G Protein Activation through Uncoupling Regulator of G Signaling Protein, AtRGS1

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Intellectual Merit: For organisms to develop normally and to react appropriately to their environment, the cells that compose organisms must communicate with each other. Signals used by cells to communicate can be hormones, light, small molecules, electrical impulses, and touch. For example, light can turn on (activate) a light receptor which then activates a few other molecular complexes that then activates many more switches, and so on to afford changes in the cell. Between the signal at the receptor and the changes occurring in the cell is a molecular switch called the heterotrimeric G protein complex. Heterotrimeric G protein-coupled signaling is used for normal development of the organism as well as reactions to pathogens causing disease. Recent investigations of heterotrimeric G protein-coupled signaling using divergent model systems reveals a potential paradigm shift on how signal activation can be regulated. For example, in metazoans and fungi, activation is the rate limiting step and is catalyzed by a ligand-stimulated, cell surface G protein coupled receptor (GPCR). In contrast, plants regulate the back reaction which is GTP hydrolysis leading to the resting state, and in Arabidopsis, this resting state is maintained by a transmembrane regulator of G Signaling (RGS) protein designated AtRGS1. In Arabidopsis, D-glucose and/or sugar metabolites are signals that are mediated in part by the heterotrimeric G protein complex. It is hypothesized that glucose and/or sugar metabolites activate the G protein pathway by regulating the coupling between AtRGS1 and AtGPA1, and it has been demonstrated that D-glucose induces a rapid internalization of AtRGS1, but not AtGPA1. Key questions to be answered include how uncoupling is achieved, the consequence of uncoupling, and the molecular mechanism for uncoupling. These questions will be examined using mathematical modeling, genetic manipulation of the signaling components, and microscopy. Broader Impacts: The broader impact is education of high school biology students on how differential cellular outcomes can be achieved through signal transduction. In collaboration with the UNC outreach program, a teaching module called "Same Genes- Different Fates" will be further developed to teach the concept of differential gene expression and the basis for different cell fates for a set of cells all having the same genes. While the module is currently designed for Biology I and II students (9th and 10th graders), it is adaptable to AP biology and college freshmen. This project will make those adaptations to the higher subject matter by adding concepts like epigenetics and signaling-induced changes into the instructions. The wet-lab part of the module utilizes transgenic plants that have gene promoters driving an enzyme activity that causes different parts of the plant to turn blue. The area of blue is dependent on the type of gene promoter and illustrates differential gene expression. Associated teacher workshops will be held to teach and promote the use of this module. The module will be distributed to high schools initially in North Carolina but eventually nationwide.