scholarly journals Spatiotemporal cytokinin signaling imaging reveals IPT3 function in nodule development in Medicago truncatula

2021 ◽  
Author(s):  
Paolo M. Triozzi ◽  
Thomas B. Irving ◽  
Henry W. Schmidt ◽  
Zachary P. Keyser ◽  
Sanhita Chakraborty ◽  
...  
Keyword(s):  
High Resolution ◽  
Medicago Truncatula ◽  
Sinorhizobium Meliloti ◽  
Root Nodules ◽  
Nodule Development ◽  
Nodule Formation ◽  
Symbiotic Association ◽  
Loss Of Function ◽  
Cytokinin Signaling ◽  
Tissue Specific

ABSTRACTMost legumes can establish a symbiotic association with soil rhizobia that triggers the development of root nodules. These nodules host the rhizobia and allow them to fix nitrogen efficiently. The perception of bacterial lipo-chitooligosaccharide (LCO) signal in the epidermis initiates a signaling cascade that allows rhizobial intracellular infection in the root and de-differentiation and activation of cell division that gives rise to the nodule. Nodule organogenesis and rhizobial infection need to be coupled in space and time for successful nodulation. The plant hormone cytokinin (CK) acts as an essential positive regulator of nodule organogenesis, and specific CK receptors are required for nodule formation. Temporal regulation of tissue-specific CK signaling and biosynthesis in response to LCOs or Sinorhizobium meliloti inoculation in Medicago truncatula remains poorly understood. In the present study, using a fluorescence-based CK sensor (TCSn::nls:tGFP), we performed a high-resolution tissue-specific temporal characterization of the CK response’s sequential activation during root infection and nodule development in M. truncatula after inoculation with S. meliloti. Loss-of-function mutants of the CK-biosynthetic gene ISOPENTENYL TRANSFERASE 3 (IPT3) showed impairment of nodulation, suggesting that IPT3 is required for nodule development in M. truncatula. Simultaneous live imaging of pIPT3::tdTOMATO and the CK sensor showed that IPT3 induction in the root stele at the base of nodule primordium contributes to CK biosynthesis, which in turn promotes expression of positive regulators of nodule organogenesis in M. truncatula.One-sentence summaryHigh-resolution spatiotemporal imaging of cytokinin signaling reveals IPT3 function during indeterminate nodule development in Medicago truncatula

scholarly journals Spatiotemporal cytokinin response imaging and ISOPENTENYLTRANSFERASE 3 function in Medicago nodule development

2021 ◽  
Author(s):  
Paolo M Triozzi ◽  
Thomas B Irving ◽  
Henry W Schmidt ◽  
Zachary P Keyser ◽  
Sanhita Chakraborty ◽  
...  
Keyword(s):  
Sinorhizobium Meliloti ◽  
Root Nodules ◽  
Nodule Development ◽  
Symbiotic Association ◽  
Loss Of Function ◽  
Tissue Specific ◽  
Temporal Regulation ◽  
Positive Regulators ◽  
Sequential Activation

Abstract Most legumes can establish a symbiotic association with soil rhizobia that trigger the development of root nodules. These nodules host the rhizobia and allow them to fix nitrogen efficiently. The perception of bacterial lipo-chitooligosaccharides (LCOs) in the epidermis initiates a signaling cascade that allows rhizobial intracellular infection in the root and de-differentiation and activation of cell division that gives rise to the nodule. Thus, nodule organogenesis and rhizobial infection need to be coupled in space and time for successful nodulation. The plant hormone cytokinin (CK) contributes to the coordination of this process, acting as an essential positive regulator of nodule organogenesis. However, the temporal regulation of tissue-specific CK signaling and biosynthesis in response to LCOs or Sinorhizobium meliloti inoculation in Medicago truncatula remains poorly understood. In this study, using a fluorescence-based CK sensor (pTCSn::nls:tGFP), we performed a high-resolution tissue-specific temporal characterization of the sequential activation of CK response during root infection and nodule development in M. truncatula after inoculation with S. meliloti. Loss-of-function mutants of the CK-biosynthetic gene ISOPENTENYLTRANSFERASE 3 (IPT3) showed impairment of nodulation, suggesting that IPT3 is required for nodule development in M. truncatula. Simultaneous live imaging of pIPT3::nls:tdTOMATO and the CK sensor showed that IPT3 induction in the pericycle at the base of nodule primordium contributes to CK biosynthesis, which in turn promotes expression of positive regulators of nodule organogenesis in M. truncatula.

scholarly journals MtBZR1 Plays an Important Role in Nodule Development in Medicago truncatula

2019 ◽  
Vol 20 (12) ◽  
pp. 2941
Author(s):  
Can Cui ◽  
Hongfeng Wang ◽  
Limei Hong ◽  
Yiteng Xu ◽  
Yang Zhao ◽  
...  
Keyword(s):  
Medicago Truncatula ◽  
Sinorhizobium Meliloti ◽  
Normal Growth ◽  
Dry Mass ◽  
Growth Conditions ◽  
Nodule Development ◽  
Functional Study ◽  
Loss Of Function ◽  
Wild Type

Brassinosteroid (BR) is an essential hormone in plant growth and development. The BR signaling pathway was extensively studied, in which BRASSINAZOLE RESISTANT 1 (BZR1) functions as a key regulator. Here, we carried out a functional study of the homolog of BZR1 in Medicago truncatula R108, whose expression was induced in nodules upon Sinorhizobium meliloti 1021 inoculation. We identified a loss-of-function mutant mtbzr1-1 and generated 35S:MtBZR1 transgenic lines for further analysis at the genetic level. Both the mutant and the overexpression lines of MtBZR1 showed no obvious phenotypic changes under normal growth conditions. After S. meliloti 1021 inoculation, however, the shoot and root dry mass was reduced in mtbzr1-1 compared with the wild type, caused by partially impaired nodule development. The transcriptomic analysis identified 1319 differentially expressed genes in mtbzr1-1 compared with wild type, many of which are involved in nodule development and secondary metabolite biosynthesis. Our results demonstrate the role of MtBZR1 in nodule development in M. truncatula, shedding light on the potential role of BR in legume–rhizobium symbiosis.

scholarly journals Specific Host-Responsive Associations Between Medicago truncatula Accessions and Sinorhizobium Strains

2017 ◽  
Vol 30 (5) ◽  
pp. 399-409 ◽  
Author(s):  
Théophile Kazmierczak ◽  
Marianna Nagymihály ◽  
Florian Lamouche ◽  
Quentin Barrière ◽  
Ibtissem Guefrachi ◽  
...  
Keyword(s):  
Medicago Truncatula ◽  
Sinorhizobium Meliloti ◽  
Root Nodules ◽  
Nodule Formation ◽  
Bacterial Strains ◽  
Symbiotic Efficiency ◽  
Model Legume ◽  
Functional Studies ◽  
Multiple Parameters ◽  
Nitrogen Fixation Activity

Legume plants interact with rhizobia to form nitrogen-fixing root nodules. Legume-rhizobium interactions are specific and only compatible rhizobia and plant species will lead to nodule formation. Even within compatible interactions, the genotype of both the plant and the bacterial symbiont will impact on the efficiency of nodule functioning and nitrogen-fixation activity. The model legume Medicago truncatula forms nodules with several species of the Sinorhizobium genus. However, the efficiency of these bacterial strains is highly variable. In this study, we compared the symbiotic efficiency of Sinorhizobium meliloti strains Sm1021, 102F34, and FSM-MA, and Sinorhizobium medicae strain WSM419 on the two widely used M. truncatula accessions A17 and R108. The efficiency of the interactions was determined by multiple parameters. We found a high effectiveness of the FSM-MA strain with both M. truncatula accessions. In contrast, specific highly efficient interactions were obtained for the A17-WSM419 and R108-102F34 combinations. Remarkably, the widely used Sm1021 strain performed weakly on both hosts. We showed that Sm1021 efficiently induced nodule organogenesis but cannot fully activate the differentiation of the symbiotic nodule cells, explaining its weaker performance. These results will be informative for the selection of appropriate rhizobium strains in functional studies on symbiosis using these M. truncatula accessions, particularly for research focusing on late stages of the nodulation process.

scholarly journals Developmental Analysis of the GATA Factor HANABA TARANU Mutants in Medicago truncatula Reveals Their Roles in Nodule Formation

2021 ◽  
Vol 12 ◽  
Author(s):  
Yiteng Xu ◽  
Hongfeng Wang ◽  
Zhichao Lu ◽  
Lizhu Wen ◽  
Zhiqun Gu ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Medicago Truncatula ◽  
Transcriptional Analysis ◽  
Nodule Development ◽  
Root Tip ◽  
Nodule Formation ◽  
Loss Of Function ◽  
Gata Factor ◽  
Nitrate Treatment ◽  
Gata Transcription Factors

Formation of nodules on legume roots results from symbiosis with rhizobial bacteria. Here, we identified two GATA transcription factors, MtHAN1 and MtHAN2, in Medicago truncatula, which are the homologs of HANABA TARANU (HAN) and HANABA TARANU LIKE in Arabidopsis thaliana. Our analysis revealed that MtHAN1 and MtHAN2 are expressed in roots and shoots including the root tip and nodule apex. We further show that MtHAN1 and MtHAN2 localize to the nucleus where they interact and that single and double loss-of-function mutants of MtHAN1 and MtHAN2 did not show any obvious phenotype in flower development, suggesting their role is different than their closest Arabidopsis homologues. Investigation of their symbiotic phenotypes revealed that the mthan1 mthan2 double mutant develop twice as many nodules as wild type, revealing a novel biological role for GATA transcription factors. We found that HAN1/2 transcript levels respond to nitrate treatment like their Arabidopsis counterparts. Global gene transcriptional analysis by RNA sequencing revealed different expression genes enriched for several pathways important for nodule development including flavonoid biosynthesis and phytohormones. In addition, further studies suggest that MtHAN1 and MtHAN2 are required for the expression of several nodule-specific cysteine-rich genes, which they may activate directly, and many peptidase and peptidase inhibitor genes. This work expands our knowledge of the functions of MtHANs in plants by revealing an unexpected role in legume nodulation.

scholarly journals The LATD Gene of Medicago truncatula Is Required for Both Nodule and Root Development

2005 ◽  
Vol 18 (6) ◽  
pp. 521-532 ◽  
Author(s):  
Lydia J. Bright ◽  
Yan Liang ◽  
David M. Mitchell ◽  
Jeanne M. Harris
Keyword(s):  
Medicago Truncatula ◽  
Root Development ◽  
Sinorhizobium Meliloti ◽  
Root Nodules ◽  
Lateral Roots ◽  
Primary Root ◽  
Evolutionary Origin ◽  
Nodule Formation ◽  
Wild Type ◽  
Model Legume

The evolutionary origins of legume root nodules are largely unknown. We have identified a gene,LATD, of the model legume Medicago truncatula, that is required for both nodule and root development, suggesting that these two developmental processes may share a common evolutionary origin. The latd mutant plants initiate nodule formation but do not complete it, resulting in immature, non-nitrogen-fixing nodules. Similarly, lateral roots initiate, but remain shortstumps. The primary root, which initially appears to be wild type, gradually ceases growth and forms an abnormal tipthat resembles that of the mutant lateral roots. Infection by the rhizobial partner, Sinorhizobium meliloti, can occur, although infection is rarely completed. Once inside latd mutant nodules, S. meliloti fails to express rhizobial genes associatedwith the developmental transition from free-living bacterium to endosymbiont, such as bacA and nex38. The infecting rhizobia also fail to express nifH and fix nitrogen. Thus, both plant and bacterial development are blocked in latd mutant roots. Based on the latd mutant phenotype, we propose that the wild-type function of the LATD gene is to maintain root meristems. The strong requirement of both nodules and lateral roots for wild-type LATD gene function supports lateral roots as a possible evolutionary origin for legume nodules.

scholarly journals Salt stress enhances early symbiotic gene expression in Medicago truncatula and induces a stress-specific set of rhizobium-responsive genes.

2021 ◽  
Author(s):  
Sanhita Chakraborty ◽  
Heather Driscoll ◽  
Juan Abrahante Lloréns ◽  
Fan Zhang ◽  
Robert Fisher ◽  
...  
Keyword(s):  
Gene Expression ◽  
Salt Stress ◽  
Medicago Truncatula ◽  
Sinorhizobium Meliloti ◽  
Root Nodules ◽  
Symbiotic Association ◽  
Symbiotic Gene ◽  
Nod Factor ◽  
Early Nodulin ◽  
Regulated Gene Expression

Salt stress is a major agricultural concern inhibiting not only plant growth but also the symbiotic association between legume roots and the soil bacteria rhizobia. This symbiotic association is initiated by a molecular dialogue between the two partners, leading to the activation of a signaling cascade in the legume host and ultimately the formation of nitrogen-fixing root nodules. Here we show that a moderate salt stress increases the responsiveness of early symbiotic genes in Medicago truncatula to its symbiotic partner, Sinorhizobium meliloti, while conversely, inoculation with S. meliloti counteracts salt-regulated gene expression, restoring one-third to control levels. Our analysis of Early Nodulin 11 shows that salt-induced expression is dynamic, Nod-factor dependent, and requires the ionic, but not the osmotic, component of salt. We demonstrate that salt stimulation of rhizobium-induced gene expression requires NSP2, which functions as a node to integrate the abiotic and biotic signals. In addition, our work reveals that inoculation with Sinorhizobium meliloti succinoglycan mutants also hyperinduces ENOD11 expression in the presence or absence of salt, suggesting a possible link between rhizobial exopolysaccharide and the plant response to salt stress. Finally, we identify an accessory set of genes that are induced by rhizobium only under conditions of salt stress and have not been previously identified as being nodulation-related genes. Our data suggests that interplay of core nodulation genes with different accessory sets, specific for different abiotic conditions, function to establish the symbiosis. Together, our findings reveal a complex and dynamic interaction between plant, microbe, and environment.

Effects of a Nod-factor-overproducing strain of Sinorhizobium meliloti on the expression of the ENOD40 gene in Melilotus alba

2002 ◽  
Vol 80 (9) ◽  
pp. 907-915 ◽  
Author(s):  
Walter F Giordano ◽  
Michelle R Lum ◽  
Ann M Hirsch
Keyword(s):  
Lateral Root ◽  
Sinorhizobium Meliloti ◽  
Cortical Cell ◽  
Host Cells ◽  
Nodule Development ◽  
Nodule Formation ◽  
Additional Increase ◽  
Nod Factor ◽  
Melilotus Alba ◽  
Different Strains

We have initiated studies on the molecular biology and genetics of white sweetclover (Melilotus alba Desr.) and its responses to inoculation with the nitrogen-fixing symbiont Sinorhizobium meliloti. Early nodulin genes such as ENOD40 serve as markers for the transition from root to nodule development even before visible stages of nodule formation are evident. Using Northern blot analysis, we found that the ENOD40 gene was expressed within 6 h after inoculation with two different strains of S. meliloti, one of which overproduces symbiotic Nod factors. Inoculation with this strain resulted in an additional increase in ENOD40 gene expression over a typical wild-type S. meliloti strain. Moreover, the increase in mRNA brought about by the Nod-factor-overproducing strain 24 h after inoculation was correlated with lateral root formation by using whole-mount in situ hybridization to localize ENOD40 transcripts in lateral root meristems and by counting lateral root initiation sites. Cortical cell divisions were not detected. We also found that nodulation occurred more rapidly on white sweetclover in response to the Nod-factor-overproducing strain, but ultimately there was no difference in nodulation efficiency in terms of nodule number or the number of roots nodulated by the two strains. Also, the two strains could effectively co-colonize the host when inoculated together, although a few host cells were occupied by both strains.Key words: ENOD40, Nod factor, Melilotus, Sinorhizobium, symbiosis.

scholarly journals Expression of Medicago truncatula Genes Responsive to Nitric Oxide in Pathogenic and Symbiotic Conditions

2008 ◽  
Vol 21 (6) ◽  
pp. 781-790 ◽  
Author(s):  
Alberto Ferrarini ◽  
Matteo De Stefano ◽  
Emmanuel Baudouin ◽  
Chiara Pucciariello ◽  
Annalisa Polverari ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Medicago Truncatula ◽  
Regulatory Networks ◽  
Sinorhizobium Meliloti ◽  
Biotic Interactions ◽  
Hypersensitive Reaction ◽  
Nodule Formation ◽  
Polymorphism Analysis ◽  
Symbiotic Interaction ◽  
Physiological Relevance

Nitric oxide (NO) is involved in diverse physiological processes in plants, including growth, development, response to pathogens, and interactions with beneficial microorganisms. In this work, a dedicated microarray representing the widest database available of NO-related transcripts in plants has been produced with 999 genes identified by a cDNA amplified fragment length polymorphism analysis as modulated in Medicago truncatula roots treated with two NO donors. The microarray then was used to monitor the expression of NO-responsive genes in M. truncatula during the incompatible interaction with the foliar pathogen Colletotrichum trifolii race 1 and during the symbiotic interaction with Sinorhizobium meliloti 1021. A wide modulation of NO-related genes has been detected during the hypersensitive reaction or during nodule formation and is discussed with special emphasis on the physiological relevance of these genes in the context of the two biotic interactions. This work clearly shows that NO-responsive genes behave differently depending on the plant organ and on the type of interaction, strengthening the need to consider regulatory networks, including different signaling molecules.

scholarly journals The Medicago truncatula N5 Gene Encoding a Root-Specific Lipid Transfer Protein Is Required for the Symbiotic Interaction with Sinorhizobium meliloti

2009 ◽  
Vol 22 (12) ◽  
pp. 1577-1587 ◽  
Author(s):  
Youry Pii ◽  
Alessandra Astegno ◽  
Elisa Peroni ◽  
Massimo Zaccardelli ◽  
Tiziana Pandolfini ◽  
...  
Keyword(s):  
Medicago Truncatula ◽  
Sinorhizobium Meliloti ◽  
Lipid Transfer Protein ◽  
Symbiotic Association ◽  
Lipid Transfer ◽  
Symbiotic Interaction ◽  
Transgenic Roots ◽  
Transfer Protein ◽  
Small Proteins

The Medicago truncatula N5 gene is induced in roots after Sinorhizobium meliloti infection and it codes for a putative lipid transfer protein (LTP), a family of plant small proteins capable of binding and transferring lipids between membranes in vitro. Various biological roles for plant LTP in vivo have been proposed, including defense against pathogens and modulation of plant development. The aim of this study was to shed light on the role of MtN5 in the symbiotic interaction between M. truncatula and S. meliloti. MtN5 cDNA was cloned and the mature MtN5 protein expressed in Escherichia coli. The lipid binding capacity and antimicrobial activity of the recombinant MtN5 protein were tested in vitro. MtN5 showed the capacity to bind lysophospholipids and to inhibit M. truncatula pathogens and symbiont growth in vitro. Furthermore, MtN5 was upregulated in roots after infection with either the fungal pathogen Fusarium semitectum or the symbiont S. meliloti. Upon S. meliloti infection, MtN5 was induced starting from 1 day after inoculation (dpi). It reached the highest concentration at 3 dpi and it was localized in the mature nodules. MtN5-silenced roots were impaired in nodulation, showing a 50% of reduction in the number of nodules compared with control roots. On the other hand, transgenic roots overexpressing MtN5 developed threefold more nodules with respect to control roots. Here, we demonstrate that MtN5 possesses biochemical features typical of LTP and that it is required for the successful symbiotic association between M. truncatula and S. meliloti.

scholarly journals PvRACK1 Loss-of-Function Impairs Cell Expansion and Morphogenesis in Phaseolus vulgaris L. Root Nodules

2011 ◽  
Vol 24 (7) ◽  
pp. 819-826 ◽  
Author(s):  
Tania Islas-Flores ◽  
Gabriel Guillén ◽  
Xóchitl Alvarado-Affantranger ◽  
Miguel Lara-Flores ◽  
Federico Sánchez ◽  
...  
Keyword(s):  
Phaseolus Vulgaris ◽  
Cell Expansion ◽  
Root Nodules ◽  
Microscopic Analysis ◽  
Nodule Development ◽  
Loss Of Function ◽  
Rhizobium Tropici ◽  
Transcript Accumulation ◽  
Transgenic Roots ◽  
Plant Signal Transduction

Receptor for activated C kinase (RACK1) is a highly conserved, eukaryotic protein of the WD-40 repeat family. Its peculiar β-propeller structure allows its interaction with multiple proteins in various plant signal-transduction pathways, including those arising from hormone responses, development, and environmental stress. During Phaseolus vulgaris root development, RACK1 (PvRACK1) mRNA expression was induced by auxins, abscissic acid, cytokinin, and gibberellic acid. In addition, during P. vulgaris nodule development, PvRACK1 mRNA was highly accumulated at 12 to 15 days postinoculation, suggesting an important role after nodule meristem initiation and Rhizobium nodule infection. PvRACK1 transcript accumulation was downregulated by a specific RNA interference construct which was expressed in transgenic roots of composite plants of P. vulgaris inoculated with Rhizobium tropici. PvRACK1 downregulated transcript levels were monitored by quantitative reverse-transcription polymerase chain reaction analysis in individual transgenic roots and nodules. We observed a clear phenotype in PvRACK1-knockdown nodules, in which nodule number and nodule cell expansion were impaired, resulting in altered nodule size. Microscopic analysis indicated that, in PvRACK1-knockdown nodules, infected and uninfected cells were considerably smaller (80 and 60%, respectively) than in control nodules. In addition, noninfected cells and symbiosomes in silenced nodules showed significant defects in membrane structure under electron microscopy analysis. These findings indicate that PvRACK1 has a pivotal role in cell expansion and in symbiosome and bacteroid integrity during nodule development.

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