Material Type  

 학위논문

 

0017162542

Date and Time of Latest Transaction  

20250211152024

ISBN  

9798346867340

DDC  

540

Author  

Tong, Yuquan.

Title/Author  

Transcriptome-Wide Mapping of Small-Molecule RNA-Binding Sites and Conversion into Degraders.

Publish Info  

[S.l.] : The Scripps Research Institute., 2024

Publish Info  

Ann Arbor : ProQuest Dissertations & Theses, 2024

Material Info  

723 p.

General Note  

Source: Dissertations Abstracts International, Volume: 86-06, Section: B.

General Note  

Advisor: Disney, Matthew D.;Griffin, Patrick R.

학위논문주기  

Thesis (Ph.D.)--The Scripps Research Institute, 2024.

Abstracts/Etc  

요약Originally considered merely as an intermediate biomolecule bridging the genome and the proteins, RNA is now established to play various key functions beyond translating proteins. Ranging from catalyzing chemical reactions to regulating gene expression, and from activating cellular responses to performing intercellular communications, the roles of RNA are evident in nearly every aspect of cellular processes in both health and disease biology. Notably, the diverse functions of RNA depend on their 3D structures, with harbors vast yet largely unexplored opportunities to be targeted by small molecules. Over the past decades, small molecules targeting RNA as potential precision medicines has garnered increasing attention in both academia and industry, as growing efforts have been devoted into this field to discover bioactive small molecules that can module RNA biology in a precise and predictable manner. Starting with antibiotics like Streptomycin targeting ribosomal RNA of bacteria, the chemical space of RNA-targeted small molecules has been expanded to include more scaffolds such as benzimidazoles, diphenylfurans, and many others. Consequently, the classes of RNA targetable by small molecules have also been expanded, including viral RNA, human mRNA, and microRNA precursors, where small molecules elicit bioactivity via a broad spectrum of mechanisms, including disrupting frameshifting, inhibiting translation, and repressing the biogenesis of mature microRNA. While the field of targeting RNA with small molecules is rapidly growing with an exciting and promising outlook, clinical success of RNA-targeted small molecules remains elusive. To bridge the gap between benchtop discovery and clinical applications and to accelerate the development of RNA-targeted small molecules as precision medicines, my thesis aims to overcome two main challenges in this field: (1) augmenting the bioactivity of RNA binders and (2) understanding small-molecule RNA interactions in live cells. The first challenge stems from the observation that many RNA binders are inactive in cells; that is, the simple binding of these small molecules does not affect the native function of target RNA and therefore is biologically silent. To address this challenge, we converted RNA binders into heterobifunctional degraders by appending them with a small molecule that locally activates endogenous RNase L to cleave the bound RNA target in a precise and stoichiometric manner (Chapters II to III). We have demonstrated the application of this strategy, dubbed RiboTAC (ribonuclease targeting chimera), to a broad range of RNA targets and diseases, ranging from human mRNA to microRNA precursors, and from neurodegenerative diseases to cancers. Notably, my thesis has reported the first literature example of converting inactive RNA binders into bioactive RiboTAC degraders, highlighting the potential and generalizability of this approach to augment the function of RNA binders even when the binding itself is biologically inactive.In the second part of my thesis (Chapters IV to V), we aim to develop an unbiased, transcriptome-wide platform to map small-molecule RNA interactions in intact biological systems. While many high-throughput assays for screening RNA binders have been established in vitro, folding RNA in a tube may not fully capture its structures and dynamics from its native cellular environment. Therefore, direct probing of small-molecule RNA interactions in live cells provide valuable information that can be leveraged to identify novel RNA binders and RNA structures, to assess the selectivity of lead RNA-targeted compounds, and to investigate the ligandability of the human transcriptome. To achieve this goal, we integrated covalent chemistry with next-generation sequencing, the former enabling mapping the binding sites of small molecules to RNA targets while the latter allowing for transcriptome-wide profiling. We further applied this platform to study RiboTACs, comparing the targets bound to the ones cleaved by RiboTACs. Importantly, our results revealed that only a small fraction of bound RNA targets are cleaved by RiboTACs, driven by factors such as the local RNA structures near the small molecule binding sites and the thermodynamic stability of bound RNA structures. These results not only led to the discovery of isoform-specific RiboTACs targeting cancer-dependent mRNAs, but also laid the foundation for understanding the selectivity and potency of RiboTACs for the rational design and optimization for this class of heterobifunctional compounds.Collectively, my thesis has showcased the potential and generalizability of RiboTACs by enhancing the potency of bioactive RNA binders, and by converting inactive RNA binders into bioactive degraders. To study factors affecting the selectivity and potency of RiboTACs in live cells, my thesis presents the development of an unbiased, transcriptome-wide platform to map small-molecule RNA binding sites and to profile RiboTAC cleavages in a massively paralleled and target-agnostic manner. Broadly, my thesis has provided a streamlined platform for augmenting the function of RNA binders and for unveiling the patterns of ligandability of the human transcriptome.

Subject Added Entry-Topical Term  

Chemistry.

Subject Added Entry-Topical Term  

Cellular biology.

Subject Added Entry-Topical Term  

Biochemistry.

Index Term-Uncontrolled  

Transcriptome

Index Term-Uncontrolled  

RNA binding sites

Index Term-Uncontrolled  

Intercellular communication

Index Term-Uncontrolled  

microRNAs

Added Entry-Corporate Name  

The Scripps Research Institute Chemical Biology

Host Item Entry  

Dissertations Abstracts International. 86-06B.

Electronic Location and Access  

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Control Number  

joongbu:658353