Date of Award
Spring 2024
Document Type
Dissertation
Degree Name
Doctor of Philosophy (PhD)
Department
Molecular Biophysics and Biochemistry
First Advisor
Pyle, Anna
Abstract
Pre-mRNA processing is a critical metabolic step in eukaryotes. RNA transcripts must be spliced, polyadenylated, and capped, among other modifications, to ensure their proper function. Group II introns are an archetypal splicing system that can catalyze their self-excision and retroelements that invade DNA. They provide a window into understanding the evolution of splicing and have played an important role in shaping the eukaryotic genome as the ancestors of spliceosomal introns, telomerases, and non-LTR retrotransposons. Group II introns consist of a well-folded, large-structured RNA that forms a ribonucleoprotein complex with their encoded maturases to carry out splicing and intron integration functions. Despite their central role in RNA metabolism, there are key questions that remained unanswered regarding their structure, function, and mechanisms of action. In this work, I primarily use biochemical, biophysical, and structural techniques to understand the splicing pathway of group II introns. Using cryo-electron microscopy, we visualized group II intron RNPs as they proceed through the branching pathway, revealing the network of interactions that dictate branch helix positioning, branch site selection, and splice site exchange. There are striking similarities with the spliceosome that highlight the pattern of conservation from group II introns to the spliceosome. Continuing with the intron lifecycle, I then investigated their function as retroelements that target new DNA insertion sites, which occurs after maturase-mediated intron excision. I characterized the interactions of the intron holoenzyme with its structured DNA targets, resulting in important findings regarding the type of molecular strategies that enable shape and sequence specific recognition. Overall, this work provides a comprehensive view of the intron life cycle, encompassing the forward splicing (intron excision) and reverse splicing (intron retrotransposition) reactions. This work sheds light on pre-mRNA splicing machines and RNP retroelement complexes, providing a foundation for understanding how ancient group II intron elements have and continue to impact eukaryotic genomes.
Recommended Citation
Chung, Kevin, "Structural insights into group II intron splicing and retrotransposition" (2024). Yale Graduate School of Arts and Sciences Dissertations. 1388.
https://elischolar.library.yale.edu/gsas_dissertations/1388
