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A γ-Glutamylcysteine Synthetase Gene from Pyrus calleryana Is Responsive to Ions and Osmotic Stresses

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Abstract

Pyrus calleryana Decne. is widely used as a pear rootstocks in Asia. Glutathione (GSH) can induce the phytochelatin synthase gene’s (PcPCS1) transcription in this plant when exposed to cadmium. This phenomenon indicated that GSH plays a vital role in protecting P. calleryana from cadmium stress. Here, we have isolated and characterized a gene encoding γ-glutamylcysteine synthetase (PcγECS), the key enzyme of GSH biosynthesis, from the leaves of P. calleryana, with the aim of expanding our knowledge of the role of GSH at the molecular level. P. calleryana plants treated with cadmium, salt, PEG6000, or mannitol responded by increasing transcription of the PcγECS gene. The response was specific for cadmium, which causes a toxicity that is thought to be mitigated through phtochelatins, and other treatments that cause damage that can be relieved by the synthesis of GSH. Feeding experiments suggested that cysteine contributed to activating the transcription of PcγECS and GSH synthesis. However, the gene’s expression was inhibited in the presence of GSH. Recombinant Escherichia coli carrying PcγECS had higher GSH synthesis levels and grew better than the control cells under cadmium, salt, or osmotic stresses. These results suggest that PcγECS may participate in responses against multiple environmental stimuli in P. calleryana.

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Abbreviations

A 600 :

The absorbance value at 600 nm

Amp:

Ampicillin

DTNB:

5,5-Dithiobis-2-nitrobenoic acid

GSH:

Glutathione

IPTG:

Isopropyl β-d-1-thiogalactopyrano-side

MEGA:

Molecular evolutionary genetics analysis

ORF:

Open reading frame

PCs:

Phytochelatins

qPCR:

Quantitative real-time PCR

RACE:

Rapid amplification of cDNA ends

γ-ECS:

γ-Glutamylcysteine synthetase (also known as glutamate-cysteine ligase)

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Acknowledgments

This work was supported by the Jiangsu Agriculture Science and Technology Innovation Fund of China [CX(12)5033] and the National Natural Sciences Foundation of China (Nos. 31101529, 31372051).

Conflict of Interest

All authors declare that they have no conflicts of interest for this paper.

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Authors and Affiliations

  1. Institute of Horticulture, Jiangsu Academy of Agricultural Sciences/Jiangsu Key Laboratory for Horticultural Crop Genetic Improvement, Nanjing, 210014, China

    Hui Li, Jin-long Han, Jing Lin, Qing-song Yang & You-hong Chang

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  1. Hui Li
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  2. Jin-long Han
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  3. Jing Lin
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  4. Qing-song Yang
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  5. You-hong Chang
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Correspondence to You-hong Chang.

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Li, H., Han, Jl., Lin, J. et al. A γ-Glutamylcysteine Synthetase Gene from Pyrus calleryana Is Responsive to Ions and Osmotic Stresses. Plant Mol Biol Rep 33, 1088–1097 (2015). https://doi.org/10.1007/s11105-014-0821-2

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