Graduation Semester and Year
Fall 2024
Language
English
Document Type
Dissertation
Degree Name
Doctor of Philosophy in Quantitative Biology
Department
Biology
First Advisor
Shawn Christensen
Second Advisor
Paul Chippindale
Third Advisor
Matthew Fujita
Fourth Advisor
Piya Ghose
Fifth Advisor
Subhrangsu Mandal
Abstract
Long Interspersed Elements (LINEs), also referred to as non-LTR retrotransposons, encode a multifunctional protein that facilitates reverse transcription of their mRNA into DNA at insertion sites through target-primed reverse transcription (TPRT). Specifically, R2 LINEs integrate into 28S rRNA genes through a sequence of DNA binding, cleavage, and synthesis reactions. While the initial stages of integration via TPRT are well-characterized, the later steps—second-strand DNA cleavage and synthesis—remain less understood.
This dissertation provides insights into the mechanistic and structural determinants of R2 retrotransposition, emphasizing the roles of Domain 6A, the N-terminal Myb and zinc finger (ZF) domains, and the structural requirements of junction DNA. Domain 6A, located within the reverse transcriptase domain, was found to be critical for DNA binding and cleavage. Mutations of key lysine residues in Domain 6A impaired these activities and disrupted interactions with Myb and ZF domains, essential for stabilizing the reverse transcriptase active site.
The N-terminal Myb and Zinc Finger (ZF) domains act as molecular scaffolds, facilitating sequence-specific DNA recognition and stabilizing protein-DNA interactions. Deletion of these domains in a truncated R2 protein, termed BoMoC, reduced DNA binding and RNA co-factor modulation, highlighting their essential roles in retrotransposition. Additionally, the junction DNA, a substrate for second-strand cleavage, was shown to exhibit sequence-specific and structural requirements.
These findings in this dissertation highlights the modular nature of the R2 protein, where distinct domains collaborate to achieve efficient DNA binding, cleavage, and integration. The study provides understanding of the structural and mechanistic determinants of R2 retrotransposition and establish a foundation for further exploration of retroelement biology.
Disciplines
Biochemistry | Genetics | Molecular Biology | Molecular Genetics
License
This work is licensed under a Creative Commons Attribution 4.0 International License.
Recommended Citation
Chatterjee, Joydeep, "STRUCTURAL AND MECHANISTIC INSIGHTS INTO DOMAIN-SPECIFIC FUNCTIONS OF THE R2 PROTEIN IN RETROTRANSPOSITION" (2024). Biology Dissertations. 223.
https://mavmatrix.uta.edu/biology_dissertations/223
Included in
Biochemistry Commons, Genetics Commons, Molecular Biology Commons, Molecular Genetics Commons
