Collaborative Research: Investigations of the Plant Photoreceptor Kinase Family of Phototropins

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Plant photoreceptors mediate a broad spectrum of responses throughout plant development, from seed germination through all phases of vegetative growth to flowering and senescence. These photoreceptors include phytochromes, cryptochromes, and phototropins. Three additional proteins, FKF1, LKP2 and ZTL, involved in regulation of plant rhythms, likely form a fourth family of plant photoreceptors. The Phototropins (phot1 and phot2) mediate several blue light responses in plants including: phototropism, chloroplast relocation, stomatal opening, leaf expansion and the rapid inhibition of growth of etiolated seedlings. They contain two light-sensing domains (LOV1 and LOV2), each non-covalently binding an FMN chromophore, and a kinase domain. Light stimulation of the LOV domains leads to activation the kinase, and autophosphorylation. Arabidopsis phototropin LOV domains and more complex constructs can be expressed in transgenic plants and heterologous systems. These constructs are photoactive, but their photochemistry can be perturbed or selectively inactivated with site-specific mutations. This project focuses on the biochemical and biophysical characterization of constructs including the chromophore-binding domains and the full-length protein, and their ability to complement function in plants. These studies will probe the mechanism of light activation including the initial photochemistry and subsequent intramolecular signal-propagation events. This research will involve the use of time-resolved optical and vibrational spectroscopy, standard and time-resolved fluorescence and circular dichroism spectroscopy, time-resolved polarized light spectroscopy, and paramagnetic and spectroscopic probes of molecular structural changes. Studies on the phototropins have uncovered a chromophore module, the LOV domain that functions as a light-driven molecular switch. These LOV domains are closely related to the superfamily of PAS domains found in proteins in a diverse range of organisms. These studies should provide a new dimension in the understanding of their functions.Integration of research and education is an essential component of this project. This integration involves training of post-doctoral fellows and graduate students and mentoring of undergraduate students, including those from underrepresented groups. Outreach to high school students and teachers will continue.