Determining the Role of Pirin1 in Light-Regulated Growth and Cellular Responses in Arabidopsis thaliana

As photoautotrophic and sessile organisms, plants are both reliant upon, and vulnerable to abiotic signals. Different wavelengths of light provide information for plant development, but these signals can also become stressors at specific levels. PRN1, a G protein α effector and a member of the cupin superfamily, is reported to be involved in the transcription of a light-regulated gene, as well as in seed germination in Arabidopsis thaliana. My work herein reveals that PRN1 can act as a quercetinase in vitro and that this activity is regulated by the activation status of the G protein α subunit. My research also demonstrates that prn1 null mutants contain more of the UV-screening quercetin compound, and thus can withstand longer amounts of UV-C radiation than wild type seedlings. This thesis pioneers the use of PRN1 in plant transgenes, where it is demonstrated that both the overexpression of PRN1, and the lack of PRN1 cause disoriented growth in response to white light. Moreover, PRN::PRN1-GFP plants indicate that PRN1 is a mainly nuclear localized protein, with diffuse expression in other cellular areas. While the PRN1 transcript and protein are not strictly diurnally or circadian regulated, Western blot analysis indicates that the protein levels in the cell vary in response to light conditions. When in vitro translated PRN1 protein has been treated briefly with a total cell extract (with only large cell debris removed), there is a shift in band size to higher molecular mass, indicating post-translational modification of PRN1. Treatment of wild type plants with MG-132, a cell permeable proteasome inhibitor, indicates that the larger molecular weight species (corresponding to the anti-PRN1 specific antibody) are likely ubiquitinated. These data reveal many activities of and indicate the regulation of PRN1, and demonstrate the important roles that this protein may play in light-regulated growth and cellular responses to light in Arabidopsis thaliana.