Exogenous Nitric Oxide Delays Plant Regeneration from Protoplast and Protonema Development in Physcomitrella patens.

Daniela Cervantes-Pérez,Ramón Alberto Batista-García,Verónica Lira-Ruan,Rigoberto Medina-Andrés,Angélica Ortega-García

doi:10.3390/plants9101380

Daniela Cervantes-Pérez, Ramón Alberto Batista-García + Show 3 more

Open Access

https://doi.org/10.3390/plants9101380

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Abstract

Nitric oxide (NO) has been recognized as a major player in the regulation of plant physiology and development. NO regulates cell cycle progression and cell elongation in flowering plants and green algae, although the information about NO function in non-vascular plants is scarce. Here, we analyze the effect of exogenous NO on Physcomitrella patens protonema growth. We find that increasing concentrations of the NO donor sodium nitroprusside (SNP) inhibit protonema relative growth rate and cell length. To further comprehend the effect of NO on moss development, we analyze the effect of SNP 5 and 10 µM on protoplast regeneration and, furthermore, protonema formation compared with untreated plants (control). Isolated protoplasts were left to regenerate for 24 h before starting the SNP treatments that lasted five days. The results show that SNP restrains the protoplast regeneration process and the formation of new protonema cells. When SNP treatments started five days after protoplast isolation, a decrease in cell number per protonema filament was observed, indicating an inhibition of cell cycle progression. Our results show that in non-vascular plants, NO negatively regulates plant regeneration, cell cycle and cell elongation.

Highlights

Nitric oxide (NO) is a versatile molecule that influences plant physiology and development from germination to senescence and in response to biotic and abiotic stresses [1,2] In land plants, NO is enzymatically generated by nitric oxide synthase-like activity (reviewed by Astier et al [3] and nitrate reductase (NR) [4]
The protonema pieces were transferred to fresh Knop medium or to the same medium supplemented with different concentrations of NO donor, sodium nitroprusside (SNP), with or without the NO scavenger
Plants appeared to be smaller as SNP concentration increased and the simultaneous addition of the NO scavenger cPTIO reduced that difference (Figure 1a)

Summary

Introduction

Nitric oxide (NO) is a versatile molecule that influences plant physiology and development from germination to senescence and in response to biotic and abiotic stresses [1,2] In land plants, NO is enzymatically generated by nitric oxide synthase-like activity (reviewed by Astier et al [3] and nitrate reductase (NR) [4]. In tomato and Arabidopsis thaliana roots, exogenous NO derived from SNP restrained primary root growth by altering the meristematic activity and inhibiting cell elongation [13,14]. An increase in endogenous NO levels at the beginning of the lag phase indicates its role in promoting active cell growth in the unicellular algae Chlorella vulgaris and Chlamydomonas sp. Physcomitrella patens is a useful model to understand complex plant physiology and developmental processes such as hormone response, stem cell formation and maintenance, and plant–pathogen interactions [19,20,21,22]. We show that increasing concentrations of the NO donor SNP decreases the protonema relative growth rate and cell elongation in a dose-dependent manner. The application of SNP on regenerating protoplasts delays plant regeneration and further chloronema development, indicating that in P. patens, NO affects both the cell cycle and cell elongation

Plant Material and Growth Conditions

Exogenous NO Treatments

Protoplast Isolation and NO Treatment

Plant Growth and Development Estimation

Results and Discussion

DAIatand