Semester

Summer

Date of Graduation

2022

Document Type

Dissertation

Degree Type

PhD

College

School of Medicine

Department

Biochemistry

Committee Chair

Aaron Robart

Committee Co-Chair

Roberta Leonardi

Committee Member

Roberta Leonardi

Committee Member

Peter Stoilov

Committee Member

John Hollander

Committee Member

Qiang Ma

Abstract

Group II introns are ancient ribozymes capable of self-excision and mobility into new genomic locations. As the ancestors of both the eukaryotic spliceosome and eukaryotic retroelements, these simple RNA machines serve as excellent models for understanding the mechanisms of both gene splicing and retroelement activity in eukaryotes. Although group II introns have been studied for decades, knowledge of the most unique and arguably evolutionarily relevant class of group II introns, IIC, is limited. Our knowledge is especially limited in terms of understanding how IIC introns mediate the transition between the two transesterification steps of forward splicing and reverse splicing into DNA. We used in vitro biochemical methods and the IIC Ta.it.I1 intron from Thermoanaerobacter italicus to: 1) determine the role of the intron encoded protein and domain II - domain VI tertiary interactions in mediating the transition between forward splicing steps, and 2) determine the role of exon binding sequence – intron binding sequence interactions, reverse transcriptase domain engagement, and DNA target heterduplexes in mediating the transition between reverse splicing steps. The results of this dissertation establish that the Ta.it.I1 intron is dependent on its intron encoded protein for producing lariat ribonucleoprotein complexes and that the transition between the first and second steps of forward and reverse splicing is complex and therefore mediated by several factors.

Embargo Reason

Publication Pending

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