Direct visualization of protein interactions in plant cells

Area of Science:

• Plant molecular biology
• Biochemistry
• Plant pathology

Background:

• Systemic acquired resistance (SAR) is a plant defense mechanism.
• NPR1/NIM1 protein is essential for SAR induction.
• NPR1/NIM1 interacts with TGA/OBF transcription factors, but in-cell monitoring methods were lacking.

Purpose of the Study:

• To develop and validate an in vivo protein fragment complementation assay (PCA) for detecting protein-protein interactions in plant cells.
• To investigate the effect of salicylic acid (SA) and its analog (INA) on the interaction between NPR1/NIM1 and TGA2 in Arabidopsis.

Main Methods:

• Developed an in planta protein fragment complementation assay (PCA) using reconstituted murine dihydrofolate reductase (mDHFR) and a fluorescent probe.
• Applied PCA to study the interaction between Arabidopsis NPR1/NIM1 and TGA2.
• Utilized salicylic acid (SA) and 2,6-dichloroisonicotinic acid (INA) to induce and monitor the protein interaction.

Main Results:

• Successfully demonstrated an in vivo PCA for detecting protein-protein interactions in plant cells.
• Showed that SA and INA induce the interaction between NPR1/NIM1 and TGA2 in Arabidopsis.
• Observed distinct species-specific responses to SA and INA.
• Localized the induced NPR1/NIM1-TGA2 interaction predominantly within the plant cell nucleus.

Conclusions:

• The developed PCA is a valuable tool for studying protein-protein interactions in planta.
• This method allows for the monitoring of SAR regulator-induced interactions.
• PCA has potential applications in agricultural research for pathway mapping and screening small molecules that modulate protein interactions.