The Non-Phototrophic Hypocotyl3 (NPH3)-domain protein NRL5 is a trafficking-associated GTPase essential for drought resistance

The mechanisms of plant resistance to low water potential (ψw) during drought are unclear but may involve signaling and trafficking at the plasma membrane as well as metabolic reprogramming, including proline accumulation. Forward genetic screening using a Proline Dehydrogenase 1 ( ProDH1) promoter:reporter line identified a mutant with extreme low ψw hypersensitivity due to a single amino acid substitution (P335L) in the Non-Phototrophic Hypocotyl3 (NPH3) domain of NPH3/RPT2-Like5 (NRL5)/Naked Pins in Yucca8 (NPY8). Further experiments found that NRL5, and other NPH3-domain proteins, are GTPases. NRL5 interacted with RAB small GTPases and the SNARE proteins VAMP721/722 and had polar localization. NRL5P335L had greatly reduced GTPase activity, impaired RAB and VAMP721/722 interaction and disrupted polar localization. These data demonstrate that NRL5-mediated restraint of proline catabolism is required for drought resistance and also more broadly define unexpected functions of the NPH3 domain such that the role of NPH3-domain proteins in signaling, trafficking, and cellular polarity can be critically re-evaluated. One-Sentence Summary A protein containing the plant-specific NPH3-domain has GTPase activity, trafficking interaction and drought resistance function. ### Competing Interest Statement The authors have declared no competing interest.