Revealing RNA Dynamics and Processing Using Advanced Microscopy Techniques

RNA mediated gene regulation has emerged as more and more important roles in central dogma. It has expanded the complexity of gene regulation at multiple different aspects. A large various of biomolecules have been found to be involved in RNA mediated regulation and it is precisely controlled in a spatiotemporal manner. Researchers have engineered RNA molecules or RNA-protein complex and introduced a new layer of regulation in cells. In addition, more research has shed a light on a large potential of targeting RNA molecules or leveraging RNA molecules for therapeutic purpose. In this thesis, combining advanced fluorescent imaging platform and biochemical assays, we aim to study RNA dynamics, modification, localization and functions in different biological system. We first used single-molecule Fluorescence Resonance Energy Transfer (sm-FRET) to study the dynamics of T-box and tRNA interaction in vitro. In the second study, we have leveraged an established deoxyribozyme (DR) and developed an RT-qPCR based method to quantify m6A modification of a particular site of a target RNA and we further made a progress on applying the DR for imaging m6A in situ. We also have found out mRNA localization inside nuclear speckle and splicing outcome can be regulated by certain RNA motifs. These studies have revealed new mechanisms of RNA regulation in different perspective and provided a new method to study RNA modifications.