Isolation of dimorphic chloroplasts from the single-cell C4 species Bienertia sinuspersici

Lung, Shiu‐Cheung; Yanagisawa, Makoto; Chuong, Simon D. X.

doi:10.1186/1746-4811-8-8

Cited by 6 publications

(3 citation statements)

References 24 publications

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“…Protoplasts during isolation from the donor tissues undergo various stresses, resulting in physical damage and modification of physiological properties of cells, reflected in decreased viability. It has been shown that a rapid and irreversible increase in the permeability of plasma membranes results in protoplast bursting, electrolyte leakage from the cells, and often cell death (Nolte et al 1990;Lung et al 2012). The decrease of protoplast viability might be a result of the damage in plasma membranes during enzyme digestion and purification (Watanabe et al 2002) but also osmotic stress, or oxidative stress during isolation and culture establishment (Tiburcio et al 1986).…”

Section: Discussionmentioning

confidence: 99%

Peptide Growth Factor Phytosulfokine-α Stimulates Cell Divisions and Enhances Regeneration from B. oleracea var. capitata L. Protoplast Culture

Kiełkowska

Adamus

2018

J Plant Growth Regul

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Protoplasts of six cabbage accessions were isolated from leaf mesophyll and cultured in the presence of 0.01, 0.1 and 1.0 µM phytosulfokine-α (PSK-α) and in a PSK-free control medium. PSK-α was applied for 10 days and later, protoplast-derived cells were cultured in the PSK-free medium. Supplementation of the culture medium with PSK-α showed a dose-dependent effect on the mitotic activity of cultured cells. On the 15th day of culture, the highest mitotic activity of protoplast-derived cells was observed in cultures treated with 0.1 µM of PSK-α, and ranged from 14 to 60% dependent on the accession. The number of multi-cell structures was also higher (90-93%) on this medium compared to the control (77-80%). Analysis of cellulose regeneration in cultured protoplasts after Calcofluor White staining showed that this process was not synchronous, but depended instead on the presence of PSK-α in the culture medium, and was more pronounced in the low-responding accession. Sustained cell divisions led to formation of microcallus colonies, subjected to regeneration on solid media. Supplementation of the regeneration media with 0.1 µM of PSK significantly increased shoot regeneration compared to the control media. Moreover, enhanced regeneration was observed from calluses developed from cells treated with PSK-α at the early stages of development and later transferred for regeneration onto the media supplemented with 0.1 µM of this peptide.

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Section: Discussionmentioning

confidence: 99%

Peptide Growth Factor Phytosulfokine-α Stimulates Cell Divisions and Enhances Regeneration from B. oleracea var. capitata L. Protoplast Culture

Kiełkowska

Adamus

2018

J Plant Growth Regul

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“…Relative expression levels of G protein subunits, LOX1, COI1, PAL , and PR1 were determined by quantitative real-time PCR (Q-RT-PCR) according to Webling and Panstruga (2012) . Total RNA was extracted from 200 mg leaf tissue thrice from each treatment by using RNeasy plant mini kit (Qiagen).…”

Section: Methodsmentioning

confidence: 99%

Pseudomonas fluorescens and Trichoderma asperellum Enhance Expression of Gα Subunits of the Pea Heterotrimeric G-protein during Erysiphe pisi Infection

et al. 2016

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We investigated the transcript accumulation patterns of all three subunits of heterotrimeric G-proteins (Gα1 and 2, Gβ, and Gγ) in pea under stimulation of two soil-inhabiting rhizosphere microbes Pseudomonas fluorescens OKC and Trichoderma asperellum T42. The microbes were either applied individually or co-inoculated and the transcript accumulation patterns were also investigated after challenging the same plants with a fungal biotrophic pathogen Erysiphe pisi. We observed that mostly the transcripts of Gα 1 and 2 subunits were accumulated when the plants were treated with the microbes (OKC and T42) either individually or co-inoculated. However, transcript accumulations of Gα subunits were highest in the T42 treatment particularly under the challenge of the biotroph. Transcript accumulations of the other two subunits Gβ and Gγ were either basal or even lower than the basal level. There was an indication for involvement of JA-mediated pathway in the same situations as activation of LOX1 and COI1 were relatively enhanced in the microbe co-inoculated treatments. Non-increment of SA content as well as transcripts of SA-dependent PR1 suggested non-activation of the SA-mediated signal transduction in the interaction of pea with E. pisi under the stimuli of OKC and T42. Gα1 and 2 transcript accumulations were further correlated with peroxidases activities, H2O2 generation and accumulation in ABA in pea leaves under OKC and T42 stimulations and all these activities were positively correlated with stomata closure at early stage of the biotroph challenge. The microbe-induced physiological responses in pea leaves finally led to reduced E. pisi development particularly in OKC and T42 co-inoculated plants. We conclude that OKC and T42 pretreatment stimulate transcript accumulations of the Gα1 and Gα2 subunits of the heterotrimeric G protein, peroxidases activities and phenol accumulation in pea during infection by E. pisi. The signal transduction was possibly mediated through JA in pea under the stimulus of the microbes and the cumulative effect of the co-inoculated microbes had a suppressive effect on E. pisi conidial development on pea leaves.

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“…Briefly, the isolated chlorenchyma cells of B. sinuspersici were incubated in an enzyme solution containing 1.5 % ( w / v ) cellulase Onozuka R10 (Yakult Honsha Co. Ltd., Tokyo, Japan) and 0.1% ( w / v ) BSA in cell-stabilizing (CS) solution [0.7 M sucrose, 25 mM HEPES-KOH (pH 6.5),5 mM KCl and 1mM CaCl 2 ] for 4 h in the dark without shaking. A homogeneous population of protoplasts was collected by washing or floating them twice on top the CS medium followed by a 2 min centrifugation step at 100 g. The purification of dimorphic chloroplasts from isolated chlorenchyma protoplasts of B. sinuspersici was obtained with the hypo-osmotic shock method according to Lung et al (2012) [ 59 ]. Total RNA was extracted from purified chloroplasts using TRIzol Reagent (Invitrogen, Burlington, Canada) according to the manufacturer’s instruction.…”

Section: Methodsmentioning

confidence: 99%

Development of C4 Biochemistry and Change in Expression of Markers for Photosystems I and II in the Single-Cell C4 Species, Bienertia sinuspersici

Yanagisawa

Chuong

2022

Plants

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Bienertia sinuspersici is one of four identified terrestrial plants that perform C4 photosynthesis within a single chlorenchyma cell via the compartmentation of organelles and photosynthetic enzymes. The patterns of accumulation of key photosynthetic enzymes and transcripts in developing leaves were examined using immunolocalization and in situ hybridization. The polypeptides of Rubisco large subunit (RbcL) and pyruvate Pi dikinase (PPDK) accumulated equally in all chloroplasts before the formation of two intracellular cytoplasmic compartments: the central (CCC) and peripheral (PCC) cytoplasmic compartments. The differential accumulation of these enzymes was not completed until the leaf had reached maturity, indicating that the transition from C3 to C4 photosynthesis occurred during leaf maturation. In mature chlorenchyma cells, RbcL accumulated 20-fold higher in the CCC than in the PCC, while PPDK exhibited a concentration gradient that was the lowest in the chloroplasts in the central region of the CCC and the highest in PCC chloroplasts. The pattern of rbcL transcript accumulation followed that of its polypeptides in developing leaves, suggesting that the expression of this gene was likely controlled by transcriptional and/or post-transcriptional processes. Immunocytochemical results examining the distribution of photosystems I and II in the chloroplasts of chlorenchyma cells from mature leaves showed that PSII is more abundant in chloroplasts of the central compartment, whereas PSI is higher in those of the peripheral compartment. The quantitative real-time PCR results of rbcL, psbA, and psaB transcripts from the isolated chloroplasts of each compartment further supported this observation. Our results suggest that multiple levels of regulation play a role in controlling the differential accumulation of photosynthetic gene expression in the dimorphic chloroplasts of single-cell C4 species during leaf development.

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Isolation of dimorphic chloroplasts from the single-cell C4 species Bienertia sinuspersici

Cited by 6 publications

References 24 publications

Peptide Growth Factor Phytosulfokine-α Stimulates Cell Divisions and Enhances Regeneration from B. oleracea var. capitata L. Protoplast Culture

Peptide Growth Factor Phytosulfokine-α Stimulates Cell Divisions and Enhances Regeneration from B. oleracea var. capitata L. Protoplast Culture

Pseudomonas fluorescens and Trichoderma asperellum Enhance Expression of Gα Subunits of the Pea Heterotrimeric G-protein during Erysiphe pisi Infection

Development of C4 Biochemistry and Change in Expression of Markers for Photosystems I and II in the Single-Cell C4 Species, Bienertia sinuspersici

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