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Cytological changes and alterations in polyamine contents induced by cadmium in tobacco BY-2 cells
Andrea Kuthanová1, Lenka Gemperlová, Sylva Zelenková
1Department of Plant Physiology, Faculty of Sciences, Charles University, Vinicná 5, 128 44 Prague 2, Czech Republic.
Abstract:
Changes in cell viability, proliferation, cell and nuclear morphology including nuclear and DNA fragmentation induced by 0.05 and 1 mM CdSO4 (Cd2+) in tobacco BY-2 cell line (Nicotiana tabacum L.) were studied in the course of 7 days. Simultaneously changes in endogenous contents of both free and conjugated forms of polyamines (PAs) were investigated for 3 days. The application of 0.05 mM Cd2+ evoked decline of cell viability to approximately 60% during the first 24 h of treatment. Later on degradation of cytoplasmic strands, formation of the stress granules and vesicles, modifications in size and shape of the nuclei, including their fragmentation, were observed in the surviving cells. Their proliferation was blocked and cells elongated. Beginning the first day of treatment TUNEL-positive nuclei were detected in cells cultivated in medium containing 0.05 mM Cd2+. Treatment with highly toxic 1 mM Cd2+ induced fast decrease of cell viability (no viable cells remained after 6-h treatment) and cell death occurred before DNA cleavage might be initiated. The exposure of tobacco BY-2 cells to 0.05 mM Cd2+ resulted in a marked accumulation of total PAs (represented by the sum of free PAs and their perchloric acid (PCA)-soluble and PCA-insoluble conjugates) during 3-day treatment. The increase in total PA contents was primarily caused by the increase in putrescine (Put) concentration. The accumulation of free spermidine (Spd) and spermine (Spm) at 12 and 24 h in 0.05 mM Cd2+ treated BY-2 cells and high contents of Spd and especially Spm determined in dead cells after I mM Cd2+ application was observed. The participation of PA conjugation with hydroxycinnamic acids and PA oxidative deamination in maintaining of free PA levels in BY-2 cells under Cd2+-induced oxidative stress is discussed.
Insights
Cadmium (Cd2+) exposure in tobacco cells (Nicotiana tabacum L.) significantly reduces cell viability and alters cell morphology. Polyamines (PAs) accumulate, particularly putrescine, suggesting a role in cellular response to cadmium-induced oxidative stress.
Area of Science:
- Plant Science
- Toxicology
- Cell Biology
Background:
- Cadmium (Cd2+) is a toxic heavy metal posing environmental and health risks.
- Understanding plant responses to heavy metal stress is crucial for agriculture and environmental remediation.
- Tobacco BY-2 cells (Nicotiana tabacum L.) are a model system for plant cell studies.
Purpose of the Study:
- To investigate the effects of cadmium sulfate (CdSO4) on tobacco BY-2 cell viability, morphology, and proliferation.
- To analyze changes in endogenous polyamine (PA) levels, including free and conjugated forms, under cadmium stress.
- To explore the role of polyamines in cellular defense mechanisms against cadmium-induced oxidative stress.
Main Methods:
- Tobacco BY-2 cells were treated with 0.05 mM and 1 mM CdSO4 for up to 7 days.
- Cell viability and proliferation were assessed using standard assays.
- Cell and nuclear morphology, including DNA fragmentation (TUNEL assay), were examined.
- Endogenous polyamine levels (free and conjugated forms) were quantified using chromatographic methods.
Main Results:
- 0.05 mM Cd2+ reduced cell viability to ~60% within 24 hours, causing nuclear fragmentation and blocked proliferation.
- 1 mM Cd2+ induced rapid cell death within 6 hours, preceding DNA fragmentation.
- Cadmium exposure led to a marked accumulation of total polyamines, primarily putrescine (Put), over 3 days.
- Accumulation of free spermidine (Spd) and spermine (Spm) was observed, with high levels in dead cells exposed to 1 mM Cd2+.
Conclusions:
- Cadmium exposure induces significant cellular damage and death in tobacco BY-2 cells, with varying responses based on concentration.
- Polyamines, especially putrescine, accumulate in response to cadmium stress, indicating their involvement in cellular defense.
- The study suggests polyamine conjugation and oxidative deamination are key mechanisms for maintaining polyamine homeostasis under cadmium-induced oxidative stress.
