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A Toolkit for High Resolution Imaging of Cell Division and Phytohormone Signaling in Legume Roots and Root Nodules

Legume plants benefit from a nitrogen-fixing symbiosis in association with rhizobia hosted in specialized root nodules. Formation of root nodules is initiated by de novo organogenesis and coordinated infection of these developing lateral root organs by rhizobia. Both bacterial infection and nodule o...

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Autores principales: Nadzieja, Marcin, Stougaard, Jens, Reid, Dugald
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6688427/
https://www.ncbi.nlm.nih.gov/pubmed/31428118
http://dx.doi.org/10.3389/fpls.2019.01000
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author Nadzieja, Marcin
Stougaard, Jens
Reid, Dugald
author_facet Nadzieja, Marcin
Stougaard, Jens
Reid, Dugald
author_sort Nadzieja, Marcin
collection PubMed
description Legume plants benefit from a nitrogen-fixing symbiosis in association with rhizobia hosted in specialized root nodules. Formation of root nodules is initiated by de novo organogenesis and coordinated infection of these developing lateral root organs by rhizobia. Both bacterial infection and nodule organogenesis involve cell cycle activation and regulation by auxin and cytokinin is tightly integrated in the process. To characterize the hormone dynamics and cell division patterns with cellular resolution during nodulation, sensitive and specific sensors suited for imaging of multicellular tissues are required. Here we report a modular toolkit, optimized in the model legume Lotus japonicus, for use in legume roots and root nodules. This toolkit includes synthetic transcriptional reporters for auxin and cytokinin, auxin accumulation sensors and cell cycle progression markers optimized for fluorescent and bright field microscopy. The developed vectors allow for efficient one-step assembly of multiple units using the GoldenGate cloning system. Applied together with a fluorescence-compatible clearing approach, these reporters improve imaging depth and facilitate fluorescence examination in legume roots. We additionally evaluate the utility of the dynamic gravitropic root response in altering the timing and location of auxin accumulation and nodule emergence. We show that alteration of auxin distribution in roots allows for preferential nodule emergence at the outer side of the bend corresponding to a region of high auxin signaling capacity. The presented tools and procedures open new possibilities for comparative mutant studies and for developing a more comprehensive understanding of legume-rhizobia interactions.
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spelling pubmed-66884272019-08-19 A Toolkit for High Resolution Imaging of Cell Division and Phytohormone Signaling in Legume Roots and Root Nodules Nadzieja, Marcin Stougaard, Jens Reid, Dugald Front Plant Sci Plant Science Legume plants benefit from a nitrogen-fixing symbiosis in association with rhizobia hosted in specialized root nodules. Formation of root nodules is initiated by de novo organogenesis and coordinated infection of these developing lateral root organs by rhizobia. Both bacterial infection and nodule organogenesis involve cell cycle activation and regulation by auxin and cytokinin is tightly integrated in the process. To characterize the hormone dynamics and cell division patterns with cellular resolution during nodulation, sensitive and specific sensors suited for imaging of multicellular tissues are required. Here we report a modular toolkit, optimized in the model legume Lotus japonicus, for use in legume roots and root nodules. This toolkit includes synthetic transcriptional reporters for auxin and cytokinin, auxin accumulation sensors and cell cycle progression markers optimized for fluorescent and bright field microscopy. The developed vectors allow for efficient one-step assembly of multiple units using the GoldenGate cloning system. Applied together with a fluorescence-compatible clearing approach, these reporters improve imaging depth and facilitate fluorescence examination in legume roots. We additionally evaluate the utility of the dynamic gravitropic root response in altering the timing and location of auxin accumulation and nodule emergence. We show that alteration of auxin distribution in roots allows for preferential nodule emergence at the outer side of the bend corresponding to a region of high auxin signaling capacity. The presented tools and procedures open new possibilities for comparative mutant studies and for developing a more comprehensive understanding of legume-rhizobia interactions. Frontiers Media S.A. 2019-08-02 /pmc/articles/PMC6688427/ /pubmed/31428118 http://dx.doi.org/10.3389/fpls.2019.01000 Text en Copyright © 2019 Nadzieja, Stougaard and Reid. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Plant Science
Nadzieja, Marcin
Stougaard, Jens
Reid, Dugald
A Toolkit for High Resolution Imaging of Cell Division and Phytohormone Signaling in Legume Roots and Root Nodules
title A Toolkit for High Resolution Imaging of Cell Division and Phytohormone Signaling in Legume Roots and Root Nodules
title_full A Toolkit for High Resolution Imaging of Cell Division and Phytohormone Signaling in Legume Roots and Root Nodules
title_fullStr A Toolkit for High Resolution Imaging of Cell Division and Phytohormone Signaling in Legume Roots and Root Nodules
title_full_unstemmed A Toolkit for High Resolution Imaging of Cell Division and Phytohormone Signaling in Legume Roots and Root Nodules
title_short A Toolkit for High Resolution Imaging of Cell Division and Phytohormone Signaling in Legume Roots and Root Nodules
title_sort toolkit for high resolution imaging of cell division and phytohormone signaling in legume roots and root nodules
topic Plant Science
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6688427/
https://www.ncbi.nlm.nih.gov/pubmed/31428118
http://dx.doi.org/10.3389/fpls.2019.01000
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