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Quantification of NLR-Induced Cell Death Using Rice Protoplasts
Ayaka Yoshihisa1, Tsutomu Kawasaki2
1Graduate School of Agriculture, Kindai University, Nara, Japan.
Methods in Molecular Biology (Clifton, N.J.)
|February 5, 2026
Summary
Researchers developed a new rice leaf protoplast assay to measure plant cell death. This method enables evaluating nucleotide-binding leucine-rich repeat receptor (NLR) activity in rice, overcoming limitations of traditional techniques.
Area of Science:
- Plant immunology
- Molecular biology
- Plant pathology
Background:
- Plant nucleotide-binding leucine-rich repeat receptors (NLRs) are crucial for detecting pathogen effectors and triggering plant immune responses, often leading to programmed cell death.
- Assessing NLR activity typically involves measuring cell death levels, commonly performed in Nicotiana benthamiana using agroinfiltration for co-expression of NLRs and effectors.
- Agroinfiltration is not effective for protein expression in rice, limiting functional studies of rice NLRs and their interactions with pathogen effectors.
Purpose of the Study:
- To establish a novel and efficient method for assessing plant cell death in rice.
- To enable the functional evaluation of rice nucleotide-binding leucine-rich repeat receptor (NLR) activity.
- To provide an alternative assay for studying cell death-inducing proteins in rice.
Main Methods:
- Development of a protocol for preparing protoplasts from rice leaves.
- Implementation of a cell death assay using polyethylene glycol (PEG)-mediated transfection of rice protoplasts.
- Co-expression of plant nucleotide-binding leucine-rich repeat receptors (NLRs) and their cognate effectors within the protoplasts.
Main Results:
- Successful preparation of viable rice leaf protoplasts.
- Demonstration of polyethylene glycol (PEG)-mediated transfection efficiency in rice protoplasts.
- Establishment of a functional cell death assay in rice protoplasts for evaluating nucleotide-binding leucine-rich repeat receptor (NLR) activity.
Conclusions:
- A novel rice leaf protoplast-based cell death assay has been developed, overcoming the limitations of agroinfiltration in rice.
- This method provides a robust platform for studying the activity of plant nucleotide-binding leucine-rich repeat receptors (NLRs) and other cell death-inducing proteins in rice.
- The developed assay facilitates research into plant immunity and host-pathogen interactions in a key cereal crop.
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