Despite its outsized role in plant defense, NPR1’s structure has remained elusive ­– much to the consternation of researchers in the field. Without detailed structure data, scientists have struggled to understand how the protein governs plant protection, the author says. “What’s really crucial and missing is an explanation of how NPR1 works on a molecular level.”

In new work that unveils how NPR1 looks and acts, the teams bridge that gap – a find that could change the face of plant breeding. The two groups report the structure of NPR1 from the common lab plant Arabidopsis thaliana, in the journal Nature.

For as long as humans have cultivated crops, they have had to fight off the numerous pests and pathogens that stymie plant growth. The water mold Phytophthora infestans, for instance, is one of the most notorious baddies ­– responsible for the Irish Potato Famine that resulted in a million deaths and two million refugees. “It’s a huge struggle that has shaped our world,” says the senior author.

Today, pathogens continue to plague bananas, avocados, and other popular crops. But tackling the problem with traditional approaches can be problematic. Chemical pesticides, for example, are often toxic to the environment. That’s one reason plant breeders are now looking to genetic solutions, like engineering plant cells to produce high levels of NPR1. The approach has proven successful in the lab and in limited field trials, but with one catch: as immunity increases, growth declines.

The researchers solved NPR1’s structure using x-ray crystallography and the imaging technique cryo-electron microscopy (cryo-EM). The team’s success stemmed from using the techniques complementarily. Cryo-EM gave the researchers a preliminary structure of NPR1, which offered crucial insight into how to prepare the protein for successful crystallography. The result: high-resolution images of NPR1 and its key functional regions.

While previous studies offered glimpses into parts of NPR1’s structure, none have been “as comprehensive as reported in this new paper,” the author says. The new images reveal that two NPR1 proteins come together, forming a structure that resembles a bird with unfurled wings. At the wing tips, NPR1 binds to molecules in the cell’s nucleus to turn on plant immune genes, Crystal structure analysis revealed a unique zinc-finger motif in BTB for interacting with ankyrin repeats and mediating NPR1 oligomerization.

The authors found that, after stimulation, salicylic-acid-induced folding and docking of the salicylic-acid-binding domain onto ankyrin repeats is required for the transcriptional cofactor activity of NPR1, providing a structural explanation for a direct role of salicylic acid in regulating NPR1-dependent gene expression.

Moreover, the structure of the TGA3₂–NPR1₂–TGA3₂ complex, DNA-binding assay and genetic data show that dimeric NPR1 activates transcription by bridging two fatty-acid-bound TGA3 dimers to form an enhanceosome. The stepwise assembly of the NPR1–TGA complex suggests possible hetero-oligomeric complex formation with other transcription factors, revealing how NPR1 reprograms the defence transcriptome.

Now, the team wants to find out how NPR1 folds into a new shape when an infection kicks the protein into action. “This study not only addressed many long-standing questions, but also points to new research directions,” the senior author says. “It’s an exciting time.”

https://www.nature.com/articles/s41586-022-04699-w