Group II introns are ribozymes with a diversity of biological functions: They catalyze their own splicing from precursor RNA and transposition into new genomic locations. At least one-third of the human genome may have originally derived from ancestral group II introns, which are the likely evolutionary predecessors of eukaryotic introns, the spliceosome and major classes of retrotransposons. Group II introns continue to play a major role in the metabolism and evolution of modern organisms and, given their ability to alter specific DNA and RNA sequences through transposition reactions, they are promising tools for biotechnology and molecular medicine. At a more basic level, group II introns have an unusual architecture that is providing a wealth of information on RNA folding, tertiary structure, and RNA-protein interactions. In order to apply group II introns in biomedical and basic research, and to understand molecular mechanisms for eukaryotic RNA splicing, it is essential that we visualize their three-dimensional structures. To this end, we propose a series of experiments to elucidate the high-resolution structures of group II introns during the multiple stages of intron folding and splicing. Specifically, we are employing crystallographic and biochemical methods to establish the folded architectural states of the intron and to visualize molecular details of the active-site. These studies are being conducted on isolated intron RNAs and on group II introns in complex with their natural partner proteins. This project is designed to yield fundamental new insights into the mechanisms by which RNA molecules fold into tertiary structures, catalyze chemical reactions and collaborate with proteins in order to carry out the basic metabolism of the cell. High-resolution structural information on group II introns will help us to understand, and potentially manipulate, molecular mechanisms for the RNA splicing reactions that control our patterns of gene expression. It will also facilitate the development of group II introns as tools for application in biotechnology and gene therapy.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Molecular Genetics A Study Section (MGA)
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Preusch, Peter C
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Yale University
Schools of Medicine
New Haven
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Zhao, Chen; Liu, Fei; Pyle, Anna Marie (2018) An ultraprocessive, accurate reverse transcriptase encoded by a metazoan group II intron. RNA 24:183-195
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Liu, Fei; Somarowthu, Srinivas; Pyle, Anna Marie (2017) Visualizing the secondary and tertiary architectural domains of lncRNA RepA. Nat Chem Biol 13:282-289
Dickey, Thayne H; Pyle, Anna M (2017) The SMAD3 transcription factor binds complex RNA structures with high affinity. Nucleic Acids Res 45:11980-11988
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Zhao, Chen; Pyle, Anna Marie (2017) The group II intron maturase: a reverse transcriptase and splicing factor go hand in hand. Curr Opin Struct Biol 47:30-39
Zhao, Chen; Pyle, Anna Marie (2016) Crystal structures of a group II intron maturase reveal a missing link in spliceosome evolution. Nat Struct Mol Biol 23:558-65
Somarowthu, Srinivas (2016) Progress and Current Challenges in Modeling Large RNAs. J Mol Biol 428:736-747
Pyle, Anna Marie (2016) Group II Intron Self-Splicing. Annu Rev Biophys 45:183-205
Marcia, Marco (2016) Using Molecular Replacement Phasing to Study the Structure and Function of RNA. Methods Mol Biol 1320:233-57

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