Phot2-regulated relocation of NPH3 mediates phototropic response to high-intensity blue light in Arabidopsis thaliana

Two redundant blue-light receptors, known as phototropins (phot1 and phot2), influence a variety of physiological responses, including phototropism, chloroplast positioning, and stomatal opening in Arabidopsis thaliana. Whereas phot1 functions in both low- and high-intensity blue light (HBL), phot2 functions primarily in HBL. Here, we aimed to elucidate phot2-specific functions by screening for HBL-insensitive mutants among mutagenized Arabidopsis phot1 mutants. One of the resulting phot2 signaling associated (p2sa) double mutants, phot1 p2sa2, exhibited phototropic defects that could be restored by constitutively expressing NON-PHOTOTROPIC HYPOCOTYL 3 (NPH3), indicating that P2SA2 was allelic to NPH3. It was observed that NPH3-GFP signal mainly localized to and clustered on the plasma membrane in darkness. This NPH3 clustering on the plasma membrane was not affected by mutations in genes encoding proteins that interact with NPH3, including PHOT1, PHOT2 and ROOT PHOTOTROPISM 2 (RPT2). However, the HBL irradiation-mediated release of NPH3 proteins into the cytoplasm was inhibited in phot1 mutants and enhanced in phot2 and rpt2-2 mutants. Furthermore, HBL-induced hypocotyl phototropism was enhanced in phot1 mutants and inhibited in the phot2 and rpt2-2 mutants. Our findings indicate that phot1 regulates the dissociation of NPH3 from the plasma membrane, whereas phot2 mediates the stabilization and relocation of NPH3 to the plasma membrane to acclimate to HBL.