CsAKT1 is a key gene for CeO2 nanoparticles improved cucumber salt tolerance: a validation from CRISPR-Cas9 lines

Abstract

<title>Abstract</title> Background Salinity is one of the main factors limiting crop growth and yield. Applications of nanomaterials such as cerium oxide nanoparticles (nanoceria) improved salt tolerance in many plant species. The known mechanisms of nano-improved crop salt tolerance are mostly at the physiological and hormonal but not the gene level. Lack of proper methods to precisely investigate the key genes involved in nano-improved crop salt tolerance is one of the main reasons. Results We found that both leaf and root application of PNC (poly acrylic acid coated nanoceria) improved cucumber salt tolerance, showing higher shoot and root dry weight, and better photosynthetic performance than control plants. Under salinity stress, PNC treated cucumber plants showed significant lower malondialdehyde (MDA) content of leaf (46.7% and 13.3% for leaf and root application of PNC, respectively) and root (31.4% and 19.9% for leaf and root application of PNC, respectively), and reactive oxygen species (ROS) level such as H₂O₂ content of leaf (46.6% and 22.0% for leaf and root application of PNC, respectively) and root (50.0% and 29.9% for leaf and root application of PNC, respectively). Further experiments showed that under salinity stress, compared with control plants, PNC treated cucumber plants had significant higher leaf (216.1% and 59.5% for leaf and root application of PNC, respectively) and root (148.2% and 71.8% for leaf and root application of PNC, respectively) K⁺ content. RNA seq showed that under salinity stress, the modulation on <italic>AKT1</italic> is more responsible for the PNC improved K⁺ maintenance in cucumber. Further, <italic>CsAKT1</italic> depletion experiment (CRISPR-Cas9 lines) confirmed that under salinity stress, <italic>CsAKT1</italic> plays an important role in potassium uptake in cucumber plants treated with PNC. Conclusions Our results showed that foliar PNC delivery enabled stronger cucumber salt tolerance than the root application is associated with the better maintained K⁺/Na⁺ ratio. CRISPR-Cas9 lines further confirmed that <italic>CsAKT1</italic> is a key gene involved in PNC-improved cucumber salt tolerance. Our work not only adds more knowledge to the mechanisms underlying nano-improved plant salt tolerance, but also suggests a CRISPR-Cas9 approach to study the key genes involved in it.