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

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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Aspartate Aminotransferase in Alfalfa Nodules: Localization of mRNA During Effective and Ineffective Nodule Development and Promoter Analysis

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi.1999.12.4.263 ◽

1999 ◽

Vol 12 (4) ◽

pp. 263-274 ◽

Cited By ~ 11

Author(s):

Hirofumi Yoshioka ◽

Robert G. Gregerson ◽

Deborah A. Samac ◽

Kim C. M. Hoevens ◽

Gian Trepp ◽

...

Keyword(s):

Aspartate Aminotransferase ◽

Root Nodules ◽

Critical Role ◽

Nodule Development ◽

Vascular Bundles ◽

Fixed Nitrogen ◽

Transcript Accumulation ◽

Specific Expression ◽

Promoter Deletion Analysis

Aspartate aminotransferase (AAT) plays a critical role in the assimilation of symbiotically fixed nitrogen into aspartate and asparagine in legume root nodules. The enzyme occurs as a cytosolic form (AAT1) and a plastid form (AAT2) in alfalfa nodules. To elucidate the functional role of each isozyme in root nodule metabolism further, in situ hybridization was used to determine the pattern of transcript accumulation from the two genes. AAT2 transcripts were localized to infected cells throughout the symbiotic zone of effective alfalfa nodules; however, expression was reduced in ineffective nodules. The AAT1 gene was expressed in the uninfected cells of the invasion zone and symbiotic zone, the nodule parenchyma, and nodule vascular bundles of both effective and ineffective nodules. The AAT1 and AAT2 promoters were evaluated in transgenic alfalfa plants containing promoter β-glucuronidase (GUS) gene fusions. Histochemical staining patterns agreed with results from in situ localization. The distribution pattern of gene transcripts suggests that AAT1 has a role in maintenance of the O2 diffusion barrier in nodules and that AAT2 plays a major role in assimilation of recently fixed nitrogen. Promoter deletion analysis of the AAT2 promoter revealed that nodule-specific expression was retained in a promoter fragment of 300 bp.

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Occurrence of polyamines in root nodules of Phaseolus vulgaris in symbiosis with Rhizobium tropici in response to salt stress

Phytochemistry ◽

10.1016/j.phytochem.2014.08.017 ◽

2014 ◽

Vol 107 ◽

pp. 32-41 ◽

Cited By ~ 13

Author(s):

Miguel López-Gómez ◽

Libertad Cobos-Porras ◽

Javier Hidalgo-Castellanos ◽

Carmen Lluch

Keyword(s):

Salt Stress ◽

Phaseolus Vulgaris ◽

Root Nodules ◽

Rhizobium Tropici

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

Plant Physiology ◽

10.1093/plphys/kiab447 ◽

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.

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Transcriptome analysis of the differential effect of the NADPH oxidase gene RbohB in Phaseolus vulgaris roots following Rhizobium tropici and Rhizophagus irregularis inoculation

BMC Genomics ◽

10.1186/s12864-019-6162-7 ◽

2019 ◽

Vol 20 (1) ◽

Cited By ~ 3

Author(s):

Citlali Fonseca-García ◽

Alejandra E. Zayas ◽

Jesús Montiel ◽

Noreide Nava ◽

Federico Sánchez ◽

...

Keyword(s):

Cell Wall ◽

Phaseolus Vulgaris ◽

Genetic Interaction ◽

Am Fungi ◽

Rhizophagus Irregularis ◽

Ros Scavenging ◽

Rhizobium Tropici ◽

Oxidase Gene ◽

Transgenic Roots ◽

Early Stages

Abstract Background Reactive oxygen species (ROS) are generated by NADPH oxidases known as respiratory burst oxidase homologs (RBOHs) in plants. ROS regulate various cellular processes, including the mutualistic interactions between legumes and nitrogen-fixing bacteria or arbuscular mycorrhizal (AM) fungi. Rboh is a multigene family comprising nine members (RbohA–I) in common bean (Phaseolus vulgaris). The RNA interference-mediated silencing of RbohB (PvRbohB-RNAi) in this species diminished its ROS production and greatly impaired nodulation. By contrast, the PvRbohB-RNAi transgenic roots showed early hyphal root colonization with enlarged fungal hypopodia; therefore, we proposed that PvRbohB positively regulates rhizobial infection (Rhizobium tropici) and inhibits AM colonization by Rhizophagus irregularis in P. vulgaris. Results To corroborate this hypothesis, an RNA-Seq transcriptomic analysis was performed to identify the differentially expressed genes in the PvRbohB-RNAi roots inoculated with Rhizobium tropici or Rhizophagus irregularis. We found that, in the early stages, root nodule symbioses generated larger changes of the transcriptome than did AM symbioses in P. vulgaris. Genes related to ROS homeostasis and cell wall flexibility were markedly upregulated in the early stages of rhizobial colonization, but not during AM colonization. Compared with AM colonization, the rhizobia induced the expression of a greater number of genes encoding enzymes involved in the metabolism of auxins, cytokinins, and ethylene, which were typically repressed in the PvRbohB-RNAi roots. Conclusions Our research provides substantial insights into the genetic interaction networks in the early stages of rhizobia and AM symbioses with P. vulgaris, as well as the differential roles that RbohB plays in processes related to ROS scavenging, cell wall remodeling, and phytohormone homeostasis during nodulation and mycorrhization in this legume.

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Carbon Metabolism in Developing Soybean Root Nodules: The Role of Carbonic Anhydrase

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi.2000.13.1.14 ◽

2000 ◽

Vol 13 (1) ◽

pp. 14-22 ◽

Cited By ~ 31

Author(s):

Nektarios Kavroulakis ◽

Emanouil Flemetakis ◽

Georgios Aivalakis ◽

Panagiotis Katinakis

Keyword(s):

In Situ Hybridization ◽

Carbonic Anhydrase ◽

Root Nodules ◽

Cell Types ◽

Nodule Development ◽

Vascular Bundles ◽

Transcript Accumulation ◽

Cortical Cells ◽

Gene Transcripts

A full-length cDNA clone encoding carbonic anhydrase (CA) was isolated from a soybean nodule cDNA library. In situ hybridization and immunolocalization were performed in order to assess the location of CA transcripts and protein in developing soybean nodules. CA transcripts and protein were present at high levels in all cell types of young nodules, whereas in mature nodules they were absent from the central tissue and were concentrated in cortical cells. The results suggested that, in the earlier stages of nodule development, CA might facilitate the recycling of CO2 while at later stages it may facilitate the diffusion of CO2 out of the nodule system. In parallel, sucrose metabolism was investigated by examination of the temporal and spatial transcript accumulation of sucrose synthase (SS) and phosphoenolpyruvate carboxylase (PEPC) genes, with in situ hybridization. In young nodules, high levels of SS gene transcripts were found in the central tissue as well as in the parenchymateous cells and the vascular bundles, while in mature nodules the levels of SS gene transcripts were much lower, with the majority of the transcripts located in the parenchyma and the pericycle cells of the vascular bundles. High levels of expression of PEPC gene transcripts were found in mature nodules, in almost all cell types, while in young nodules lower levels of transcripts were detected, with the majority of them located in parenchymateous cells as well as in the vascular bundles. These data suggest that breakdown of sucrose may take place in different sites during nodule development.

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

10.1101/2021.04.23.441163 ◽

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

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Rhizobium paranaense sp. nov., an effective N2-fixing symbiont of common bean (Phaseolus vulgaris L.) with broad geographical distribution in Brazil

INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY ◽

10.1099/ijs.0.064543-0 ◽

2014 ◽

Vol 64 (Pt_9) ◽

pp. 3222-3229 ◽

Cited By ~ 41

Author(s):

Rebeca Fuzinatto Dall’Agnol ◽

Renan Augusto Ribeiro ◽

Jakeline Renata Marçon Delamuta ◽

Ernesto Ormeño-Orrillo ◽

Marco Antonio Rogel ◽

...

Keyword(s):

Phaseolus Vulgaris ◽

Common Bean ◽

Novel Species ◽

Sequence Similarity ◽

Root Nodules ◽

Biochemical Properties ◽

Rhizobium Tropici ◽

Content Type ◽

Link Type ◽

Whole Genomes

Nitrogen (N), the nutrient most required for plant growth, is key for good yield of agriculturally important crops. Common bean (Phaseolus vulgaris L.) can benefit from bacteria collectively called rhizobia, which are capable of fixing atmospheric nitrogen (N2) in root nodules and supplying it to the plant. Common bean is amongst the most promiscuous legume hosts; several described species, in addition to putative novel ones have been reported as able to nodulate this legume, although not always effectively in terms of fixing N2. In this study, we present data indicating that Brazilian strains PRF 35T, PRF 54, CPAO 1135 and H 52, currently classified as Rhizobium tropici , represent a novel species symbiont of common bean. Morphological, physiological and biochemical properties differentiate these strains from other species of the genus Rhizobium , as do BOX-PCR profiles (less than 60 % similarity), multilocus sequence analysis with recA, gyrB and rpoA (less than 96.4 % sequence similarity), DNA–DNA hybridization (less than 50 % DNA–DNA relatedness), and average nucleotide identity of whole genomes (less than 92.8.%). The novel species is effective in nodulating and fixing N2 with P. vulgaris, Leucaena leucocephala and Leucaena esculenta. We propose the name Rhizobium paranaense sp. nov. for this novel taxon, with strain PRF 35T ( = CNPSo 120T = LMG 27577T = IPR-Pv 1249T) as the type strain.

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Actin Monoubiquitylation Is Induced in Plants in Response to Pathogens and Symbionts

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi.2001.14.11.1267 ◽

2001 ◽

Vol 14 (11) ◽

pp. 1267-1273 ◽

Cited By ~ 25

Author(s):

Edgar Dantán-González ◽

Yvonne Rosenstein ◽

Carmen Quinto ◽

Federico Sánchez

Keyword(s):

Phaseolus Vulgaris ◽

Posttranslational Modifications ◽

Plant Pathogens ◽

Viral Infections ◽

Root Nodules ◽

Nodule Development ◽

Actin Reorganization ◽

Culture Cells ◽

Microbe Interactions ◽

Host Microbe Interactions

Most dramatic examples of actin reorganization have been described during host-microbe interactions. Plasticity of actin is, in part, due to posttranslational modifications such as phosphorylation or ubiquitylation. Here, we show for the first time that actins found in root nodules of Phaseolus vulgaris are modified transiently during nodule development by monoubiquitylation. This finding was extended to root nodules of other legumes and to other plants infected with mycorrhiza or plant pathogens such as members of the genera Pseudomonas and Phytophthora. However, neither viral infections nor diverse stressful conditions (heat shock, wounding, or osmotic stress) induced this response. Additionally, this phenomenon was mimicked by the addition of a yeast elicitor or H2O2 to Phaseolus vulgaris suspension culture cells. This modification seems to provide increased stability of the microfilaments to proteolytic degradation and seems to be found in fractions in which the actin cytoskeleton is associated with membranes. All together, these data suggest that actin monoubiquitylation may be considered an effector mechanism of a general plant response against microbes.

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