Genetically encoded calcium indicators for fluorescence imaging in the moss <i>Physcomitrella</i>: <scp>GC</scp>a<scp>MP</scp>3 provides a bright new look

Kleist, Thomas; Cartwright, Heather; Perera, Adele; Christianson, Michael L.; Lemaux, Peggy G.; Luan, Sheng

doi:10.1111/pbi.12769

Cited by 19 publications

(12 citation statements)

References 10 publications

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“…Recently developed genetically encoded calcium indicator probes allow to trace calcium modulations in vivo via FRET (Förster Energy Resonance Transfer) or GFP-linked techniques. At their core, each sensor contains an engineered calcium binding calmodulin variant and calmodulin-binding peptide fused to one or two fluorophores that optically report Ca 2+ -dependent conformational changes [33]. Bascom et al [21].…”

Section: Discussionmentioning

confidence: 99%

Stay in Touch—The Cortical ER of Moss Protonemata in Osmotic Stress Situations

Harant

Lang

2020

Plants

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Plasmolysis is usually introduced to cell biology students as a tool to illustrate the plasma membrane: hypertonic solutions cause the living protoplast to shrink by osmotic water loss; hence, it detaches from the surrounding cell wall. What happens, however, with the subcellular structures in the cell cortex during this process of turgor loss? Here, we investigated the cortical endoplasmic reticulum (ER) in moss protonema cells of Physcomitrella patens in a cell line carrying a transgenic ER marker (GFP-HDEL). The plasma membrane was labelled simultaneously with the fluorescent dye FM4-64 to achieve structural separation. By placing the protonemata in a hypertonic mannitol solution (0.8 M), we were able to follow the behaviour of the cortical ER and the protoplast during plasmolysis by confocal laser scanning microscopy (CLSM). The protoplast shape and structural changes of the ER were further examined after depolymerisation of actin microfilaments with latrunculin B (1 µM). In its natural state, the cortical ER is a dynamic network of fine tubes and cisternae underneath the plasma membrane. Under acute and long-term plasmolysis (up to 45 min), changes in the protoplast form and the cortical ER, as well as the formation of Hechtian strands and Hechtian reticula, were observed. The processing of the high-resolution z-scans allowed the creation of 3D models and gave detailed insight into the ER of living protonema cells before, during and after plasmolysis.

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Section: Discussionmentioning

confidence: 99%

Stay in Touch—The Cortical ER of Moss Protonemata in Osmotic Stress Situations

Harant

Lang

2020

Plants

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“…Dual excitation ratiometric MatryoshCaMP6s and R‐GECO1‐mTurquoise sensors have been recently generated and successfully expressed in Arabidopsis plants (Ast et al, ; Waadt, Krebs, Kudla, & Schumacher, ). For the sake of clarity, we can state that intensiometric biosensors exhibit a higher signal change compared with a FRET‐based sensor (e.g., Cameleon) in response to several stimuli (Keinath et al, ; Kleist et al, ; Nguyen, Kurenda, Stolz, Chételat, & Farmer, ; Toyota et al, ; Vincent et al, ; Waadt et al, ). However, at the present time, the Cameleon sensor, thanks to its FRET‐based ratiometric properties, has shown to be particularly suited for the quantification of Ca 2+ levels in different genetic backgrounds (wild type vs. mutants; Behera et al,; Corso, Doccula, de Melo, Costa, & Verbruggen, ; Costa et al, ; Shkolnik, Nuriel, Bonza, Costa, & Fromm, ) as well as different growth conditions within the same genotype (e.g., Behera et al, ; Matthus et al, ; Quiles‐Pando et al, ; Tian, Zhang, Yang, Wang, & Zhang, ) so is thus probably more indicated for the study of Ca 2+ dynamics/homeostasis in different nutritional conditions.…”

Section: Methods To Measure Ca2+ Dynamics In Vivomentioning

confidence: 99%

Harnessing the new emerging imaging technologies to uncover the role of Ca²⁺ signalling in plant nutrient homeostasis

Vigani

Costa

2019

Plant Cell & Environment

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Increasing crop yields by using ecofriendly practices is of high priority to tackle problems regarding food security and malnutrition worldwide. A sustainable crop production requires a limited use of fertilizer and the employment of plant varieties with improved ability to acquire nutrients from soil. To reach these goals, the scientific community aims to understand plant nutrients homeostasis by deciphering the nutrient sensing and signalling mechanisms of plants. Several lines of evidence about the involvement of Ca 2+ as the signal of an impaired nutrient availability have been reported. Ca 2+ signalling is a tightly regulated process that requires specific protein toolkits to perceive external stimuli and to induce the specific responses in the plant needed to survive. Here, we summarize both older and recent findings concerning the involvement of Ca 2+ signalling in the homeostasis of nutrients. In this review, we present new emerging technologies, based on the use of genetically encoded Ca 2+ sensors and advanced microscopy, which offer the chance to perform in planta analyses of Ca 2+ dynamics at cellular resolution. The harnessing of these technologies with different genetic backgrounds and subjected to different nutritional stresses will provide important insights to the still little-known mechanisms of nutrient sensing in plants.

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“…The necessary cell-cell (apoplastic e.g. Ca 2+ -mediated) or cell-to-cell (plasmodesmata-mediated) communication makes regeneration an attractive developmental Model Organisms in Plant Genetics process to study this cell crosstalk [71][72][73]. The phytohormone ABA is a key responsible of the dynamic regulation of the permeability of plasmodesmata in response to changing environments, such as wounding.…”

Section: Regenerationmentioning

confidence: 99%

Mosses: Accessible Systems for Plant Development Studies

Floriach-Clark¹,

Tang²,

Willemsen³

2022

Model Organisms in Plant Genetics

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Mosses are a cosmopolitan group of land plants, sister to vascular plants, with a high potential for molecular and cell biological research. The species Physcomitrium patens has helped gaining better understanding of the biological processes of the plant cell, and it has become a central system to understand water-to-land plant transition through 2D-to-3D growth transition, regulation of asymmetric cell division, shoot apical cell establishment and maintenance, phyllotaxis and regeneration. P. patens was the first fully sequenced moss in 2008, with the latest annotated release in 2018. It has been shown that many gene functions and networks are conserved in mosses when compared to angiosperms. Importantly, this model organism has a simplified and accessible body structure that facilitates close tracking in time and space with the support of live cell imaging set-ups and multiple reporter lines. This has become possible thanks to its fully established molecular toolkit, with highly efficient PEG-assisted, CRISPR/Cas9 and RNAi transformation and silencing protocols, among others. Here we provide examples on how mosses exhibit advantages over vascular plants to study several processes and their future potential to answer some other outstanding questions in plant cell biology.

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Genetically encoded calcium indicators for fluorescence imaging in the moss Physcomitrella: GCaMP3 provides a bright new look

Cited by 19 publications

References 10 publications

Stay in Touch—The Cortical ER of Moss Protonemata in Osmotic Stress Situations

Stay in Touch—The Cortical ER of Moss Protonemata in Osmotic Stress Situations

Harnessing the new emerging imaging technologies to uncover the role of Ca²⁺ signalling in plant nutrient homeostasis

Mosses: Accessible Systems for Plant Development Studies

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Genetically encoded calcium indicators for fluorescence imaging in the moss Physcomitrella: GCaMP3 provides a bright new look

Cited by 19 publications

References 10 publications

Stay in Touch—The Cortical ER of Moss Protonemata in Osmotic Stress Situations

Stay in Touch—The Cortical ER of Moss Protonemata in Osmotic Stress Situations

Harnessing the new emerging imaging technologies to uncover the role of Ca2+ signalling in plant nutrient homeostasis

Mosses: Accessible Systems for Plant Development Studies

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Product

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Harnessing the new emerging imaging technologies to uncover the role of Ca²⁺ signalling in plant nutrient homeostasis