Description
- Abstract:
- Nearly half of the human genome is comprised of mobile genetic elements known as transposons. Most transposons in humans can no longer create new copies of themselves, with the exception of the long interspersed nuclear element-1 (LINE-1 or L1). As the only autonomous retrotransposon in humans, L1 uses an RNA intermediate and the proteins encoded by its own sequence to “copy” and “paste” itself back into DNA in a process known as retrotransposition. Throughout evolution, germline L1 retrotransposition and L1-driven retrotransposition of non-autonomous retrotransposons has resulted in these sequences accounting for 28% of the human genome. While L1 is usually repressed in somatic tissues to protect genomic integrity, L1 expression often occurs in cases of cellular dysfunction, such as during aging, and contributes to disease progression. Therefore, preventing L1 activity by targeting its proteins presents a unique avenue for developing therapeutics applicable to a variety of diseases. L1 encodes the RNA-binding protein ORF1 and the enzymatic protein ORF2, which contains endonuclease (EN) and reverse transcriptase (RT) activities required for retrotransposition. Previous evidence has demonstrated the efficacy of pharmacological RT inhibition in reducing the harmful impacts of L1, such as inflammation in senescent cells and aged tissues. However, development of RT-specific inhibitors without the off-target concerns of existing inhibitors is hampered by the absence of an RT domain structure and difficulties in biochemical characterization. On the other hand, the EN domain structure has been solved, making it amenable to computational and biochemical screening. Additionally, the EN initiates retrotransposition and L1-induced DNA damage is mediated by EN activity. Here we describe the discovery of the first reported small molecule EN inhibitors. We identified these inhibitors using a combination of in silico and in vitro screening methods. We then determined the cellular effects of these inhibitors on L1 retrotransposition, DNA damage, and inflammation. Our results also suggest that the EN contributes to the production of inflammatory L1 sequences in senescent cells. These EN inhibitors will be useful tools for evaluating the role of the EN and L1 activity overall in disease, and as potential initial candidates for the design of L1-targeted therapeutics.
- Notes:
- Thesis (Ph. D.)--Brown University, 2023
Citation
D'Ordine, Alexandra Marie,
"Identification and Characterization of Small Molecule Inhibitors of the Endonuclease Encoded by the LINE-1 Retrotransposon"
(2023).
Molecular Biology, Cell Biology, and Biochemistry Theses and Dissertations.
Brown Digital Repository. Brown University Library.
https://repository.library.brown.edu/studio/item/bdr:fc75ctre/
Relations
Collection:
-
Molecular Biology, Cell Biology, and Biochemistry Theses and Dissertations
Theses and Dissertations for the Molecular Biology, Cell Biology, and Biochemistry department....