Methods for Medium-Scale Study of Biological Effects of Strigolactone-Like Molecules on the Moss Physcomitrium (Physcomitrella) patens

Guillory, Ambre; Bonhomme, Sandrine

doi:10.1007/978-1-0716-1429-7_12

Cited by 5 publications

(5 citation statements)

References 6 publications

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“…In the moss P. patens , the biological activity of SL analogs was previously assayed by counting the number of filaments per plant, grown for 2 weeks in the dark following compound application (Guillory and Bonhomme, 2021 ). Both (±)-GR24 and (+)-GR24 enantiomer led to a decrease in filament number, in WT plants and in the Ppccd8 mutant, where the activity was more pronounced (Hoffmann et al, 2014 ; Lopez-Obando et al, 2021 ).…”

Section: Resultsmentioning

confidence: 99%

“…Assays on Physcomitrium patens were performed on plants grown in 24-well plates, starting from very small pieces of moss tissues as described by Guillory and Bonhomme ( 2021 ). As for pea rms1 , the Ppccd8 SL synthesis mutant was used for assays, since the effect of the compounds was better seen in this mutant vs. wild type (WT; Lopez-Obando et al, 2021 ).…”

Section: Methodsmentioning

confidence: 99%

“…A single picture of each well was taken under an Axio Zoom microscope (Zeiss) with a dedicated program. Filaments were counted using ImageJ software ( http://imagej.nih.gov/ij/ ) as described by Guillory and Bonhomme ( 2021 ). Twenty-four plants were tested in each treatment.…”

Section: Methodsmentioning

confidence: 99%

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Expansion of the Strigolactone Profluorescent Probes Repertory: The Right Probe for the Right Application

et al. 2022

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Strigolactones (SLs) are intriguing phytohormones that not only regulate plant development and architecture but also interact with other organisms in the rhizosphere as root parasitic plants (Striga, Orobanche, and Phelipanche) and arbuscular mycorrhizal fungi. Starting with a pioneering work in 2003 for the isolation and identification of the SL receptor in parasitic weeds, fluorescence labeling of analogs has proven a major strategy to gain knowledge in SL perception and signaling. Here, we present novel chemical tools for understanding the SL perception based on the enzymatic properties of SL receptors. We designed different profluorescent SL Guillaume Clavé (GC) probes and performed structure-activity relationship studies on pea, Arabidopsis thaliana, and Physcomitrium (formerly Physcomitrella) patens. The binding of the GC probes to PsD14/RMS3, AtD14, and OsD14 proteins was tested. We demonstrated that coumarin-based profluorescent probes were highly bioactive and well-adapted to dissect the enzymatic properties of SL receptors in pea and a resorufin profluorescent probe in moss, contrary to the commercially available fluorescein profluorescent probe, Yoshimulactone Green (YLG). These probes offer novel opportunities for the studies of SL in various plants.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Expansion of the Strigolactone Profluorescent Probes Repertory: The Right Probe for the Right Application

et al. 2022

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“…For red light experiments, plants were grown on PpNO 3 medium in Magenta pots at 25°C, in continuous red-light (∼45 µmol µmol/m 2 /s). Cellophanes of appropriate sizes were used for monitoring of protonema extension and branching, as well as for the cultures launched in 6-well plates for gene expression studies (Guillory and Bonhomme 2021). Analysis of caulonema growth in the dark was performed in 24-well plates, with ∼2 weeks of growth in control conditions before incubation (± treatment) in the dark and placed vertically for ∼10 days (Guillory and Bonhomme 2021).…”

Section: Methodsmentioning

confidence: 99%

“…P. patens protonema branching is inhibited by auxin (Thelander et al 2018), but promoted by cytokinins (Cove et al 2006). Auxin also positively regulates the chloronema to caulonema filament transition, while cytokinins positively regulate bud initiation (for a review: Guillory and Bonhomme, 2021). Four AP2-type transcriptional factors, the APB genes, are central players in this hormonal crosstalk that regulates protonema cell identity and division (Aoyama et al 2012).…”

Section: Smxl Proteins Likely Retained a Common Molecular Function Al...mentioning

confidence: 99%

Physcomitrium patensSMXL homologs are PpMAX2-dependent negative regulators of growth

Guillory

López-Obando

Bouchenine

et al. 2023

Preprint

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SMXL proteins are a plant-specific clade of type I HSP100/Clp-ATPases. SMXL genes are found in virtually all land plant genomes. However, they have mainly been studied in angiosperms. In Arabidopsis thaliana, three SMXL functional subclades have been identified: SMAX1/SMXL2, SMXL345 and SMXL678. Out of these, two subclades ensure transduction on endogenous hormone signals: SMAX1/SMXL2 are involved in KAI2-ligand (KL) signaling, while SMXL678 are involved in strigolactones (SLs) signaling. Many questions remain regarding the mode of action of these proteins, as well as their ancestral role. In light of recent discoveries in the liverwort Marchantia polymorpha, we addressed this second question by investigating the function of the four SMXL genes of the moss Physcomitrium patens. We demonstrate that PpSMXL proteins are negative regulators of growth, involved in the likely conserved ancestral MAX2-dependent KL signaling pathway. However, PpSMXL proteins expressed in A. thaliana unexpectedly cannot replace SMAX1/SMXL2 function in KL signaling, whereas they can functionally replace SMXL4/5 and restore root growth. Therefore, the molecular function of SMXL could be conserved, but not their interaction network. Moreover, one PpSMXL clade also positively regulates transduction of the SL signal in P. patens, this function most probably having an independent evolutionary origin to angiosperms SMXL678.

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The Physcomitrium (Physcomitrella) patens PpKAI2L receptors for strigolactones and related compounds function via MAX2-dependent and -independent pathways

et al. 2021

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In angiosperms, the α/β hydrolase DWARF14 (D14), along with the F-box protein MORE AXILLARY GROWTH2 (MAX2), perceives strigolactones (SL) to regulate developmental processes. The key SL biosynthetic enzyme CAROTENOID CLEAVAGE DIOXYGENASE8 (CCD8) is present in the moss Physcomitrium patens, and PpCCD8-derived compounds regulate moss extension. The PpMAX2 homolog is not involved in the SL response, but 13 PpKAI2LIKE (PpKAI2L) genes homologous to the D14 ancestral paralog KARRIKIN INSENSITIVE2 (KAI2) encode candidate SL receptors. In Arabidopsis thaliana, AtKAI2 perceives karrikins and the elusive endogenous KAI2-Ligand (KL). Here, germination assays of the parasitic plant Phelipanche ramosa suggested that PpCCD8-derived compounds are likely non-canonical SLs. (+)-GR24 SL analog is a good mimic for PpCCD8-derived compounds in P. patens, while the effects of its enantiomer (−)-GR24, a KL mimic in angiosperms, are minimal. Interaction and binding assays of seven PpKAI2L proteins pointed to the stereoselectivity towards (−)-GR24 for a single clade of PpKAI2L (eu-KAI2). Enzyme assays highlighted the peculiar behavior of PpKAI2L-H. Phenotypic characterization of Ppkai2l mutants showed that eu-KAI2 genes are not involved in the perception of PpCCD8-derived compounds but act in a PpMAX2-dependent pathway. By contrast, mutations in PpKAI2L-G, and -J genes abolished the response to the (+)-GR24 enantiomer, suggesting that PpKAI2L-G, and -J proteins are receptors for moss SLs.

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Methods for Medium-Scale Study of Biological Effects of Strigolactone-Like Molecules on the Moss Physcomitrium (Physcomitrella) patens

Cited by 5 publications

References 6 publications

Expansion of the Strigolactone Profluorescent Probes Repertory: The Right Probe for the Right Application

Expansion of the Strigolactone Profluorescent Probes Repertory: The Right Probe for the Right Application

Physcomitrium patensSMXL homologs are PpMAX2-dependent negative regulators of growth

The Physcomitrium (Physcomitrella) patens PpKAI2L receptors for strigolactones and related compounds function via MAX2-dependent and -independent pathways

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