Fahy, Deirdre and Sanad, Marwa, N. M. E. and Duscha, Kerstin and Lyons, Madison and Liu, Fuqua and Bozhkov, Peter and Kunz, Hans-Henning and Hu, Jinping and Neuhaus, H. Ekkehard and Steel, Patrick G. and Smertenko, Andrei (2017) 'Impact of salt stress, cell death, and autophagy on peroxisomes : quantitative and morphological analyses using small fluorescent probe N-BODIPY.', Scientific reports., 7 . p. 39069.
Plant peroxisomes maintain a plethora of key life processes including fatty acid β-oxidation, photorespiration, synthesis of hormones, and homeostasis of reactive oxygen species (ROS). Abundance of peroxisomes in cells is dynamic; however mechanisms controlling peroxisome proliferation remain poorly understood because measuring peroxisome abundance is technically challenging. Counting peroxisomes in individual cells of complex organs by electron or fluorescence microscopy is expensive and time consuming. Here we present a simple technique for quantifying peroxisome abundance using the small probe Nitro-BODIPY, which in vivo fluoresces selectively inside peroxisomes. The physiological relevance of our technique was demonstrated using salinity as a known inducer of peroxisome proliferation. While significant peroxisome proliferation was observed in wild-type Arabidopsis leaves following 5-hour exposure to NaCl, no proliferation was detected in the salt-susceptible mutants fry1-6, sos1-14, and sos1-15. We also found that N-BODIPY detects aggregation of peroxisomes during final stages of programmed cell death and can be used as a marker of this stage. Furthermore, accumulation of peroxisomes in an autophagy-deficient Arabidopsis mutant atg5 correlated with N-BODIPY labeling. In conclusion, the technique reported here enables quantification of peroxisomes in plant material at various physiological settings. Its potential applications encompass identification of genes controlling peroxisome homeostasis and capturing stress-tolerant genotypes.
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|Publisher Web site:||https://doi.org/10.1038/srep39069|
|Publisher statement:||This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/ © The Author(s) 2017|
|Date accepted:||17 November 2016|
|Date deposited:||15 March 2017|
|Date of first online publication:||01 February 2017|
|Date first made open access:||15 March 2017|
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