Caliph medic farming, 41 bacterial OTUs were identified in this project from within the interior of the roots of this plant by pyrosequencing of the small ribosomal subunit gene (16S rDNA) using a cultivation-independent approach. In addition, the differential abundance of these bacteria was studied following exposure of the plants to salinity stress. About 29,064 high-quality reads were obtained from the sequencing of six libraries prepared from control and salinity-treated tissues. Statistical analysis revealed that the abundance of ~70% of the OTUs was significantly (p ≤ 0.05) altered in roots that were exposed to salinity stress. Sequence analysis showed a similarity between some of the identified species and other, known, growth-promoting bacteria, marine and salt-stressed soil-borne bacteria, and nitrogen-fixing bacterial isolates. Determination of the amendments to the bacterial community due to salinity stress in Caliph medic provides a crucial step toward developing an understanding of the association of these endophytes, under salt stress conditions, in this model plant. To provide direct evidence regarding their growth promoting activity, a group of endophytic bacteria were isolated from inside of plant roots using a cultivation-dependent approach. Several of these isolates were able to produce ACC-deaminase, ammonia and IAA; and to solubilize Zn+2 and PO4-3. This data is consistent with the

  15. CdSe/ZnS Quantum Dots trigger DNA repair and antioxidant enzyme systems in Medicago sativa cells in suspension culture

    PubMed Central

    2013-01-01

    Background Nanoparticles appear to be promising devices for application in the agriculture and food industries, but information regarding the response of plants to contact with nano-devices is scarce. Toxic effects may be imposed depending on the type and concentration of nanoparticle as well as time of exposure. A number of mechanisms may underlie the ability of nanoparticles to cause genotoxicity, besides the activation of ROS scavenging mechanisms. In a previous study, we showed that plant cells accumulate 3-Mercaptopropanoic acid-CdSe/ZnS quantum dots (MPA-CdSe/ZnS QD) in their cytosol and nucleus and increased production of ROS in a dose dependent manner when exposed to QD and that a concentration of 10 nM should be cyto-compatible. Results When Medicago sativa cells were exposed to 10, 50 and 100 nM MPA-CdSe/ZnS QD a correspondent increase in the activity of Superoxide dismutase, Catalase and Glutathione reductase was registered. Different versions of the COMET assay were used to assess the genotoxicity of MPA-CdSe/ZnS QD. The number of DNA single and double strand breaks increased with increasing concentrations of MPA-CdSe/ZnS QD. At the highest concentrations, tested purine bases were more oxidized than the pyrimidine ones. The transcription of the DNA repair enzymes Formamidopyrimidine DNA glycosylase, Tyrosyl-DNA phosphodiesterase I and DNA Topoisomerase I was up-regulated in the presence of increasing concentrations of MPA-CdSe/ZnS QD. Conclusions Concentrations as low as 10 nM MPA-CdSe/ZnS Quantum Dots are cytotoxic and genotoxic to plant cells, although not lethal. This sets a limit for the concentrations to be used when practical applications using nanodevices of this type on plants are being considered. This work describes for the first time the genotoxic effect of Quantum Dots in plant cells and demonstrates that both the DNA repair genes (Tdp1β, Top1β and Fpg) and the ROS scavenging mechanisms are activated when MPA-CdSe/ZnS QD contact M. sativa

  16. Effects of Salt Stress on Amino Acid, Organic Acid, and Carbohydrate Composition of Roots, Bacteroids, and Cytosol of Alfalfa (Medicago sativa L.) 1

    PubMed Central

    Fougère, Francoise; Le Rudulier, Daniel; Streeter, John G.

    1991-01-01

    Ethanol-soluble organic acid, carbohydrate, and amino acid constituents of alfalfa (Medicago sativa) roots and nodules (cytosol and bacteroids) have been identified by gas-liquid chromatography and high performance liquid chromatography. Among organic acids, citrate was the predominant compound in roots and cytosol, with malonate present in the highest concentration in bacteroids. These two organic acids together with malate and succinate accounted for more than 85% of the organic acid pool in nodules and for 97% in roots. The major carbohydrates in roots, nodule cytosol, and bacteroids were (descending order of concentration): sucrose, pinitol, glucose, and ononitol. Maltose and trehalose appeared to be present in very low concentrations. Asparagine, glutamate, alanine, γ-aminobutyrate, and proline were the major amino acids in cytosol and bacteroids. In addition to these solutes, serine and glutamine were well represented in roots. When alfalfa plants were subjected to 0.15 m sodium chloride stress for 2 weeks, total organic acid concentration in nodules and roots were depressed by more than 40%, whereas lactate concentration increased by 11, 27, and 94% in cytosol, roots, and bacteroids, respectively. In bacteroids, lactate became the most abundant organic acid and might contribute partly to the osmotic adjustment. On the other hand, salt stress induced a large increase in the amino acid and carbohydrate pools. Within the amino acids, proline showed the largest increase, 11.3-, 12.8-, and 8.0-fold in roots, cytosol, and bacteroids, respectively. Its accumulation reflected an osmoregulatory mechanism not only in roots but also in nodule tissue. In parallel, asparagine concentration was greatly enhanced; this amide remained the major nitrogen solute and, in bacteroids, played a significant role in osmoregulation. On the contrary, the salt treatment had a very limited effect on the concentration of other amino acids. Among carbohydrates, pinitol concentration was

  17. Quantitative Resistance to Verticillium Wilt in Medicago truncatula Involves Eradication of the Fungus from Roots and Is Associated with Transcriptional Responses Related to Innate Immunity

    PubMed Central

    Toueni, Maoulida; Ben, Cécile; Le Ru, Aurélie; Gentzbittel, Laurent; Rickauer, Martina

    2016-01-01

    Resistance mechanisms to Verticillium wilt are well-studied in tomato, cotton, and Arabidopsis, but much less in legume plants. Because legume plants establish nitrogen-fixing symbioses in their roots, resistance to root-attacking pathogens merits particular attention. The interaction between the soil-borne pathogen Verticillium alfalfae and the model legume Medicago truncatula was investigated using a resistant (A17) and a susceptible (F83005.5) line. As shown by histological analyses, colonization by the pathogen was initiated similarly in both lines. Later on, the resistant line A17 eliminated the fungus, whereas the susceptible F83005.5 became heavily colonized. Resistance in line A17 does not involve homologs of the well-characterized tomato Ve1 and V. dahliae Ave1 genes. A transcriptomic study of early root responses during initial colonization (i.e., until 24 h post-inoculation) similarly was performed. Compared to the susceptible line, line A17 displayed already a significantly higher basal expression of defense-related genes prior to inoculation, and responded to infection with up-regulation of only a small number of genes. Although fungal colonization was still low at this stage, the susceptible line F83005.5 exhibited a disorganized response involving a large number of genes from different functional classes. The involvement of distinct phytohormone signaling pathways in resistance as suggested by gene expression patterns was supported by experiments with plant hormone pretreatment before fungal inoculation. Gene co-expression network analysis highlighted five main modules in the resistant line, whereas no structured gene expression was found in the susceptible line. One module was particularly associated to the inoculation response in A17. It contains the majority of differentially expressed genes, genes associated with PAMP perception and hormone signaling, and transcription factors. An in silico analysis showed that a high number of these genes also

  18. Subcellular localisation of Medicago truncatula 9/13-hydroperoxide lyase reveals a new localisation pattern and activation mechanism for CYP74C enzymes

    PubMed Central

    De Domenico, Stefania; Tsesmetzis, Nicolas; Di Sansebastiano, Gian Pietro; Hughes, Richard K; Casey, Rod; Santino, Angelo

    2007-01-01

    Background Hydroperoxide lyase (HPL) is a key enzyme in plant oxylipin metabolism that catalyses the cleavage of polyunsaturated fatty acid hydroperoxides produced by the action of lipoxygenase (LOX) to volatile aldehydes and oxo acids. The synthesis of these volatile aldehydes is rapidly induced in plant tissues upon mechanical wounding and insect or pathogen attack. Together with their direct defence role towards different pathogens, these compounds are believed to play an important role in signalling within and between plants, and in the molecular cross-talk between plants and other organisms surrounding them. We have recently described the targeting of a seed 9-HPL to microsomes and putative lipid bodies and were interested to compare the localisation patterns of both a 13-HPL and a 9/13-HPL from Medicago truncatula, which were known to be expressed in leaves and roots, respectively. Results To study the subcellular localisation of plant 9/13-HPLs, a set of YFP-tagged chimeric constructs were prepared using two M. truncatula HPL cDNAs and the localisation of the corresponding chimeras were verified by confocal microscopy in tobacco protoplasts and leaves. Results reported here indicated a distribution of M.truncatula 9/13-HPL (HPLF) between cytosol and lipid droplets (LD) whereas, as expected, M.truncatula 13-HPL (HPLE) was targeted to plastids. Notably, such endocellular localisation has not yet been reported previously for any 9/13-HPL. To verify a possible physiological significance of such association, purified recombinant HPLF was used in activation experiments with purified seed lipid bodies. Our results showed that lipid bodies can fully activate HPLF. Conclusion We provide evidence for the first CYP74C enzyme, to be targeted to cytosol and LD. We also showed by sedimentation and kinetic analyses that the association with LD or lipid bodies can result in the protein conformational changes required for full activation of the enzyme. This activation

  19. Nod Factor Effects on Root Hair-Specific Transcriptome of Medicago truncatula: Focus on Plasma Membrane Transport Systems and Reactive Oxygen Species Networks.

    PubMed

    Damiani, Isabelle; Drain, Alice; Guichard, Marjorie; Balzergue, Sandrine; Boscari, Alexandre; Boyer, Jean-Christophe; Brunaud, Véronique; Cottaz, Sylvain; Rancurel, Corinne; Da Rocha, Martine; Fizames, Cécile; Fort, Sébastien; Gaillard, Isabelle; Maillol, Vincent; Danchin, Etienne G J; Rouached, Hatem; Samain, Eric; Su, Yan-Hua; Thouin, Julien; Touraine, Bruno; Puppo, Alain; Frachisse, Jean-Marie; Pauly, Nicolas; Sentenac, Hervé

    2016-01-01

    Root hairs are involved in water and nutrient uptake, and thereby in plant autotrophy. In legumes, they also play a crucial role in establishment of rhizobial symbiosis. To obtain a holistic view of Medicago truncatula genes expressed in root hairs and of their regulation during the first hours of the engagement in rhizobial symbiotic interaction, a high throughput RNA sequencing on isolated root hairs from roots challenged or not with lipochitooligosaccharides Nod factors (NF) for 4 or 20 h was carried out. This provided a repertoire of genes displaying expression in root hairs, responding or not to NF, and specific or not to legumes. In analyzing the transcriptome dataset, special attention was paid to pumps, transporters, or channels active at the plasma membrane, to other proteins likely to play a role in nutrient ion uptake, NF electrical and calcium signaling, control of the redox status or the dynamic reprogramming of root hair transcriptome induced by NF treatment, and to the identification of papilionoid legume-specific genes expressed in root hairs. About 10% of the root hair expressed genes were significantly up- or down-regulated by NF treatment, suggesting their involvement in remodeling plant functions to allow establishment of the symbiotic relationship. For instance, NF-induced changes in expression of genes encoding plasma membrane transport systems or disease response proteins indicate that root hairs reduce their involvement in nutrient ion absorption and adapt their immune system in order to engage in the symbiotic interaction. It also appears that the redox status of root hair cells is tuned in response to NF perception. In addition, 1176 genes that could be considered as "papilionoid legume-specific" were identified in the M. truncatula root hair transcriptome, from which 141 were found to possess an ortholog in every of the six legume genomes that we considered, suggesting their involvement in essential functions specific to legumes. This

  20. Genes Conferring Copper Resistance in Sinorhizobium meliloti CCNWSX0020 Also Promote the Growth of Medicago lupulina in Copper-Contaminated Soil

    PubMed Central

    Li, Zhefei; Ma, Zhanqiang; Hao, Xiuli; Rensing, Christopher

    2014-01-01

    Sinorhizobium meliloti CCNWSX0020, isolated from root nodules of Medicago lupulina growing in gold mine tailings in the northwest of China, displayed both copper resistance and growth promotion of leguminous plants in copper-contaminated soil. Nevertheless, the genetic and biochemical mechanisms responsible for copper resistance in S. meliloti CCNWSX0020 remained uncharacterized. To investigate genes involved in copper resistance, an S. meliloti CCNWSX0020 Tn5 insertion library of 14,000 mutants was created. Five copper-sensitive mutants, named SXa-1, SXa-2, SXc-1, SXc-2, and SXn, were isolated, and the disrupted regions involved were identified by inverse PCR and subsequent sequencing. Both SXa-1 and SXa-2 carried a transposon insertion in lpxXL (SM0020_18047), encoding the LpxXL C-28 acyltransferase; SXc-1 and SXc-2 carried a transposon insertion in merR (SM0020_29390), encoding the regulatory activator; SXn contained a transposon insertion in omp (SM0020_18792), encoding a hypothetical outer membrane protein. The results of reverse transcriptase PCR (RT-PCR) combined with transposon gene disruptions revealed that SM0020_05862, encoding an unusual P-type ATPase, was regulated by the MerR protein. Analysis of the genome sequence showed that this P-type ATPase did not contain an N-terminal metal-binding domain or a CPC motif but rather TPCP compared with CopA from Escherichia coli. Pot experiments were carried out to determine whether growth and copper accumulation of the host plant M. lupulina were affected in the presence of the wild type or the different mutants. Soil samples were subjected to three levels of copper contamination, namely, the uncontaminated control and 47.36 and 142.08 mg/kg, and three replicates were conducted for each treatment. The results showed that the wild-type S. meliloti CCNWSX0020 enabled the host plant to grow better and accumulate copper ions. The plant dry weight and copper content of M. lupulina inoculated with the 5 copper

  1. The B-3 ethylene response factor MtERF1-1 mediates resistance to a subset of root pathogens in Medicago truncatula without adversely affecting symbiosis with rhizobia.

    PubMed

    Anderson, Jonathan P; Lichtenzveig, Judith; Gleason, Cynthia; Oliver, Richard P; Singh, Karam B

    2010-10-01

    The fungal necrotrophic pathogen Rhizoctonia solani is a significant constraint to a range of crops as diverse as cereals, canola, and legumes. Despite wide-ranging germplasm screens in many of these crops, no strong genetic resistance has been identified, suggesting that alternative strategies to improve resistance are required. In this study, we characterize moderate resistance to R. solani anastomosis group 8 identified in Medicago truncatula. The activity of the ethylene- and jasmonate-responsive GCC box promoter element was associated with moderate resistance, as was the induction of the B-3 subgroup of ethylene response transcription factors (ERFs). Genes of the B-1 subgroup showed no significant response to R. solani infection. Overexpression of a B-3 ERF, MtERF1-1, in Medicago roots increased resistance to R. solani as well as an oomycete root pathogen, Phytophthora medicaginis, but not root knot nematode. These results indicate that targeting specific regulators of ethylene defense may enhance resistance to an important subset of root pathogens. We also demonstrate that overexpression of MtERF1-1 enhances disease resistance without apparent impact on nodulation in the A17 background, while overexpression in sickle reduced the hypernodulation phenotype. This suggests that under normal regulation of nodulation, enhanced resistance to root diseases can be uncoupled from symbiotic plant-microbe interactions in the same tissue and that ethylene/ERF regulation of nodule number is distinct from the defenses regulated by B-3 ERFs. Furthermore, unlike the stunted phenotype previously described for Arabidopsis (Arabidopsis thaliana) ubiquitously overexpressing B-3 ERFs, overexpression of MtERF1-1 in M. truncatula roots did not show adverse effects on plant development.

  2. Henrique da Rocha Lima*

    PubMed Central

    Bernardes Filho, Fred; Avelleira, João Carlos Regazzi

    2015-01-01

    Brazilian physician and researcher Henrique da Rocha Lima was born in 1879 in the city of Rio de Janeiro, where he studied medicine and obtained the degree of M.D. in 1901. He specialized in Clinical Medicine in Germany and was the ambassador in European countries of the scientific medicine that emerged from the Oswaldo Cruz Institute in the early twentieth century. Rocha Lima has discovered the causative agent of typhus and had a major contribution to the studies of yellow fever, Chagas disease, Carrión’s disease and histoplasmosis. His genius, his research and his discoveries projected his name, and, with it, the image of Brazil in the international scientific scene. PMID:26131867

  3. Phytotoxicity of three plant-based biodiesels, unmodified castor oil, and Diesel fuel to alfalfa (Medicago sativa L.), lettuce (Lactuca sativa L.), radish (Raphanus sativus), and wheatgrass (Triticum aestivum).

    PubMed

    Bamgbose, Ifeoluwa; Anderson, Todd A

    2015-12-01

    The wide use of plant-based oils and their derivatives, in particular biodiesel, have increased extensively over the past decade to help alleviate demand for petroleum products and improve the greenhouse gas emissions profile of the transportation sector. Biodiesel is regarded as a clean burning alternative fuel produced from livestock feeds and various vegetable oils. Although in theory these animal and/or plant derived fuels should have less environmental impact in soil based on their simplified composition relative to Diesel, they pose an environmental risk like Diesel at high concentrations when disposed. The aim of the present study was to ascertain the phytotoxicity of three different plant-derived biodiesels relative to conventional Diesel. For phytotoxicological analysis, we used seeds of four crop plants, Medicago sativa, Lactuca sativa, Raphanus sativus, and Triticum aestivum to analyze the germination of seeds in contaminated soil samples. The toxicological experiment was conducted with two different soil textures: sandy loam soil and silt loam soil. The studied plant-based biodiesels were safflower methyl-ester, castor methyl ester, and castor ethyl-ester. Biodiesel toxicity was more evident at high concentrations, affecting the germination and survival of small-seeded plants to a greater extent. Tolerance of plants to the biodiesels varied between plant species and soil textures. With the exception of R. sativus, all plant species were affected and exhibited some sensitivity to the fuels, such as delayed seedling emergence and slow germination (average=10 days) at high soil concentrations (0.85% for Diesel and 1.76% for the biodiesels). Tolerance of plants to soil contamination had a species-specific nature, and on average, decreased in the following order: Raphanus sativus (0-20%)>Triticum aestivum (10-40%) ≥ Medicago sativa> Lactuca sativa (80-100%). Thus, we conclude that there is some phytotoxicity associated with plant-based biodiesels. Further

  4. Phytotoxicity of three plant-based biodiesels, unmodified castor oil, and Diesel fuel to alfalfa (Medicago sativa L.), lettuce (Lactuca sativa L.), radish (Raphanus sativus), and wheatgrass (Triticum aestivum).

    PubMed

    Bamgbose, Ifeoluwa; Anderson, Todd A

    2015-12-01

    The wide use of plant-based oils and their derivatives, in particular biodiesel, have increased extensively over the past decade to help alleviate demand for petroleum products and improve the greenhouse gas emissions profile of the transportation sector. Biodiesel is regarded as a clean burning alternative fuel produced from livestock feeds and various vegetable oils. Although in theory these animal and/or plant derived fuels should have less environmental impact in soil based on their simplified composition relative to Diesel, they pose an environmental risk like Diesel at high concentrations when disposed. The aim of the present study was to ascertain the phytotoxicity of three different plant-derived biodiesels relative to conventional Diesel. For phytotoxicological analysis, we used seeds of four crop plants, Medicago sativa, Lactuca sativa, Raphanus sativus, and Triticum aestivum to analyze the germination of seeds in contaminated soil samples. The toxicological experiment was conducted with two different soil textures: sandy loam soil and silt loam soil. The studied plant-based biodiesels were safflower methyl-ester, castor methyl ester, and castor ethyl-ester. Biodiesel toxicity was more evident at high concentrations, affecting the germination and survival of small-seeded plants to a greater extent. Tolerance of plants to the biodiesels varied between plant species and soil textures. With the exception of R. sativus, all plant species were affected and exhibited some sensitivity to the fuels, such as delayed seedling emergence and slow germination (average=10 days) at high soil concentrations (0.85% for Diesel and 1.76% for the biodiesels). Tolerance of plants to soil contamination had a species-specific nature, and on average, decreased in the following order: Raphanus sativus (0-20%)>Triticum aestivum (10-40%) ≥ Medicago sativa> Lactuca sativa (80-100%). Thus, we conclude that there is some phytotoxicity associated with plant-based biodiesels. Further

  5. ABA-Mediated Inhibition of Germination Is Related to the Inhibition of Genes Encoding Cell-Wall Biosynthetic and Architecture: Modifying Enzymes and Structural Proteins in Medicago truncatula Embryo Axis

    PubMed Central

    Gimeno-Gilles, Christine; Lelièvre, Eric; Viau, Laure; Malik-Ghulam, Mustafa; Ricoult, Claudie; Niebel, Andreas; Leduc, Nathalie; Limami, Anis M.

    2009-01-01

    Radicle emergence and reserves mobilization are two distinct programmes that are thought to control germination. Both programs are influenced by abscissic acid (ABA) but how this hormone controls seed germination is still poorly known. Phenotypic and microscopic observations of the embryo axis of Medicago truncatula during germination in mitotic inhibition condition triggered by 10 μM oryzalin showed that cell division was not required to allow radicle emergence. A suppressive subtractive hybridization showed that more than 10% of up-regulated genes in the embryo axis encoded proteins related to cell-wall biosynthesis. The expression of α-expansins, pectin-esterase, xylogucan-endotransglycosidase, cellulose synthase, and extensins was monitored in the embryo axis of seeds germinated on water, constant and transitory ABA. These genes were overexpressed before completion of germination in the control and strongly inhibited by ABA. The expression was re-established in the ABA transitory-treatment after the seeds were transferred back on water and proceeded to germination. This proves these genes as contributors to the completion of germination and strengthen the idea that cell-wall loosening and remodeling in relation to cell expansion in the embryo axis is a determinant feature in germination. Our results also showed that ABA controls germination through the control of radicle emergence, namely by inhibiting cell-wall loosening and expansion. PMID:19529818

  6. The C2H2 Transcription Factor REGULATOR OF SYMBIOSOME DIFFERENTIATION Represses Transcription of the Secretory Pathway Gene VAMP721a and Promotes Symbiosome Development in Medicago truncatula[W][OPEN

    PubMed Central

    Sinharoy, Senjuti; Torres-Jerez, Ivone; Bandyopadhyay, Kaustav; Kereszt, Attila; Pislariu, Catalina I.; Nakashima, Jin; Benedito, Vagner A.; Kondorosi, Eva; Udvardi, Michael K.

    2013-01-01

    Transcription factors (TFs) are thought to regulate many aspects of nodule and symbiosis development in legumes, although few TFs have been characterized functionally. Here, we describe REGULATOR OF SYMBIOSOME DIFFERENTIATION (RSD) of Medicago truncatula, a member of the Cysteine-2/Histidine-2 (C2H2) family of plant TFs that is required for normal symbiosome differentiation during nodule development. RSD is expressed in a nodule-specific manner, with maximal transcript levels in the bacterial invasion zone. A tobacco (Nicotiana tabacum) retrotransposon (Tnt1) insertion rsd mutant produced nodules that were unable to fix nitrogen and that contained incompletely differentiated symbiosomes and bacteroids. RSD protein was localized to the nucleus, consistent with a role of the protein in transcriptional regulation. RSD acted as a transcriptional repressor in a heterologous yeast assay. Transcriptome analysis of an rsd mutant identified 11 genes as potential targets of RSD repression. RSD interacted physically with the promoter of one of these genes, VAMP721a, which encodes vesicle-associated membrane protein 721a. Thus, RSD may influence symbiosome development in part by repressing transcription of VAMP721a and modifying vesicle trafficking in nodule cells. This establishes RSD as a TF implicated directly in symbiosome and bacteroid differentiation and a transcriptional regulator of secretory pathway genes in plants. PMID:24082011

  7. Nodule-enhanced expression of a sucrose phosphate synthase gene member (MsSPSA) has a role in carbon and nitrogen metabolism in the nodules of alfalfa (Medicago sativa L.).

    PubMed

    Aleman, Lorenzo; Ortega, Jose Luis; Martinez-Grimes, Martha; Seger, Mark; Holguin, Francisco Omar; Uribe, Diana J; Garcia-Ibilcieta, David; Sengupta-Gopalan, Champa

    2010-01-01

    Sucrose phosphate synthase (SPS) catalyzes the first step in the synthesis of sucrose in photosynthetic tissues. We characterized the expression of three different isoforms of SPS belonging to two different SPS gene families in alfalfa (Medicago sativa L.), a previously identified SPS (MsSPSA) and two novel isoforms belonging to class B (MsSPSB and MsSPSB3). While MsSPSA showed nodule-enhanced expression, both MsSPSB genes exhibited leaf-enhanced expression. Alfalfa leaf and nodule SPS enzymes showed differences in chromatographic and electrophoretic migration and differences in V (max) and allosteric regulation. The root nodules in legume plants are a strong sink for photosynthates with its need for ATP, reducing power and carbon skeletons for dinitrogen fixation and ammonia assimilation. The expression of genes encoding SPS and other key enzymes in sucrose metabolism, sucrose phosphate phosphatase and sucrose synthase, was analyzed in the leaves and nodules of plants inoculated with Sinorhizobium meliloti. Based on the expression pattern of these genes, the properties of the SPS isoforms and the concentration of starch and soluble sugars in nodules induced by a wild type and a nitrogen fixation deficient strain, we propose that SPS has an important role in the control of carbon flux into different metabolic pathways in the symbiotic nodules.

  8. Leonardo da Vinci and the Downburst.

    NASA Astrophysics Data System (ADS)

    Gedzelman, Stanley David

    1990-05-01

    Evidence from the drawings, experiments, and writings of Leonardo da Vinci are presented to demonstrate that da Vinci recognized and, possibly, discovered the downburst and understood its associated airflow. Other early references to vortex flows resembling downbursts are mentioned.

  9. Gene-Silencing-Induced Changes in Carbohydrate Conformation in Relation to Bioenergy Value and Carbohydrate Subfractions in Modeled Plant (Medicago sativa) with Down-Regulation of HB12 and TT8 Transcription Factors

    PubMed Central

    Li, Xinxin; Hannoufa, Abdelali; Zhang, Yonggen; Yu, Peiqiang

    2016-01-01

    Gene silencing with RNA interference (RNAi) technology may be capable of modifying internal structure at a molecular level. This structural modification could affect biofunctions in terms of biodegradation, biochemical metabolism, and bioactive compound availability. The objectives of this study were to (1) Detect gene silencing-induced changes in carbohydrate molecular structure in an alfalfa forage (Medicago sativa spp. sativa: alfalfa) with down-regulation of genes that encode transcription factors TT8 and HB12; (2) Determine gene silencing-induced changes in nutrient bioutilization and bioavailability in the alfalfa forage (Medicago sativa); and (3) Quantify the correlation between gene silencing-induced molecular structure changes and the nutrient bioutilization and bioavailability in animals of ruminants. The experimental treatments included: T1 = Non-transgenic and no-gene silenced alfalfa forage (code “NT”); T2 = HB12-RNAi forage with HB12 gene down regulation (code “HB12”); T3 = TT8-RNAi forage with TT8 gene down regulation (code “TT8”). The HB12 and TT8 gene silencing-induced molecular structure changes were determined by non-invasive and non-destructive advanced molecular spectroscopy in a middle infrared radiation region that focused on structural, non-structural and total carbohydrate compounds. The nutrient bioutilization and bioavailability of the modified forage were determined using NRC-2001 system in terms of total digestive nutrient (TDN), truly digestible fiber (tdNDF), non-fiber carbohydrate (tdNDF), fatty acid (tdFA), crude protein (tdCP) and bioenergy profiles (digestible energy, metabolizable energy, net energy) for ruminants. The carbohydrate subfractions were evaluated using the updated CNCPS 6.0 system. The results showed that gene silencing significantly affected tdNFC (42.3 (NT) vs. 38.7 (HB12) vs. 37.4% Dry Matter (TT8); p = 0.016) and tdCP (20.8 (NT) vs. 19.4 (HB12) vs. 22.3% DM (TT8); p = 0.009). The gene-silencing also

  10. Gene-Silencing-Induced Changes in Carbohydrate Conformation in Relation to Bioenergy Value and Carbohydrate Subfractions in Modeled Plant (Medicago sativa) with Down-Regulation of HB12 and TT8 Transcription Factors.

    PubMed

    Li, Xinxin; Hannoufa, Abdelali; Zhang, Yonggen; Yu, Peiqiang

    2016-05-13

    Gene silencing with RNA interference (RNAi) technology may be capable of modifying internal structure at a molecular level. This structural modification could affect biofunctions in terms of biodegradation, biochemical metabolism, and bioactive compound availability. The objectives of this study were to (1) Detect gene silencing-induced changes in carbohydrate molecular structure in an alfalfa forage (Medicago sativa spp. sativa: alfalfa) with down-regulation of genes that encode transcription factors TT8 and HB12; (2) Determine gene silencing-induced changes in nutrient bioutilization and bioavailability in the alfalfa forage (Medicago sativa); and (3) Quantify the correlation between gene silencing-induced molecular structure changes and the nutrient bioutilization and bioavailability in animals of ruminants. The experimental treatments included: T1 = Non-transgenic and no-gene silenced alfalfa forage (code "NT"); T2 = HB12-RNAi forage with HB12 gene down regulation (code "HB12"); T3 = TT8-RNAi forage with TT8 gene down regulation (code "TT8"). The HB12 and TT8 gene silencing-induced molecular structure changes were determined by non-invasive and non-destructive advanced molecular spectroscopy in a middle infrared radiation region that focused on structural, non-structural and total carbohydrate compounds. The nutrient bioutilization and bioavailability of the modified forage were determined using NRC-2001 system in terms of total digestive nutrient (TDN), truly digestible fiber (tdNDF), non-fiber carbohydrate (tdNDF), fatty acid (tdFA), crude protein (tdCP) and bioenergy profiles (digestible energy, metabolizable energy, net energy) for ruminants. The carbohydrate subfractions were evaluated using the updated CNCPS 6.0 system. The results showed that gene silencing significantly affected tdNFC (42.3 (NT) vs. 38.7 (HB12) vs. 37.4% Dry Matter (TT8); p = 0.016) and tdCP (20.8 (NT) vs. 19.4 (HB12) vs. 22.3% DM (TT8); p = 0.009). The gene-silencing also affected

  11. Gene-Silencing-Induced Changes in Carbohydrate Conformation in Relation to Bioenergy Value and Carbohydrate Subfractions in Modeled Plant (Medicago sativa) with Down-Regulation of HB12 and TT8 Transcription Factors.

    PubMed

    Li, Xinxin; Hannoufa, Abdelali; Zhang, Yonggen; Yu, Peiqiang

    2016-01-01

    Gene silencing with RNA interference (RNAi) technology may be capable of modifying internal structure at a molecular level. This structural modification could affect biofunctions in terms of biodegradation, biochemical metabolism, and bioactive compound availability. The objectives of this study were to (1) Detect gene silencing-induced changes in carbohydrate molecular structure in an alfalfa forage (Medicago sativa spp. sativa: alfalfa) with down-regulation of genes that encode transcription factors TT8 and HB12; (2) Determine gene silencing-induced changes in nutrient bioutilization and bioavailability in the alfalfa forage (Medicago sativa); and (3) Quantify the correlation between gene silencing-induced molecular structure changes and the nutrient bioutilization and bioavailability in animals of ruminants. The experimental treatments included: T1 = Non-transgenic and no-gene silenced alfalfa forage (code "NT"); T2 = HB12-RNAi forage with HB12 gene down regulation (code "HB12"); T3 = TT8-RNAi forage with TT8 gene down regulation (code "TT8"). The HB12 and TT8 gene silencing-induced molecular structure changes were determined by non-invasive and non-destructive advanced molecular spectroscopy in a middle infrared radiation region that focused on structural, non-structural and total carbohydrate compounds. The nutrient bioutilization and bioavailability of the modified forage were determined using NRC-2001 system in terms of total digestive nutrient (TDN), truly digestible fiber (tdNDF), non-fiber carbohydrate (tdNDF), fatty acid (tdFA), crude protein (tdCP) and bioenergy profiles (digestible energy, metabolizable energy, net energy) for ruminants. The carbohydrate subfractions were evaluated using the updated CNCPS 6.0 system. The results showed that gene silencing significantly affected tdNFC (42.3 (NT) vs. 38.7 (HB12) vs. 37.4% Dry Matter (TT8); p = 0.016) and tdCP (20.8 (NT) vs. 19.4 (HB12) vs. 22.3% DM (TT8); p = 0.009). The gene-silencing also affected

  12. The structure of Medicago truncatula δ1-pyrroline-5-carboxylate reductase provides new insights into regulation of proline biosynthesis in plants

    SciTech Connect

    Ruszkowski, Milosz; Nocek, Boguslaw; Forlani, Giuseppe; Dauter, Zbigniew

    2015-10-30

    The two pathways for proline biosynthesis in higher plants share the last step, the conversion of δ1-pyrroline-5-carboxylate (P5C) to L-proline, which is catalyzed by P5C reductase (P5CR, EC 1.5.1.2) with the use of NAD(P)H as a coenzyme. There is increasing amount of evidence to suggest a complex regulation of P5CR activity at the post-translational level, yet the molecular basis of these mechanisms is unknown. Here we report the three-dimensional structure of the P5CR enzyme from the model legume Medicago truncatula (Mt). The crystal structures of unliganded MtP5CR decamer, and its complexes with the products NAD+, NADP+, and L-proline were refined using x-ray diffraction data (at 1.7, 1.85, 1.95, and 2.1 Å resolution, respectively). Based on the presented structural data, the coenzyme preference for NADPH over NADH was explained, and NADPH is suggested to be the only coenzyme used by MtP5CR in vivo. Moreover, the insensitivity of MtP5CR to feed-back inhibition by proline, revealed by enzymatic analysis, was correlated with structural features. Additionally, a mechanism for the modulation of enzyme activity by chloride anions is discussed, as well as the rationale for the possible development of effective enzyme inhibitors.

  13. Stress Responses in Alfalfa (Medicago sativa L.) (XIV. Changes in the Levels of Phenylpropanoid Pathway Intermediates in Relation to Regulation of L-Phenylalanine Ammonia-Lyase in Elicitor-Treated Cell-Suspension Cultures).

    PubMed Central

    Orr, J. D.; Edwards, R.; Dixon, R. A.

    1993-01-01

    We have used high-resolution gas chromatography to determine the levels of trans-cinnamic acid (CA) and trans-4-coumaric acid (4CA) in alfalfa (Medicago sativa L.) cell-suspension cultures to address the role of these phenylpropanoid pathway intermediates as potential negative regulators of phenylalanine ammonia-lyase (PAL) in vivo. Exogenous addition of CA to elicitor-treated cultures resulted in rapid increases in endogenous CA, 4CA, and CA-conjugate levels associated with inhibition of the appearance of PAL transcripts. Treatment of elicited cultures with [alpha]-aminooxy-[beta]-phenylpropionic acid (AOPP), a potent and specific inhibitor of PAL activity in vivo, resulted in reductions of CA and 4CA, with concomitant increases in PAL transcripts and extractable enzyme activity. In contrast, treatment with tetcyclacis, an inhibitor of CA 4-hydroxylase, resulted in increased CA and CA-conjugate levels, decreased 4CA levels, and decreased PAL transcript levels and enzyme activity. In tetcyclasis-treated cells, the inhibition of PAL transcript appearance preceded the increase in the levels of free CA and its conjugates. In elicited cells in which the phenylpropanoid pathway was not perturbed by metabolic inhibitors, PAL transcripts accumulated rapidly and transiently, beginning to decline by 2 h postelicitation. Changes in levels of total free or conjugated CA or 4CA did not consistently correlate with these changes in transcript levels. We propose that regulation of PAL transcript levels by endogenous phenylpropanoid pathway intermediates could involve compartmentalized pools that may exist because of the microsomal localization of cinnamic acid 4-hydroxylase. PMID:12231735

  14. Characterization of Medicago truncatula (barrel medic) hydroperoxide lyase (CYP74C3), a water-soluble detergent-free cytochrome P450 monomer whose biological activity is defined by monomer–micelle association

    PubMed Central

    Hughes, Richard K.; Belfield, Eric J.; Muthusamay, Mylrajan; Khan, Anuja; Rowe, Arthur; Harding, Stephen E.; Fairhurst, Shirley A.; Bornemann, Stephen; Ashton, Ruth; Thorneley, Roger N. F.; Casey, Rod

    2006-01-01

    We describe the detailed biochemical characterization of CYP74C3 (cytochrome P450 subfamily 74C3), a recombinant plant cytochrome P450 enzyme with HPL (hydroperoxide lyase) activity from Medicago truncatula (barrel medic). Steady-state kinetic parameters, substrate and product specificities, RZ (Reinheitszahl or purity index), molar absorption coefficient, haem content, and new ligands for an HPL are reported. We show on the basis of gel filtration, sedimentation velocity (sedimentation coefficient distribution) and sedimentation equilibrium (molecular mass) analyses that CYP74C3 has low enzyme activity as a detergent-free, water-soluble, monomer. The enzyme activity can be completely restored by re-activation with detergent micelles, but not detergent monomers. Corresponding changes in the spin state equilibrium, and probably co-ordination of the haem iron, are novel for cytochrome P450 enzymes and suggest that detergent micelles have a subtle effect on protein conformation, rather than substrate presentation, which is sufficient to improve substrate binding and catalytic-centre activity by an order of magnitude. The kcat/Km of up to 1.6×108 M−1·s−1 is among the highest recorded, which is remarkable for an enzyme whose reaction mechanism involves the scission of a C–C bond. We carried out both kinetic and biophysical studies to demonstrate that this effect is a result of the formation of a complex between a protein monomer and a single detergent micelle. Association with a detergent micelle rather than oligomeric state represents a new mechanism of activation for membrane-associated cytochrome P450 enzymes. Highly concentrated and monodispersed samples of detergent-free CYP74C3 protein may be well suited for the purposes of crystallization and structural resolution of the first plant cytochrome P450 enzyme. PMID:16454766

  15. Toxicogenomic Responses of the Model Legume Medicago truncatula to Aged Biosolids Containing a Mixture of Nanomaterials (TiO₂, Ag, and ZnO) from a Pilot Wastewater Treatment Plant.

    PubMed

    Chen, Chun; Unrine, Jason M; Judy, Jonathan D; Lewis, Ricky W; Guo, Jing; McNear, David H; Tsyusko, Olga V

    2015-07-21

    Toxicogenomic responses in Medicago truncatula A17 were monitored following exposure to biosolids-amended soils. Treatments included biosolids produced using a pilot wastewater treatment plant with either no metal introduced into the influent (control); bulk/ionic TiO2, ZnO, and AgNO3 added to influent (bulk/dissolved treatment); or Ag, ZnO, and TiO2 engineered nanomaterials added to influent (ENM treatment) and then added to soil, which was aged in the field for 6 months. In our companion study, we found inhibition of nodulation in the ENM but not in the bulk/dissolved treatment. Gene expression profiling revealed highly distinct profiles with more than 10-fold down-regulation in 239 genes in M. truncatula roots from the ENM treatment, while gene expression patterns were similar between bulk/dissolved and control treatments. In response to ENM exposure, many of the identified biological pathways, gene ontologies, and individual genes are associated with nitrogen metabolism, nodulation, metal homeostasis, and stress responses. Expression levels of nine genes were independently confirmed with qRT-PCR. Exposure to ENMs induced unique shifts in expression profiles and biological pathways compared with bulk/dissolved treatment, despite the lack of difference in bioavailable metal fractions, metal oxidation state, and coordination environment between ENM and bulk/dissolved biosolids. As populations of Sinorhizobium meliloti Rm2011 were similar in bulk/dissolved and ENM treatments, our results suggest that inhibition of nodulation in the ENM treatment was primarily due to phytotoxicity, likely caused by enhanced bioavailability of Zn ions. PMID:26065335

  16. An Oxalyl-CoA Dependent Pathway of Oxalate Catabolism Plays a Role in Regulating Calcium Oxalate Crystal Accumulation and Defending against Oxalate-Secreting Phytopathogens in Medicago truncatula

    PubMed Central

    Foster, Justin; Luo, Bin; Nakata, Paul A.

    2016-01-01

    Considering the widespread occurrence of oxalate in nature and its broad impact on a host of organisms, it is surprising that so little is known about the turnover of this important acid. In plants, oxalate oxidase is the most well studied enzyme capable of degrading oxalate, but not all plants possess this activity. Recently, an Acyl Activating Enzyme 3 (AAE3), encoding an oxalyl-CoA synthetase, was identified in Arabidopsis. AAE3 has been proposed to catalyze the first step in an alternative pathway of oxalate degradation. Whether this enzyme and proposed pathway is important to other plants is unknown. Here, we identify the Medicago truncatula AAE3 (MtAAE3) and show that it encodes an oxalyl-CoA synthetase activity exhibiting high activity against oxalate with a Km = 81 ± 9 μM and Vmax = 19 ± 0.9 μmoles min-1mg protein-1. GFP-MtAAE3 localization suggested that this enzyme functions within the cytosol of the cell. Mtaae3 knock-down line showed a reduction in its ability to degrade oxalate into CO2. This reduction in the capacity to degrade oxalate resulted in the accumulation of druse crystals of calcium oxalate in the Mtaae3 knock-down line and an increased susceptibility to oxalate-secreting phytopathogens such as Sclerotinia sclerotiorum. Taken together, these results suggest that AAE3 dependent turnover of oxalate is important to different plants and functions in the regulation of tissue calcium oxalate crystal accumulation and in defense against oxalate-secreting phytopathogens. PMID:26900946

  17. Toxicogenomic Responses of the Model Legume Medicago truncatula to Aged Biosolids Containing a Mixture of Nanomaterials (TiO₂, Ag, and ZnO) from a Pilot Wastewater Treatment Plant.

    PubMed

    Chen, Chun; Unrine, Jason M; Judy, Jonathan D; Lewis, Ricky W; Guo, Jing; McNear, David H; Tsyusko, Olga V

    2015-07-21

    Toxicogenomic responses in Medicago truncatula A17 were monitored following exposure to biosolids-amended soils. Treatments included biosolids produced using a pilot wastewater treatment plant with either no metal introduced into the influent (control); bulk/ionic TiO2, ZnO, and AgNO3 added to influent (bulk/dissolved treatment); or Ag, ZnO, and TiO2 engineered nanomaterials added to influent (ENM treatment) and then added to soil, which was aged in the field for 6 months. In our companion study, we found inhibition of nodulation in the ENM but not in the bulk/dissolved treatment. Gene expression profiling revealed highly distinct profiles with more than 10-fold down-regulation in 239 genes in M. truncatula roots from the ENM treatment, while gene expression patterns were similar between bulk/dissolved and control treatments. In response to ENM exposure, many of the identified biological pathways, gene ontologies, and individual genes are associated with nitrogen metabolism, nodulation, metal homeostasis, and stress responses. Expression levels of nine genes were independently confirmed with qRT-PCR. Exposure to ENMs induced unique shifts in expression profiles and biological pathways compared with bulk/dissolved treatment, despite the lack of difference in bioavailable metal fractions, metal oxidation state, and coordination environment between ENM and bulk/dissolved biosolids. As populations of Sinorhizobium meliloti Rm2011 were similar in bulk/dissolved and ENM treatments, our results suggest that inhibition of nodulation in the ENM treatment was primarily due to phytotoxicity, likely caused by enhanced bioavailability of Zn ions.

  18. The major histocompatibility complex genes impact pain response in DA and DA.1U rats.

    PubMed

    Guo, Yuan; Yao, Fan-Rong; Cao, Dong-Yuan; Li, Li; Wang, Hui-Sheng; Xie, Wen; Zhao, Yan

    2015-08-01

    Our recent studies have shown that the difference in basal pain sensitivity to mechanical and thermal stimulation between Dark-Agouti (DA) rats and a novel congenic DA.1U rats is major histocompatibility complex (MHC) genes dependent. In the present study, we further used DA and DA.1U rats to investigate the role of MHC genes in formalin-induced pain model by behavioral, electrophysiological and immunohistochemical methods. Behavioral results showed biphasic nociceptive behaviors increased significantly following the intraplantar injection of formalin in the hindpaw of DA and DA.1U rats. The main nociceptive behaviors were lifting and licking, especially in DA rats (P<0.001 and P<0.01). The composite pain scores (CPS) in DA rats were significantly higher than those in DA.1U rats in both phases of the formalin test (P<0.01). Electrophysiological results also showed the biphasic increase in discharge rates of C and Aδ fibers of L5 dorsal root in the two strains, and the net change of the discharge rate of DA rats was significantly higher than that of DA.1U rats (P<0.05). The mechanical thresholds decreased after formalin injection in both strains (P<0.01), and the net change in the mechanical threshold in DA was greater than that in DA.1U rats (P<0.05). The expression of RT1-B, representation of MHC class II molecule, in laminae I-II of L4/5 spinal cord in DA rats was significantly higher than that in DA.1U rats in the respective experimental group (P<0.05). These results suggested that both DA and DA.1U rats exhibited nociceptive responses in formalin-induced pain model and DA rats were more sensitive to noxious chemical stimulus than DA.1U rats, indicating that MHC genes might contribute to the difference in pain sensitivity.

  19. Unraveling the Influence of Arbuscular Mycorrhizal Colonization on Arsenic Tolerance in Medicago: Glomus mosseae is More Effective than G. intraradices, Associated with Lower Expression of Root Epidermal Pi Transporter Genes

    PubMed Central

    Christophersen, Helle M.; Smith, F. Andrew; Smith, Sally E.

    2012-01-01

    We used medic (Medicago truncatula) to investigate effects of inoculation with two arbuscular mycorrhizal (AM) fungi and application of arsenate (AsV) and phosphate (Pi) on mechanisms underlying increased tolerance (in terms of growth) of AM plants to AsV. We tested the hypotheses that (1) inoculation with AM fungi results in down-regulation of MtPht1;1 and MtPht1;2 genes (encoding high-affinity Pi and AsV uptake systems in the direct root epidermal pathway) and up-regulation of the AM-induced MtPht1;4 (responsible for transfer of Pi from the arbuscular interface to cortical cells), and (2) these changes are involved in decreased As uptake relative to P uptake and hence increased As tolerance. We also measured expression of MtMT4, a Pi starvation-inducible gene, other genes encoding Pi uptake systems (MtPht 1;5 and MtPht1;6) and arsenate reductase (MtACR) and phytochelatin synthase (MtPCS), to gain insights into broader aspects of P transfers in AM plants and possible detoxification mechanisms. Medic responded slightly to AM colonization in terms of growth in the absence of As, but positively in terms of P uptake. Both growth and P responses in AM plants were positive when As was applied, indicating As tolerance relative to non-mycorrhizal (NM) plants. All AM plants showed high expression of MtPT4 and those inoculated with Glomus mosseae showed higher selectivity against As (shown by P/As molar ratios) and much lower expression of MtPht1;1 (and to some extent MtPht1;2) than Glomus intraradices-inoculated or NM plants. Results are consistent with increased P/As selectivity in AM plants (particularly those inoculated with G. mosseae) as a consequence of high P uptake but little or no As uptake via the AM pathway. However, the extent to which selectivity is dependent on down-regulation of direct Pi and AsV uptake through epidermal cells is still not clear. Marked up-regulation of a PCS gene and an ACR gene in AM plants may also be involved and requires further

  20. Virtual observatory publishing with DaCHS

    NASA Astrophysics Data System (ADS)

    Demleitner, M.; Neves, M. C.; Rothmaier, F.; Wambsganss, J.

    2014-11-01

    The Data Center Helper Suite DaCHS is an integrated publication package for building VO and Web services, supporting the entire workflow from ingestion to data mapping to service definition. It implements all major data discovery, data access, and registry protocols defined by the VO. DaCHS in this sense works as glue between data produced by the data providers and the standard protocols and formats defined by the VO. This paper discusses central elements of the design of the package and gives two case studies of how VO protocols are implemented using DaCHS' concepts.

  1. Origin of the DA and non-DA white dwarf stars

    NASA Technical Reports Server (NTRS)

    Shipman, Harry L.

    1989-01-01

    Various proposals for the bifurcation of the white dwarf cooling sequence are reviewed. 'Primordial' theories, in which the basic bifurcation of the white dwarf sequence is rooted in events predating the white dwarf stage of stellar evolution, are discussed, along with the competing 'mixing' theories in which processes occurring during the white dwarf stage are responsible for the existence of DA or non-DA stars. A new proposal is suggested, representing a two-channel scenario. In the DA channel, some process reduces the hydrogen layer mass to the value of less than 10 to the -7th. The non-DA channel is similar to that in the primordial scenario. These considerations suggest that some mechanism operates in both channels to reduce the thickness of the outermost layer of the white dwarf. It is also noted that accretion from the interstellar medium has little to do with whether a particular white dwarf becomes a DA or a non-DA star.

  2. Genetic Engineering of Alfalfa (Medicago sativa L.).

    PubMed

    Wang, Dan; Khurshid, Muhammad; Sun, Zhan Min; Tang, Yi Xiong; Zhou, Mei Liang; Wu, Yan Min

    2016-01-01

    Alfalfa is excellent perennial legume forage for its extensive ecological adaptability, high nutrition value, palatability and biological nitrogen fixation. It plays a very important role in the agriculture, animal husbandry and ecological construction. It is cultivated in all continents. With the development of modern plant breeding and genetic engineering techniques, a large amount of work has been carried out on alfalfa. Here we summarize the recent research advances in genetic engineering of alfalfa breeding, including transformation, quality improvement, stress resistance and as a bioreactor. The review article can enables us to understand the research method, direction and achievements of genetic engineering technology of Alfalfa.

  3. The Medicago HapMap Project

    SciTech Connect

    Bharti, Arvind

    2010-06-03

    Arvind Bharti from the National Center for Genome Resources discusses resequencing 400 plant and bacteria genotypes on June 3, 2010 at the "Sequencing, Finishing, Analysis in the Future" meeting in Santa Fe, NM

  4. The Supernova Impostor SN 2010da

    NASA Astrophysics Data System (ADS)

    Binder, Breanna A.; Williams, Benjamin F.; Kong, Albert K. H.; Plucinsky, Paul P.; Gaetz, Terrance J.; Skillman, Evan D.; Dolphin, Andrew E.

    2016-01-01

    Supernova impostors are optical transients that, despite being assigned a supernova designation, do not signal the death of a massive star or accreting white dwarf. Instead, many impostors are thought to be major eruptions from luminous blue variables. Although the physical cause of these eruptions is still debated, tidal interactions from a binary companion has recently gained traction as a possible explanation for observations of some supernova impostors. In this talk, I will discuss the particularly interesting impostor SN 2010da, which exhibits high-luminosity, variable X-ray emission. The X-ray emission is consistent with accretion onto a neutron star, making SN 2010da a likely high mass X-ray binary in addition to a supernova impostor. SN 2010da is a unique laboratory for understanding both binary interactions as drivers of massive star eruptions and the evolutionary processes that create high mass X-ray binaries.

  5. The local effect of octreotide on mechanical pain sensitivity is more sensitive in DA rats than DA.1U rats.

    PubMed

    Yao, Fan-Rong; Wang, Hui-Sheng; Guo, Yuan; Zhao, Yan

    2016-02-01

    A recent study by the authors indicated that major histocompatibility complex (MHC) genes are associated with the differences in basal pain sensitivity and in formalin model between Dark-Agouti (DA) and novel congenic DA.1U rats, which have the same genetic background as DA rats except for the u alleles of MHC. The objective of the present study is to investigate whether there is a difference in the pristane-induced arthritis (PIA) model and local analgesic effect of octreotide (OCT) between DA and DA.1U rats. The hindpaw mechanical withdrawal threshold (MWT) and heat withdrawal latency (HWL) were observed. The C unit firings of the tibial nerve evoked by non-noxious and noxious toe movements were recorded by electrophysiological methods in normal and PIA models in DA and DA.1U rats before and after local OCT administration. The expression of somatostatin receptor 2A (SSTR2A) was observed by immunohistochemistry. The results demonstrate that DA rats have a higher mechanical sensitivity than DA.1U rats after PIA. Local OCT administration significantly elevated MWT in DA rats under normal and PIA sate, but not in DA.1U rats. The electrophysiological experiments showed OCT significantly attenuated the firings of C units evoked by non-noxious and noxious stimulation in DA rats more than those in DA.1U rats both in normal and PIA states. In addition, the expression of SSTR2A in the dorsal horn of the spinal cord was significantly higher in DA than in DA.1U rats. All of the findings suggest a higher local analgesic effect of OCT in DA rats than DA.1U rats, which might be associated with the MHC genes.

  6. Common colds on Tristan da Cunha

    PubMed Central

    Shibli, M.; Gooch, S.; Lewis, H. E.; Tyrrell, D. A. J.

    1971-01-01

    Eight epidemics of respiratory disease have been observed among islanders of Tristan da Cunha. They seem to be initiated by the arrival of ships and transmission seemed to occur as a result of close human contact but could not always be traced. Islanders suffered from less colds than those in less isolated communities. PMID:5282927

  7. DA White Dwarfs in the Kepler Field

    NASA Astrophysics Data System (ADS)

    Doyle, T. F.; Howell, S. B.; Petit, V.; Lépine, S.

    2016-10-01

    We present 16 new, and confirm 7 previously identified, DA white dwarfs in the Kepler field through ground-based spectroscopy with the Hale 200″, Kitt Peak 4-meter, and Bok 2.3-meter telescopes. Using atmospheric models we determine their effective temperatures and surface gravities to constrain their position with respect to the ZZ Ceti (DA pulsator) instability strip, and look for the presence or absence of pulsation with Kepler's unprecedented photometry. Our results are as follows: i) From our measurements of temperature and surface gravity, 12 of the 23 DA white dwarfs from this work fall well outside of the instability strip. The Kepler photometry available for 11 of these WDs allows us to confirm that none are pulsating. One of these eleven happens to be a presumed binary, KIC 11604781, with a period of ˜5 days. ii) The remaining 11 DA white dwarfs are instability strip candidates, potentially falling within the current, empirical instability strip, after accounting for uncertainties. These WDs will help constrain the strip's location further, as eight are near the blue edge and three are near the red edge of the instability strip. Four of these WDs do not have Kepler photometry, so ground-based photometry is needed to determine the pulsation nature of these white dwarfs. The remaining seven have Kepler photometry available, but do not show any periodicity on typical WD pulsation timescales.

  8. How to Think Like Leonardo da Vinci

    ERIC Educational Resources Information Center

    Caouette, Ralph

    2008-01-01

    To be effective and relevant in twenty-first-century learning, art needs to be more inclusive. In this article, the author discusses how teachers can find a good example in Leonardo da Vinci for building an art program. His art, design, and curiosity are the perfect foundation for any art program, at any level. (Contains 3 resources and 3 online…

  9. The PAN-DA data acquisition system

    SciTech Connect

    Petravick, D.; Berg, D.; Berman, E.; Bernett, M.; Constanta-Fanourakis, P.; Dorries, T.; Haire, M.; Kaczar, K; MacKinnon, B.; Moore, C.; Nicinski, T.; Oleynik, G.; Pordes, R.; Sergey, G.; Votava, M.; White, V.

    1989-05-01

    The Online and Data Acquisition software groups at Fermi National Accelerator Laboratory have extended the VAXONLINE data acquisition package to include a VME based data path. The resulting environment, PAN-DA, provides a high throughput for logging, filtering, formatting and selecting events. 10 refs., 1 fig.

  10. Hidden sketches by Leonardo da Vinci revealed

    NASA Astrophysics Data System (ADS)

    Dumé, Belle

    2009-02-01

    Three drawings on the back of Leonardo da Vinci's The Virgin and Child with St Anne (circa 1508) have been discovered by researchers led by Michel Menu from the Centre de Recherche et de Restauration des Musées de France (C2RMF) and the Louvre Museum in Paris.

  11. Impact of whole-genome and tandem duplications in the expansion and functional diversification of the F-box family in legumes (Fabaceae).

    PubMed

    Bellieny-Rabelo, Daniel; Oliveira, Antônia Elenir Amâncio; Venancio, Thiago Motta

    2013-01-01

    F-box proteins constitute a large gene family that regulates processes from hormone signaling to stress response. F-box proteins are the substrate recognition modules of SCF E3 ubiquitin ligases. Here we report very distinct trends in family size, duplication, synteny and transcription of F-box genes in two nitrogen-fixing legumes, Glycine max (soybean) and Medicago truncatula (alfafa). While the soybean FBX genes emerged mainly through segmental duplications (including whole-genome duplications), M. truncatula genome is dominated by locally-duplicated (tandem) F-box genes. Many of these young FBX genes evolved complex transcriptional patterns, including preferential transcription in different tissues, suggesting that they have probably been recruited to important biochemical pathways (e.g. nodulation and seed development).

  12. 32 CFR 516.25 - DA Form 4.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 32 National Defense 3 2010-07-01 2010-07-01 true DA Form 4. 516.25 Section 516.25 National Defense Department of Defense (Continued) DEPARTMENT OF THE ARMY AID OF CIVIL AUTHORITIES AND PUBLIC RELATIONS LITIGATION Reporting Legal Proceedings to HQDA § 516.25 DA Form 4. (a) General. The DA Form 4 (See figure...

  13. A Day in the Life at DaVita Academy

    ERIC Educational Resources Information Center

    Weinstein, Margery

    2010-01-01

    When a company name means "giving life," the bar for learning and development programs is held high. In this article, the author describes what it takes to graduate from DaVita Academy, the soft skills training program dialysis services company DaVita offers all its employees. DaVita's chief executive officer, Kent Thiry, states that the Academy…

  14. PanDA for COMPASS at JINR

    NASA Astrophysics Data System (ADS)

    Petrosyan, A. Sh.

    2016-09-01

    PanDA (Production and Distributed Analysis System) is a workload management system, widely used for data processing at experiments on Large Hadron Collider and others. COMPASS is a high-energy physics experiment at the Super Proton Synchrotron. Data processing for COMPASS runs locally at CERN, on lxbatch, the data itself stored in CASTOR. In 2014 an idea to start running COMPASS production through PanDA arose. Such transformation in experiment's data processing will allow COMPASS community to use not only CERN resources, but also Grid resources worldwide. During the spring and summer of 2015 installation, validation and migration work is being performed at JINR. Details and results of this process are presented in this paper.

  15. The DA{phi}NE-Light Facility

    SciTech Connect

    Burattini, Emilio; Cinque, Gianfelice; Dabagov, Sultan; Grilli, Antonio; Marcelli, Augusto; Pace, Elisabetta; Piccinini, Massimo; Raco, Agostino; Monti, Francesca

    2004-05-12

    The new Synchrotron Radiation facility at Frascati exploits the intense photon emission from DA{phi}NE, the 0,51 GeV storage ring circulating over 1 A of electrons. Among the three beamlines commissioned, the Synchrotron INfrared Beamline At Da{phi}ne (SINBAD) is fully operational by a brilliant SR beam spanning the entire IR. Recently, the soft X-ray beamline has been characterized and, once implemented the double-crystal monocromator, X-ray Absorption Spectroscopy is applied on material standards in the distinguishing energy region below 4 keV. An UltraViolet line, presently dedicated to photobiology dosimetry, has also given first results on cell irradiation in the UVB band.

  16. [Leonardo da Vinci--a dyslectic genius?].

    PubMed

    Røsstad, Anna

    2002-12-10

    Leonardo da Vinci's texts consist almost exclusively of scientific notes. Working on a book on Leonardo's art, I studied all Leonardo's published texts carefully for any new information. In some prefaces I came to suspect that Leonardo might have suffered from dyslexia. This article considers the question of whether it is possible to find indications of dyslexia in Leonardo's texts and in the accounts of his life.

  17. The Real Code of Leonardo da Vinci

    PubMed Central

    Ose, Leiv

    2008-01-01

    Leonardo da Vinci was born in Italy. Among the researchers and scientists, he is favourably known for his remarkable efforts in scientific work. His investigations of atherosclerosis judiciously combine three separate fields of research. In 1506, he finished his masterpiece, painting of Mona Lisa. A careful clinical examination of the famous painting reveals a yellow irregular leather-like spot at the inner end of the left upper eyelid and a soft bumpy well-defined swelling of the dorsum of the right hand beneath the index finger about 3 cm long. This is probably the first case of familial hypercholesterolemia (FH). The FH code of Leonardo da Vinci was given immense consideration by scientists like Carl Muller, who described the xanthomas tuberosum and angina pectoris. On the contrary, Akira Endo searched for microbial metabolites that would inhibit HMG-CoA reductase, the rate-limiting enzyme in the synthesis of cholesterol and finally, Michael Brown and Joseph Goldstein published a remarkable series of elegant and insightful papers in the 70s and 80s. They established that the cellular uptake of low-density lipoprotein (LDL) essentially requires the LDL receptor. In conclusion: this was the real Code of Leonardo da Vinci. PMID:19924278

  18. Leonardo da Vinci's studies of the heart.

    PubMed

    Shoja, Mohammadali M; Agutter, Paul S; Loukas, Marios; Benninger, Brion; Shokouhi, Ghaffar; Namdar, Husain; Ghabili, Kamyar; Khalili, Majid; Tubbs, R Shane

    2013-08-20

    Leonardo da Vinci's detailed drawings are justly celebrated; however, less well known are his accounts of the structures and functions of the organs. In this paper, we focus on his illustrations of the heart, his conjectures about heart and blood vessel function, his experiments on model systems to test those conjectures, and his unprecedented conclusions about the way in which the cardiovascular system operates. In particular, da Vinci seems to have been the first to recognize that the heart is a muscle and that systole is the active phase of the pump. He also seems to have understood the functions of the auricles and pulmonary veins, identified the relationship between the cardiac cycle and the pulse, and explained the hemodynamic mechanism of valve opening and closure. He also described anatomical variations and changes in structure and function that occurred with age. We outline da Vinci's varied career and suggest ways in which his personality, experience, skills and intellectual heritage contributed to these advances in understanding. We also consider his influence on later studies in anatomy and physiology.

  19. Turbocharging the DA465 gasoline engine

    NASA Astrophysics Data System (ADS)

    Zhang, Peng-Qi; Zong, Li-Jun; Wang, Yin-Yan

    2008-06-01

    In order to improve performance of the DA465Q gasoline engine, a substantial amount of research was done to optimize its turbocharging system. The research led to the GT12 turbocharger being selected and its turbocharging parameters being settled. Based on these tests, rational matching was worked out for respective components of the turbocharging system. Results show that this turbocharger allows the engine to easily meet the proposed requirements for power and economic performance, giving insight into further performance improvements for gasoline engines.

  20. The Case: Bunche-Da Vinci Learning Partnership Academy

    ERIC Educational Resources Information Center

    Eisenberg, Nicole; Winters, Lynn; Alkin, Marvin C.

    2005-01-01

    The Bunche-Da Vinci case described in this article presents a situation at Bunche Elementary School that four theorists were asked to address in their evaluation designs (see EJ791771, EJ719772, EJ791773, and EJ792694). The Bunche-Da Vinci Learning Partnership Academy, an elementary school located between an urban port city and a historically…

  1. 40 CFR 60.52Da - Recordkeeping requirements.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 6 2010-07-01 2010-07-01 false Recordkeeping requirements. 60.52Da Section 60.52Da Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR PROGRAMS (CONTINUED) STANDARDS OF PERFORMANCE FOR NEW STATIONARY SOURCES Standards of Performance for Electric Utility Steam Generating Units for...

  2. Asteroid 2012 DA14 To Whiz Past Earth Safely

    NASA Video Gallery

    The small near-Earth asteroid 2012 DA14 will pass very close to Earth on Feb. 15, 2013. Asteroid 2012 DA14 will be closest to Earth at about 11:24 a.m. PST (2:24 p.m. EST and 1924 UTC), on Feb. 15,...

  3. DA-6034 Induces [Ca(2+)]i Increase in Epithelial Cells.

    PubMed

    Yang, Yu-Mi; Park, Soonhong; Ji, Hyewon; Kim, Tae-Im; Kim, Eung Kweon; Kang, Kyung Koo; Shin, Dong Min

    2014-04-01

    DA-6034, a eupatilin derivative of flavonoid, has shown potent effects on the protection of gastric mucosa and induced the increases in fluid and glycoprotein secretion in human and rat corneal and conjunctival cells, suggesting that it might be considered as a drug for the treatment of dry eye. However, whether DA-6034 induces Ca(2+) signaling and its underlying mechanism in epithelial cells are not known. In the present study, we investigated the mechanism for actions of DA-6034 in Ca(2+) signaling pathways of the epithelial cells (conjunctival and corneal cells) from human donor eyes and mouse salivary gland epithelial cells. DA-6034 activated Ca(2+)-activated Cl(-) channels (CaCCs) and increased intracellular calcium concentrations ([Ca(2+)]i) in primary cultured human conjunctival cells. DA-6034 also increased [Ca(2+)]i in mouse salivary gland cells and human corneal epithelial cells. [Ca(2+)]i increase of DA-6034 was dependent on the Ca(2+) entry from extracellular and Ca(2+) release from internal Ca(2+) stores. Interestingly, these effects of DA-6034 were related to ryanodine receptors (RyRs) but not phospholipase C/inositol 1,4,5-triphosphate (IP3) pathway and lysosomal Ca(2+) stores. These results suggest that DA-6034 induces Ca(2+) signaling via extracellular Ca(2+) entry and RyRs-sensitive Ca(2+) release from internal Ca(2+) stores in epithelial cells.

  4. Leonardo da Vinci's contributions to neuroscience.

    PubMed

    Pevsner, Jonathan

    2002-04-01

    Leonardo da Vinci (1452-1519) made far-reaching contributions to many areas of science, technology and art. Leonardo's pioneering research into the brain led him to discoveries in neuroanatomy (such as those of the frontal sinus and meningeal vessels) and neurophysiology (he was the first to pith a frog). His injection of hot wax into the brain of an ox provided a cast of the ventricles, and represents the first known use of a solidifying medium to define the shape and size of an internal body structure. Leonardo developed an original, mechanistic model of sensory physiology. He undertook his research with the broad goal of providing physical explanations of how the brain processes visual and other sensory input, and integrates that information via the soul.

  5. Da Costa's syndrome or neurocirculatory asthenia.

    PubMed Central

    Paul, O

    1987-01-01

    The syndrome variously called Da Costa's syndrome, effort syndrome, neurocirculatory asthenia, etc has been studied for more than 100 years by many distinguished physicians. Originally identified in men in wartime, it has been widely recognised as a common chronic condition in both sexes in civilian life. Although the symptoms may seem to appear after infections and various physical and psychological stresses, neurocirculatory asthenia is most often encountered as a familial disorder that is unrelated to these factors, although they may aggravate an existing tendency. Respiratory complaints (including breathlessness, with and without effort, and smothering sensations) are almost universal, and palpitation, chest discomfort, dizziness and faintness, and fatigue are common. The physical examination is normal. The aetiology is obscure but patients usually have a normal life span. Reassurance and measures to improve physical fitness are helpful. PMID:3314950

  6. 75 FR 52292 - Airworthiness Directives; Diamond Aircraft Industries GmbH Models DA 40 and DA 40F Airplanes

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-08-25

    ... ``significant rule'' under the DOT Regulatory Policies and Procedures (44 FR 11034, February 26, 1979); and 3... Federal Aviation Administration 14 CFR Part 39 RIN 2120-AA64 Airworthiness Directives; Diamond Aircraft... new airworthiness directive (AD) for all Diamond Aircraft Industries GmbH Models DA 40 and DA...

  7. 75 FR 75868 - Airworthiness Directives; Diamond Aircraft Industries GmbH Models DA 40 and DA 40F Airplanes

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-12-07

    ... Register on August 25, 2010 (75 FR 52292). That NPRM proposed to require a retrofit of the rear passenger... 12866, (2) Is not a ``significant rule'' under DOT Regulatory Policies and Procedures (44 FR 11034... Aircraft Industries GmbH Models DA 40 and DA 40F Airplanes AGENCY: Federal Aviation Administration,...

  8. Metal Lines in DA White Dwarfs

    NASA Astrophysics Data System (ADS)

    Zuckerman, B.; Koester, D.; Reid, I. N.; Hünsch, M.

    2003-10-01

    We report Keck telescope HIRES echelle observations of DA white dwarfs in a continuation of an extensive search for metals. These spectra are supplemented with new JHK magnitudes that are used to determine improved atmospheric parameters. Of the DA white dwarfs not in binary or common proper motion systems, about 25% show Ca II lines. For these, Ca abundances are determined from comparison with theoretical equivalent widths from model atmosphere calculations; in a few cases we also obtain Mg, Fe, Si, and Al abundances. If Ca is not observed, we generally determine very stringent upper limits. We compare the data to predictions of previously published models involving the accretion/diffusion of interstellar matter and of comets. The derived abundances are not obviously compatible with the predictions of either model, which up to now could only be tested with traces of metals in helium-rich white dwarfs. By modifying certain assumptions in the published interstellar accretion model we are able to match the distribution of the elements in the white dwarf atmospheres, but, even so, tests of other expectations from this scenario are less successful. Because comet accretion appears unlikely to be the primary cause of the DAZ phenomenon, the data suggest that no more than about 20% of F-type main-sequence stars are accompanied by Oort-like comet clouds. This represents the first observational estimate of this fraction. A plausible alternative to the accretion of cometary or interstellar matter is disruption and accretion of asteroidal material, a model first suggested in 1990 to explain excess near-infrared emission from the DAZ G29-38. An asteroidal debris model to account for the general DAZ phenomenon does not presently disagree with the HIRES data, but neither is there any compelling evidence in support of such a model. The HIRES data indicate that in close red dwarf/white dwarf binaries not known to be cataclysmic variables there is, nonetheless, significant mass

  9. Characterization of the isolated 20 kDa and 50 kDa fragments of the myosin head.

    PubMed

    Muhlrad, A; Kasprzak, A A; Ue, K; Ajtai, K; Burghardt, T P

    1986-01-30

    We have isolated two proteolytic fragments of subfragment 1 (S-1) of myosin from rabbit skeletal muscle. These fragments, identified by their molecular weights of 20 and 50 kDa, may be functional domains that, when isolated, retain their specific function. We have studied several structural and functional features of the 20 and 50 kDa fragments. Considerable secondary structure in both fragments has been observed in CD spectrum studies. Previously CD spectra showed 64% ordered structure for the 20 kDa fragment (Muhlrad and Morales, M.F. (1984) Proc. Natl. Acad. Sci. 81, 1003) and here we show 71% ordered structures for the 50 kDa fragment. Fluorescence lifetime studies of tryptophan residues in the 50 kDa fragment and 1,5-IAEDANS-labeled SH-1 in the 20 kDa fragment are used to investigate the tertiary structure of the fragments. We find the tertiary structure relating to this measurement of both fragments to be intact; however, the reaction of 1,5-IAEDANS with SH-1 on the isolated 20 kDa fragment is less specific than with S-1. Furthermore, the fragments showed a tendency to aggregate. The domain concept of S-1 was supported by the characteristic biochemical function of the isolated fragments. Both of the fragments were effective in competing with S-1 for binding to actin in acto-S-1 ATPase measurements. From these studies and in direct binding measurement the 20 kDa fragment proved to bind with higher affinity to actin than did the 50 kDa fragment.

  10. Tree Branching: Leonardo da Vinci's Rule versus Biomechanical Models

    PubMed Central

    Minamino, Ryoko; Tateno, Masaki

    2014-01-01

    This study examined Leonardo da Vinci's rule (i.e., the sum of the cross-sectional area of all tree branches above a branching point at any height is equal to the cross-sectional area of the trunk or the branch immediately below the branching point) using simulations based on two biomechanical models: the uniform stress and elastic similarity models. Model calculations of the daughter/mother ratio (i.e., the ratio of the total cross-sectional area of the daughter branches to the cross-sectional area of the mother branch at the branching point) showed that both biomechanical models agreed with da Vinci's rule when the branching angles of daughter branches and the weights of lateral daughter branches were small; however, the models deviated from da Vinci's rule as the weights and/or the branching angles of lateral daughter branches increased. The calculated values of the two models were largely similar but differed in some ways. Field measurements of Fagus crenata and Abies homolepis also fit this trend, wherein models deviated from da Vinci's rule with increasing relative weights of lateral daughter branches. However, this deviation was small for a branching pattern in nature, where empirical measurements were taken under realistic measurement conditions; thus, da Vinci's rule did not critically contradict the biomechanical models in the case of real branching patterns, though the model calculations described the contradiction between da Vinci's rule and the biomechanical models. The field data for Fagus crenata fit the uniform stress model best, indicating that stress uniformity is the key constraint of branch morphology in Fagus crenata rather than elastic similarity or da Vinci's rule. On the other hand, mechanical constraints are not necessarily significant in the morphology of Abies homolepis branches, depending on the number of daughter branches. Rather, these branches were often in agreement with da Vinci's rule. PMID:24714065

  11. Cloning of the 52-kDa chitinase gene from Serratia marcescens KCTC2172 and its proteolytic cleavage into an active 35-kDa enzyme.

    PubMed

    Gal, S W; Choi, J Y; Kim, C Y; Cheong, Y H; Choi, Y J; Lee, S Y; Bahk, J D; Cho, M J

    1998-03-01

    A chitinase gene (pCHI52) encoding the 52-kDa chitinase was isolated from a Serratia marcescens KCTC2172 cosmid library. This chitinase gene consists of 2526 bp with an open reading frame that encodes 485 amino acids. Escherichia coli harboring the pCHI52 gene secreted not only a 52-kDa but also a 35-kDa chitinase into the culture supernatant. We purified both 52-kDa and 35-kDa chitinases using a chitin affinity column and Sephacryl-S-300 gel filtration chromatography. We determined that the 17 N-terminal amino acid sequences of the 52-kDa and the 35-kDa chitinase are identical. Furthermore, a protease obtained from S. marcescens KCTC2172 cleaved the 52-kDa chitinase into the 35-kDa protein with chitinase activity. These results suggest that the 35-kDa chitinase derives from the 52-kDa chitinase by post-translational proteolytic modification. The optimal reaction temperature of 45 degrees C and the optimal pH of 5.5 were identical for both enzymes. The specific activities of the 52-kDa and 35-kDa chitinases on natural swollen chitin were 67 mumol min-1 mg-1 and 60 mumol min-1 mg-1, respectively.

  12. PKC phosphorylates residues in the N-terminal of the DA transporter to regulate amphetamine-induced DA efflux.

    PubMed

    Wang, Qiang; Bubula, Nancy; Brown, Jason; Wang, Yunliang; Kondev, Veronika; Vezina, Paul

    2016-05-27

    The DA transporter (DAT), a phosphoprotein, controls extracellular dopamine (DA) levels in the central nervous system through transport or reverse transport (efflux). Multiple lines of evidence support the claim that PKC significantly contributes to amphetamine-induced DA efflux. Other signaling pathways, involving CaMKII and ERK, have also been shown to regulate DAT mediated efflux. Here we assessed the contribution of putative PKC residues (S4, S7, S13) in the N-terminal of the DAT to amphetamine-induced DA efflux by transfecting DATs containing different serine to alanine (S-A) point mutations into DA pre-loaded HEK-293 cells and incubating these cells in amphetamine (2μM). The effects of a S-A mutation at the non-PKC residue S12 and a threonine to alanine (T-A) mutation at the ERK T53 residue were also assessed for comparison. WT-DATs were used as controls. In an initial experiment, we confirmed that inhibiting PKC with Go6976 (130nM) significantly reduced amphetamine-induced DA efflux. In subsequent experiments, cells transfected with the S4A, S12A, S13A, T53A and S4,7,13A mutants showed a reduction in amphetamine-induced DA efflux similar to that observed with Go6976. Interestingly, cells transfected with the S7A mutant, identified by some as a PKC-PKA residue, showed unperturbed WT-DAT levels of amphetamine-induced DA efflux. These results indicate that phosphorylation by PKC of select residues in the DAT N-terminal can regulate amphetamine-induced efflux. PKC can act either independently or in concert with other kinases such as ERK to produce this effect.

  13. A cosmologia no ensino da geografia

    NASA Astrophysics Data System (ADS)

    Santos, S. C.; Chiaradia, A. P. M.

    2003-08-01

    O principal objetivo deste trabalho é auxiliar o professor de Geografia em sala de aula no ensino de tópicos relacionados com a Cosmologia. A idéia deste trabalho surgiu quando foi constatado que o professor de Geografia tem dificuldades de ensinar este tópico. Esta constatação foi feita por uma das autoras ao lecionar este tópico no ensino fundamental e em discussões com outros professores de Geografia. Da mesma maneira que ocorria desde os tempos mais antigos, os alunos têm muito interesse em conhecer os fenômenos que ocorrem no Cosmo, porém os livros didáticos de Geografia utilizados em sala de aula não são ricos em informações sobre este assunto. Assim, o professor de Geografia tem poucas informações para discutir este assunto em sala de aula e não dá a devida importância para este tópico. Então, foi desenvolvido um material de apoio para professores de Geografia sobre a origem do Universo, sua evolução e seu possível futuro evolutivo segundo as mais recentes teorias, com base em perguntas feitas pelos alunos de ensino fundamental e as informações trazidas nos livros didáticos Não cabe a este material inovar e tão pouco trazer uma metodologia de ensino de Cosmologia. Neste material o professor de Geografia pode encontrará um banco de informações, que constitui no estabelecimento de conceitos, teorias e hipóteses, sobre a Cosmologia, em linguagem simples e de fácil entendimento. Para desenvolvê-lo, foram feitas pesquisas não exaustivas em livros e revistas científicas, compilação e discussão em forma cronológica das teorias aceitas sobre modelos cosmológicos. Portanto, este material será apresentado neste trabalho.

  14. Geomagnetic observations on tristan da cunha, south atlantic ocean

    USGS Publications Warehouse

    Matzka, J.; Olsen, N.; Maule, C.F.; Pedersen, L.W.; Berarducci, A.M.; Macmillan, S.

    2009-01-01

    Few geomagnetic ground observations exist of the Earth's strongest core field anomaly, the South Atlantic Anomaly (SAA). The geomagnetic repeat station on the island Tristan da Cunha, located half-way between South Africa and South America at 37?? 05' S, 12?? 18' W, is therefore of crucial importance. We have conducted several sets of repeat station measurements during magnetically quiet conditions (Kp 2o or less) in 2004. The procedures are described and the results are compared to those from earlier campaigns and to the predictions of various global field models. Features of the local crustal bias field and the solar quiet daily variation are discussed. We also evaluate the benefit of continuous magnetic field recordings from Tristan da Cunha, and argue that such a data set is a very valuable addition to geomagnetic satellite data. Recently, funds were set up to establish and operate a magnetometer station on Tristan da Cunha during the Swarm magnetic satellite mission (2011-2014).

  15. 1991 DA: An asteroid in a bizarre orbit

    NASA Technical Reports Server (NTRS)

    Steel, Duncan; Mcnaught, Robert H.; Asher, David

    1992-01-01

    Asteroidal object 1991 DA has an orbit of high inclination, crossing the planets from Mars to Uranus. This is unique for an asteroid, but not unusual for a comet of the Halley-type: it therefore seems likely that 1991 DA is an extinct or dormant comet. Previous CCD imaging has shown no indication of a coma; spectroscopic observations of 1991 DA which lack any evidence of strong comet-like emissions are reported. Numerical integrations of the orbit of this object were performed which show that is has been remarkably stable for the past approximately 20,000 yr, but chaotic before that. This may allow a new estimate to be made of the physical lifetimes of comets.

  16. The future of PanDA in ATLAS distributed computing

    NASA Astrophysics Data System (ADS)

    De, K.; Klimentov, A.; Maeno, T.; Nilsson, P.; Oleynik, D.; Panitkin, S.; Petrosyan, A.; Schovancova, J.; Vaniachine, A.; Wenaus, T.

    2015-12-01

    Experiments at the Large Hadron Collider (LHC) face unprecedented computing challenges. Heterogeneous resources are distributed worldwide at hundreds of sites, thousands of physicists analyse the data remotely, the volume of processed data is beyond the exabyte scale, while data processing requires more than a few billion hours of computing usage per year. The PanDA (Production and Distributed Analysis) system was developed to meet the scale and complexity of LHC distributed computing for the ATLAS experiment. In the process, the old batch job paradigm of locally managed computing in HEP was discarded in favour of a far more automated, flexible and scalable model. The success of PanDA in ATLAS is leading to widespread adoption and testing by other experiments. PanDA is the first exascale workload management system in HEP, already operating at more than a million computing jobs per day, and processing over an exabyte of data in 2013. There are many new challenges that PanDA will face in the near future, in addition to new challenges of scale, heterogeneity and increasing user base. PanDA will need to handle rapidly changing computing infrastructure, will require factorization of code for easier deployment, will need to incorporate additional information sources including network metrics in decision making, be able to control network circuits, handle dynamically sized workload processing, provide improved visualization, and face many other challenges. In this talk we will focus on the new features, planned or recently implemented, that are relevant to the next decade of distributed computing workload management using PanDA.

  17. Situação da Mulher na Astronomia Brasileira

    NASA Astrophysics Data System (ADS)

    Silva, Adriana V. R.

    2007-07-01

    O conteúdo desse texto surgiu de uma apresentação de mesmo título que fiz na XXXI Reunião Anual da Sociedade Astronômica Brasileira (SAB) em 2005. Esse tema foi inspirado originalmente pela minha participação no "2nd UIPAP International Conference on Women in Physics" realizado entre 23 e 25 de maio de 2005 no Rio de Janeiro. Essa é uma conferência internacional que acontece de três em três anos, sendo que a primeira ocorreu em 2002 na cidade de Paris, França. Participei dessa conferência como membro da delegação da Sociedade Brasileira de Física e um dos trabalhos que apresentei versava sobre a situação das mulheres na Astronomia brasileira, cujos resultados principais discorro a seguir. A situação das astrônomas, baseada nos dados dos sócios da SAB coletados no final de 2004, é comparada com a das físicas brasileiras e também com as nossas colegas americanas. Os dados identificam ainda uma maior evasão da carreira por parte das mulheres do que os homens. Alguns dos possíveis motivos da evasão são discutidos, como o desejo de constituir família e/ou isolamento. Resultados um tanto preocupantes com relação à distribuição de bolsas de produtividade do CNPq também são apresentados. As principais discussões e estratégias recomendadas nesse congresso são mencionadas de forma resumida ao final.

  18. Publishing Planetary Data to the VO using DaCHS

    NASA Astrophysics Data System (ADS)

    Demleitner, M.

    2014-04-01

    Publishing to the Virtual Observatory has many advantages, but between standards compliance, registration, data organisation, and server operation it is also requires a nontrivial effort if starting from scratch. The DaCHS software from the German Astrophysical Virtual Observatory offers a packaged solution letting publishers concentrate on working with their own data and metadata as much as possible. For the planetary community, publishing is further facilitated by built-in support for the EPN-TAP data model. This contribution will show a typical publishing workflow together with a brief overview of how DaCHS meshes in with the VO and Web-based data usage.

  19. The DaCHS Multi-protocol VO Server

    NASA Astrophysics Data System (ADS)

    Demleitner, M.

    2014-05-01

    GAVO's Data Center Helper Suite (DaCHS) is a suite of tools for publishing data to the Virtual Observatory. It implements all major VO protocols (SCS, SIAP, SSAP, TAP, OAI-PMH). The integrated management and ingestion component allow defining metadata, structure, and services once and re-use the definition throughout the publication cycle from initial metadata aquisition to registry record generation. It has been driving GAVO's data center since 2008 and is now deployed in multiple locations around the globe. This poster briefly describes the design of the system as well as a bird's-eye view of data publishing with DaCHS.

  20. Potential effects of Thiram on Medicago - R. meliloti symbiotic association.

    PubMed

    Sirois, J C; Peterson, E A; Miller, R W

    1981-01-01

    The effects of Thiram and 2 commercial Thiram formulations on the growth and respiration of rhizobia were tested to compare the extent of bacteriostasis under controlled conditions. Although bacteriostasis was measurable at all concentrations tested, liquid cultures grew to maximum optical density in Thiram suspensions containing less than 10 micrograms/ml. Percentage germination, root elongation, and subsequent nodulation by R. meliloti of 2 cultivars of alfalfa, were determined in thiram suspensions to determine potential physiological effects of the fungicide on the host plant. Conditions were identified which produced enhancement or inhibition of germination, root elongation and development of nodular nitrogenase activity. At concentrations of the fungicide recommended for seed application, only minor, temporary bacteriostasis was observed as a possible negative effect while germination rates of fungi-contaminated seed were markedly increased.

  1. Characterization of Medicago truncatula reduced calcium oxalate crystal mutant alleles

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Calcium oxalate crystal formation is common in plants. Formation of these crystals has been shown to function in plant defense, calcium regulation, and aluminum tolerance. Although calcium oxalate is common and plays important roles in plant development, our understanding of how these crystals form ...

  2. 40 CFR 60.48Da - Compliance provisions.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ...)), by the hourly heat input rate (measured according to the provisions of § 60.49Da(n)), and dividing... shall conduct the performance test required under § 60.8 using the appropriate methods in appendix A of... second sampling site shall be located at the outlet from the heat recovery steam generating...

  3. 40 CFR 60.48Da - Compliance provisions.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... emission rate in lb/MMBtu (measured by the CEMS required under § 60.49Da(c) and (d)), by the hourly heat... section may be used: (1) The owner or operator of an affected duct burner shall conduct the performance... located at the outlet from the heat recovery steam generating unit. Measurements shall be taken at...

  4. 40 CFR 60.48Da - Compliance provisions.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ...)), by the hourly heat input rate (measured according to the provisions of § 60.49Da(n)), and dividing... shall conduct the performance test required under § 60.8 using the appropriate methods in appendix A of... second sampling site shall be located at the outlet from the heat recovery steam generating...

  5. Leonardo da Vinci and the sinuses of Valsalva.

    PubMed

    Robicsek, F

    1991-08-01

    Recent studies indicate that eddy currents generated by the sinuses of Valsalva play an important role in the physiologic closure of the aortic valve. This process is briefly discussed and evidence is presented that this fact was well known and elaborated upon by the renaissance artist Leonardo da Vinci. This fact is illustrated with his words and drawings.

  6. Studying and Working Abroad. Leonardo da Vinci Series: Good Practices.

    ERIC Educational Resources Information Center

    Commission of the European Communities, Brussels (Belgium). Directorate-General for Education and Culture.

    This document profiles recent successful examples of students studying and working abroad as part of the European Commission's Leonardo da Vinci program, which is designed to give students across the European Union the opportunity to experience vocational training in a foreign country. The following examples are presented: (1) 3 Finnish students…

  7. 40 CFR 60.47Da - Commercial demonstration permit.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... Steam Generating Units for Which Construction is Commenced After September 18, 1978 § 60.47Da Commercial... potential combustion concentration (80 percent reduction) for each 24-hour period of steam generator... owner or operator of a fluidized bed combustion electric utility steam generator (atmospheric...

  8. Infrared applications for portable devices using IrDA transceivers

    NASA Astrophysics Data System (ADS)

    Krishnan, Sanjay M.; Pamidighantam, Ramana V.; Tan, Wee S.

    2001-11-01

    The internet has been the mantra in last decade and digital convergence is the latest talk on the tech streets. Features that once clearly discerned a mobile phone from a PDA or a PDA from a laptop are being replaced with multi-functional overlapping features and there is a growing need for wireless access and interoperability. Infrared communications has been addressing these needs for quite some time. The technology has matured through standardization efforts of IrDA (Infrared Data Association) and is widely used in PDAs, mobile phones, laptops, printers and digital cameras to name a few. This paper explores some new application scenarios for an IrDA transceiver. The first part of the paper describes a reference architecture for using infrared wireless communication for MP3 file exchange and the various possible user models. The second part of the paper discusses various configurations for implementing an ambient light detection scheme using IrDA transceivers, to turn off the backlighting of a LCD panel of a PDA or mobile phone in the presence of ambient light. The third part of the paper discusses various techniques of enabling extended link distance operation of IrDA transceivers for toy and remote control applications.

  9. Training and Health. Leonardo da Vinci Series: Good Practices.

    ERIC Educational Resources Information Center

    Commission of the European Communities, Brussels (Belgium). Directorate-General for Education and Culture.

    This document profiles programs in the fields of health and medicine that are offered through the European Commission's Leonardo da Vinci program. The following programs are profiled: (1) CYTOTRAIN (a transnational vocational training program in cervical cancer screening); (2) Apollo (a program of open and distance learning for paramedical…

  10. Women and Technical Professions. Leonardo da Vinci Series: Good Practices.

    ERIC Educational Resources Information Center

    Commission of the European Communities, Brussels (Belgium). Directorate-General for Education and Culture.

    This document profiles programs for women in technical professions that are offered through the European Commission's Leonardo da Vinci program. The following programs are profiled: (1) Artemis and Diana (vocational guidance programs to help direct girls toward technology-related careers); (2) CEEWIT (an Internet-based information and…

  11. 32 CFR 643.121 - Private organizations on DA installations.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ....121 Section 643.121 National Defense Department of Defense (Continued) DEPARTMENT OF THE ARMY... DA installations. (a) AR 210-1 defines and classifies private organizations, such as thrift shops and child-care centers, located on Army installations and provides policy guidance for their...

  12. 32 CFR 643.121 - Private organizations on DA installations.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ....121 Section 643.121 National Defense Department of Defense (Continued) DEPARTMENT OF THE ARMY... DA installations. (a) AR 210-1 defines and classifies private organizations, such as thrift shops and child-care centers, located on Army installations and provides policy guidance for their...

  13. 32 CFR 643.121 - Private organizations on DA installations.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ....121 Section 643.121 National Defense Department of Defense (Continued) DEPARTMENT OF THE ARMY... DA installations. (a) AR 210-1 defines and classifies private organizations, such as thrift shops and child-care centers, located on Army installations and provides policy guidance for their...

  14. 32 CFR 643.121 - Private organizations on DA installations.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ....121 Section 643.121 National Defense Department of Defense (Continued) DEPARTMENT OF THE ARMY... DA installations. (a) AR 210-1 defines and classifies private organizations, such as thrift shops and child-care centers, located on Army installations and provides policy guidance for their...

  15. The Potential da Vinci in All of Us

    ERIC Educational Resources Information Center

    Petto, Sarah; Petto, Andrew

    2009-01-01

    The study of the human form is fundamental to both science and art curricula. For vertebrates, perhaps no feature is more important than the skeleton to determine observable form and function. As Leonard da Vinci's famous Proportions of the Human Figure (Virtruvian Man) illustrates, the size, shape, and proportions of the human body are defined by…

  16. 40 CFR 60.48Da - Compliance provisions.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... combined cycle systems, either of the procedures described in paragraph (j)(1) or (2) of this section may... carbon dioxide (CO2)) and meet the requirements of § 60.49Da. Alternatively, data from a NOX emission... shall constitute the NOX emission rate from the duct burner of the combined cycle system. (k)...

  17. Da Que Hablar (Something To Talk About), 1991-1993.

    ERIC Educational Resources Information Center

    Da Que Hablar, 1993

    1993-01-01

    This document consists of all 14 issues of a bimonthly serial, from its inception in May 1991 through November 1993. "Da Que Hablar" provides numerous authentic materials from magazines and newspapers to stimulate discussion in Spanish in the foreign language classroom. The articles cover topics such as current events, cultural issues, sports,…

  18. 40 CFR 60.45Da - Standard for mercury (Hg).

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 6 2010-07-01 2010-07-01 false Standard for mercury (Hg). 60.45Da... for mercury (Hg). (a) For each coal-fired electric utility steam generating unit other than an IGCC... gases that contain mercury (Hg) emissions in excess of each Hg emissions limit in paragraphs...

  19. Modelagem do vento e da fotosfera de AG Carinae

    NASA Astrophysics Data System (ADS)

    Groh, J. H.; Damineli, A.

    2003-08-01

    A trajetória evolutiva das estrelas de alta massa depende fortemente de suas taxas de perda de massa. Apesar do rápido progresso no estudo destas estrelas, a taxa de perda de massa e outros parâmetros físicos básicos, como a temperatura superficial e a velocidade terminal do vento ainda não estão bem determinados. Isto ocorre devido à presença de ventos irregulares, rápidos e fortes ao redor destas estrelas, tornando a interpretação dos seus espectros uma tarefa difícil. Assim, a modelagem do vento e da fotosfera dessas estrelas está sendo cada vez mais usada para obter tais parâmetros a partir dos espectros. O aumento da taxa de perda de massa durante a fase LBV (Variáveis Luminosas Azuis), comparado com outros tipos de estrelas, tem sido atribuído a instabilidades do tipo S Doradus. Dispomos de uma base de dados espectroscópicos cobrindo 22 anos de observações de AG Carinae, incluindo um ciclo S Doradus completo, com espectros CCD em alta resolução na faixa óptica e infravermelha. Utilizamos o programa desenvolvido por Schmutz (1997) para uma análise preliminar desse ciclo, obtendo a taxa de perda de massa a partir da linha do Ha. Não existe uma correlação clara da taxa de perda de massa com mudanças da temperatura efetiva, do raio da estrela e do fluxo na banda V. A estrela atingiu seu mínimo fotométrico (raio mínimo) em 1990 e o máximo fotométrico (raio máximo) em 1995, enquanto que o fluxo máximo da linha do Ha ocorreu em 1996. Além disso a taxa de perda de massa não segue esse ciclo, contrariamente às idéias correntes. Para fazer um modelo mais realista estamos usando o programa CMFGEN (Hillier & Miller), que trata a fotosfera e o vento estelar de forma consistente, considerando a radiação fora do equilíbrio termodinâmico (NLTE) e com blanketting total de linhas. Simulamos o espectro de AG Carinae em duas épocas extremas do ciclo S Dor para testar os resultados obtidos com o modelo mais simplificado.

  20. Roles and Delegation of Authority (R/DA) System

    SciTech Connect

    ABBOTT,JOHN P.; HUTCHINS,JAMES C.; SCHOCH,DAVID G.

    1999-11-01

    The processes of defining managerial roles and providing for delegation of authority are essential to any enterprise. At most large organizations, these processes are defined in policy manuals and through sets of standard operating procedures for many, if not all, business and administrative functions. Many of these staff-initiated, administrative functions require the routing of documents for approval to one or more levels of management. These employee-oriented, back office types of workflows tend to require more flexibility in determining to whom these documents should go to, while, at the same time, providing the responsible parties with the flexibility to delegate their approval authority or allow others to review their work. Although this practice is commonplace in manual, paper-based processes that exist in many organizations, it is difficult to provide the same flexibility in the more structured, electronic-based, workflow systems. The purpose of this report is to present a framework or architecture for creating a R/DA system and provide some insights associated with its design and utilization. To improve understanding and clarify subsequent discussion, the goals and requirements for the major R/DA system components, namely, the database and interface modules, are initially presented along with the identification of important concepts and the definition of critical terms. Next high-level functions relating the types of inputs to the outputs of the R/DA interface module are introduced and discussed. Then the relationships between the major R/DA modules and the primary components associated with its creation and maintenance are presented and analyzed. Finally, some conclusions are drawn relative to the advantages associated with developing a R/DA system for use in implementing an enterprise-wide, work-facilitating information system.

  1. 32 CFR 635.23 - DA Form 4833 (Commander's Report of Disciplinary or Administrative Action).

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... DA Form 4833 within 45 days of receipt. (1) Appropriate blocks will be checked and blanks annotated... the disposition on DA Form 4833 as appropriate. That portion of the signature block of DA Form 4833... the format and at the frequency specified by the supported commander....

  2. In vivo breakdown products of the 32 kDa thylakoid herbicide binding protein. [Spirodela oligorrhiza

    SciTech Connect

    Greenberg, B.M.; Mattoo, A.K.; Gaba, V.; Edelman, M.

    1986-04-01

    The 32 kDa herbicide binding protein of PSII is degraded rapidly in the light. This phenomenon was investigated in Spirodela oligorrhiza. When fronds were radiolabeled in the presence of cycloheximide only the chloroplast-encoded proteins were labeled. Under these conditions, a breakdown product of 23.5 kDa was observed. This polypeptide was further degraded with kinetics similar to that of the 32 kDa protein. The 23.5 kDa polypeptide cross-reacted with an antibody specific to the 32 kDa protein. By protease digestion it was determined that the 23.5 kDa polypeptide is the intact N-terminal piece of the 32 kDa protein. Thus, this product corresponds to the membrane anchor of the 32 kDa protein. Using the same antibody, breakdown products of 16 kDa, 14 kDa and 12 kDa were observed. These products have also been observed in Zea mays and Solanum nigrum. Using DCMU during pulse-chase experiments at different light intensities, evidence was obtained that the 23.5 kDa breakdown product is generated in vivo.

  3. PanDaTox: a tool for accelerated metabolic engineering

    SciTech Connect

    Amitai, Gil; Sorek, Rotem

    2012-04-18

    Metabolic engineering is often facilitated by cloning of genes encoding enzymes from various heterologous organisms into E. coli. Such engineering efforts are frequently hampered by foreign genes that are toxic to the E. coli host. We have developed PanDaTox (www.weizmann.ac.il/pandatox), a web-based resource that provides experimental toxicity information for more than 1.5 million genes from hundreds of different microbial genomes. The toxicity predictions, which were extensively experimentally verified, are based on serial cloning of genes into E. coli as part of the Sanger whole genome shotgun sequencing process. PanDaTox can accelerate metabolic engineering projects by allowing researchers to exclude toxic genes from the engineering plan and verify the clonability of selected genes before the actual metabolic engineering experiments are conducted.

  4. [Lobectomy for lung cancer using the Da Vinci surgical system].

    PubMed

    Nakamura, Hiroshige

    2014-05-01

    Robot-assisted surgery using the da Vinci surgical system has attracted attention because of excellent operability without shaking by joint forceps under the clear vision of a three-dimensional high-definition camera in lung cancer surgery. Although this form of advanced medical care is not yet approved for insurance coverage, it is at the stage of clinical research and expected to be useful in hilar exposure, lymph node dissection, and suturing of the lung parenchyma or bronchus. Lung cancer surgery with the da Vinci system has the advantage of combining thoracotomy and minimally invasive surgery in video-assisted thoracic surgery. However, safety management, education, and significant cost are problems to be resolved. Several important issues such as sharing knowledge and technology of robotic surgery, education, training, development of new instruments, and acquisition of advanced medical insurance are discussed for the future development of robotic surgical systems. PMID:24946522

  5. A novel 29-kDa chicken heat shock protein.

    PubMed

    Einat, M F; Haberfeld, A; Shamay, A; Horev, G; Hurwitz, S; Yahav, S

    1996-12-01

    The family of small heat shock proteins is the more variable among the highly conserved superfamily of heat shock proteins (HSP). Using a metabolic labeling procedure with tissue explants, we have detected in chickens a new member of the small HSP family with an apparent molecular weight of 29-kDa. This protein was induced in broiler chickens' heart muscle and lungs following an in vivo heat stress. The 29-kDa band appears after 3 h of heat stress, much later than the induction of HSP 90, HSP 70, and HSP 27. The late onset of induction suggests that HSP 29 plays a more specific role of a "second stage defense protein". PMID:9000279

  6. The Da Vinci code dynamically de-coded.

    PubMed

    Cohen, Mariam

    2005-01-01

    The novel The Da Vinci Code, by Dan Brown has been on best-seller lists for over two years. An examination of Brown's previous novels reveals a well-designed plot line shared by all four novels that not only makes them good "thrillers" but also creates a mythological structure to the novels that draws on common unconscious fantasies in the same way that fairy tales do. One aspect of this mythological structure is the use of evil conspiracies (and benign ones as well) for the protagonist to overcome. In addition, The Da Vinci Code presents a religious theme involving legends about Mary Magdalene. This theme touches on the role of a feminine aspect to divinity in allowing for an erotic connection with the divine.

  7. TA-DA: A Tool for Astrophysical Data Analysis

    NASA Astrophysics Data System (ADS)

    Da Rio, Nicola; Robberto, Massimo

    2012-12-01

    We present the Tool for Astrophysical Data Analysis (TA-DA), a new software aimed to greatly simplify and improve the analysis of stellar photometric data in comparison with theoretical models, and allow the derivation of stellar parameters from multi-band photometry. Its flexibility allows one to address a number of such problems: from the interpolation of stellar models, or sets of stellar physical parameters in general, to the computation of synthetic photometry in arbitrary filters or units; from the analysis of observed color-magnitude diagrams to a Bayesian derivation of stellar parameters (and extinction) based on multi-band data. TA-DA is available as a pre-compiled Interactive Data Language widget-based application; its graphical user interface makes it considerably user-friendly. In this paper, we describe the software and its functionalities.

  8. TA-DA: A TOOL FOR ASTROPHYSICAL DATA ANALYSIS

    SciTech Connect

    Da Rio, Nicola; Robberto, Massimo

    2012-12-01

    We present the Tool for Astrophysical Data Analysis (TA-DA), a new software aimed to greatly simplify and improve the analysis of stellar photometric data in comparison with theoretical models, and allow the derivation of stellar parameters from multi-band photometry. Its flexibility allows one to address a number of such problems: from the interpolation of stellar models, or sets of stellar physical parameters in general, to the computation of synthetic photometry in arbitrary filters or units; from the analysis of observed color-magnitude diagrams to a Bayesian derivation of stellar parameters (and extinction) based on multi-band data. TA-DA is available as a pre-compiled Interactive Data Language widget-based application; its graphical user interface makes it considerably user-friendly. In this paper, we describe the software and its functionalities.

  9. [Lobectomy for lung cancer using the Da Vinci surgical system].

    PubMed

    Nakamura, Hiroshige

    2014-05-01

    Robot-assisted surgery using the da Vinci surgical system has attracted attention because of excellent operability without shaking by joint forceps under the clear vision of a three-dimensional high-definition camera in lung cancer surgery. Although this form of advanced medical care is not yet approved for insurance coverage, it is at the stage of clinical research and expected to be useful in hilar exposure, lymph node dissection, and suturing of the lung parenchyma or bronchus. Lung cancer surgery with the da Vinci system has the advantage of combining thoracotomy and minimally invasive surgery in video-assisted thoracic surgery. However, safety management, education, and significant cost are problems to be resolved. Several important issues such as sharing knowledge and technology of robotic surgery, education, training, development of new instruments, and acquisition of advanced medical insurance are discussed for the future development of robotic surgical systems.

  10. 16-kDa prolactin and bromocriptine in postpartum cardiomyopathy.

    PubMed

    Hi