Plant phototropism is an adaptive response to changes in light direction, quantity, and quality that results in optimization of photosynthetic light harvesting, as well as water and nutrient acquisition. Though several components of the phototropic signal response pathway have been identified in recent years, including the blue light (BL) receptors phototropin1 (phot1) and phot2, much remains unknown. Here, we show that the phot1-interacting protein NONPHOTOTROPIC HYPOCOTYL3 (NPH3) functions as a substrate adapter in a CULLIN3-based E3 ubiquitin ligase, CRL3 NPH3 . Under low-intensity BL, CRL3 NPH3 mediates the mono/multiubiquitination of phot1, likely marking it for clathrin-dependent internalization from the plasma membrane. In high-intensity BL, phot1 is both mono/multi-and polyubiquitinated by CRL3 NPH3 , with the latter event targeting phot1 for 26S proteasome-mediated degradation. Polyubiquitination and subsequent degradation of phot1 under high-intensity BL likely represent means of receptor desensitization, while mono/multiubiquitination-stimulated internalization of phot1 may be coupled to BL-induced relocalization of hormone (auxin) transporters.

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Summary Reactive oxygen species (ROS) and the photoreceptor protein phytochrome B (phyB) play a key role in plant acclimation to stress. However, how phyB that primarily functions in the nuclei impacts ROS signaling mediated by respiratory burst oxidase homolog (RBOH) proteins that reside on the plasma membrane, during stress, is unknown. Arabidopsis thaliana and Oryza sativa mutants, RNA‐Seq, bioinformatics, biochemistry, molecular biology, and whole‐plant ROS imaging were used to address this question. Here, we reveal that phyB and RBOHs function as part of a key regulatory module that controls apoplastic ROS production, stress‐response transcript expression, and plant acclimation in response to excess light stress. We further show that phyB can regulate ROS production during stress even if it is restricted to the cytosol and that phyB, respiratory burst oxidase protein D (RBOHD), and respiratory burst oxidase protein F (RBOHF) coregulate thousands of transcripts in response to light stress. Surprisingly, we found that phyB is also required for ROS accumulation in response to heat, wounding, cold, and bacterial infection. Our findings reveal that phyB plays a canonical role in plant responses to biotic and abiotic stresses, regulating apoplastic ROS production, possibly while at the cytosol, and that phyB and RBOHD/RBOHF function in the same regulatory pathway.

Plants are sessile organisms and therefore are limited in their ability to gather resources. Therefore, they have evolved several mechanisms that aid them in their quest to gather light, water and nutrients. One such mechanism is phototropism, a plants ability to bend towards or away from a light source. This mechanism is mediated by the blue light photoreceptor phototropin (phot). Arabidopsis thaliana contains two phototropins, phot1 which is the primary photoreceptor under low intensity light and phot2, which acts redundantly with phot1 under high intensity light. The perception of blue light by the phototropins (phot1 and phot2) initiates signaling events that lead to a lateral redistribution of the plant hormone auxin; which ultimately results in differential growth and the bending response. In addition to phototropism, these proteins mediate several other growth and developmental responses such as leaf movement, chloroplast movement, and stomatal opening. A second protein which is critical to the phototropic response and interacts with phot1 is NONPHOTOTROPIC HYPOCOTYL3 (NPH3). In addition to phototropism, NPH3 has a role in phot1-mediated leaf movement. NPH3 has been shown to act as a substrate adapter in an E3 ubiquitin ligase complex with the protein CULLIN3 (CUL3). This CRL3NPH3 complex is responsible for ubiquitinating the phot1 photoreceptor in a blue light fashion. This ubiquitination has been shown to be necessary for the bending response, but it's role in receptor ubiquitination is still not fully understood. To better understand this component of phot1-mediated phototropism, we characterized an allelic series of NPH3 mutants to further understand the role of this substrate adapter in this mechanism. Additionally, we characterized several mutant plant lines containing a mutant phot1 protein in which critical ubiquitination sites were mutated in an effort to render the protein unable to be ubiquitinated to further understand phot1 ubiquitination.