9509081 Chang Specific RNA-protein interactions are crucial in a variety of biological processes. These interactions are accomplished by proteins which can recognize a specific RNA structure rather than a particular RNA sequence. At present, identification of the physiologically relevant RNA ligand for a given RNA-binding protein is laborious and difficult. As the number of proteins predicted to bind RNA continues to grow, there is an urgent need for a reliable method for identifying their RNA ligands. To this end, this project will develop a "3-hybrid" genetic system, building on the concept of the well-known yeast 2-hybrid system. A "3-hybrid" test system consisting of two fusion proteins (the first and the second hybrids) and a "bipartite" RNA molecule (the third hybrid) will be produced in a yeast reporter strain. If the specific "sandwich-like" protein-RNA-protein interaction can be achieved in vivo, then GAL4 transcriptional activity may be reconstituted, leading to the transcriptional activation of a reporter gene. The long term goal of this research is to develop this in vivo screening method for identifying the RNA ligand for any given RNA-binding protein, or vice versa. %%% Specific RNA-protein interactions are crucial in a variety of biological processes. These interactions are accomplished by proteins which can recognize a specific RNA structure rather than a particular RNA sequence. At present, identification of the physiologically relevant RNA ligand for a given RNA-binding protein is laborious and difficult. As the number of proteins predicted to bind RNA continues to grow, there is an urgent need for a reliable method for identifying their RNA ligands. To this end, a "3-hybrid" genetic system will be produced in yeast so that specific protein-RNA-protein interactions in vivo will result in the transcriptional activation of a reporter gene. The goal of this research is to develop this in vivo screening method for identifying the RNA ligand for any given RNA-binding protein, or vice versa. ***
