Development of in Vitro Genetic System to Study RNA Editing

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9520253 Landweber The objective of this research is to develop the use of in vitro selection as a model system to explore he possible link between RNA catalysis and kinetoplastid RNA editing. RNA editing is the bizarre and complex addition or deletion of uridine residues which occurs in the mitochondria of some of the earliest eukaryotes, the kinetoplastid protozoa. The approach will be to develop a successful in vitro selection system to analyze the mechanism and evolution of RNA editing. These experiments will address the antiquity of RNA editing and the role of RNA catalysts in evolution. One hypothesis is that RNA editing is an ancient process, and the interaction of several short loci to produce one final transcript may reflect some of the steps involved in the first assembly of genes. Specifically, these experiments will test the hypothesis that RNA editing may have been one of the key players in the RNA world by analyzing whether each of its steps can be catalyzed by RNA, a characteristic of some of the earliest hypothesized biological mechanisms. This research will determine whether the first step of RNA editing, a transesterification similar to other RNA catalyzed ligation reactions, can be catalyzed by an RNA molecule. Through a series of in vitro selection experiments, which simulate evolution in a test tube, the experiments will explore the world of possible primitive RNA editing enzymes, by selecting the desired catalytic molecules from a pool of random sequences. Armed with these preliminary experiments, we will then be prepared to analyze the later steps of RNA editing, with the final goal of evolving a complete RNA catalyzed RNA editing system in vitro. These experiments will test whether a molecule capable of editing RNA could have arisen under fairly simple conditions: a model for the evolution of complex molecular processes. Ultimately, this will allow one to design new RNA enzymes that can edit RNA of any specific sequence. %%% The objective od this research is to deve lop the use of in vitro selection to explore the possible link between RNA catalysis and kinetoplastid RNA editing. RNA editing is the bizarre and complex addition or deletion of uridine residues which occurs in the mitochondria of some of the earliest eukaryotes, the kinetoplastid protozoa. The approach will be to develop a successful in vitro selection system to analyze the mechanism and evolution of RNA editing. These experiments will test whether a molecule capable of editing RNA could have arisen under fairly simple conditions: a model for the evolution of complex molecular processes. Through a series of in vitro selection experiments, which simulate evolution in a test tube, the experiments will explore the world of possible primitive RNA editing enzymes, by seleating new catalytic molecules from a pool of random sequences. Ultimately, this will allow one to design new RNA enzymes that can edit RNA of any specific sequence. ***