Molecular Analysis Transcription Factors Regulating phyA Gene Expression

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9513590 Quail This proposal is focused on the previously cloned transcription factor, GT-2. The specific objectives are: (a) to perform further detailed analyses of the functional activities of GT-2, including transcriptional activation analysis using a transfection assay to define domains of GT-2 involved in transactivation, and a series of biochemical assays to assess capacity for, and map any domains involved in, multimerization; (b) to undertake a structural analysis of the previously defined twin minimal DNA-binding domains of GT-2 using high resolution nuclear magnetic resonance spectroscopy for the purpose of beginning to understand the molecular basis of sequence-specific DNA recognition by this novel class of transciption factor; and (c) to screen cDNA libraries for other protein factors that interact with GT-2 using in vitro interaction cloning and the yeast two hybrid system for the purpose of approaching the question of the mechanism by which GT-2 enhances transcriptional activity. %%% Plants respond to light by activating certain genes that govern growth and differentiation. The mechanism involves a light-activated receptor in the cell that sends a chemical signal to certain genes that are specifically light activated including the receptor itself. A protein that binds close to the DNA that contains the code for the receptor has been discovered. This protein is unique to plants and has DNA binding regions at both ends. This protein accelerates the production of the receptor by binding to the DNA and by binding to other proteins that help produce the message. The research will determine what parts of the protein are responsible for its ability to accelerate the production of the message. Also, the structure of the protein will be analyzed to determine how each segment of the molecule is arranged into a molecular blueprint. ***