RNA-Guided RNA Modifications: Biogenesis, Functions, and Applications

Post-transcriptional modifications are ubiquitous in both protein-coding and noncoding RNAs (ncRNAs), playing crucial functional roles in diverse biological processes across all kingdoms of life. These RNA modifications can be achieved through two distinct mechanisms: RNA-independent and RNA-guided (also known as RNA-dependent). In the RNA-independent mechanism, modifications are directly introduced onto RNA molecules by enzymes without the involvement of other RNA molecules, while the cellular RNA-guided RNA modification system exists in the form of RNA-protein complexes, wherein one guide RNA collaborates with a set of proteins, including the modifying enzyme. The primary function of guide RNAs lies in their ability to bind to complementary regions within the target RNAs, orchestrating the installation of specific modifications. Both mechanisms offer unique advantages and are critical to the diverse and dynamic landscape of RNA modifications. RNA-independent modifications provide rapid and direct modification of RNA molecules, while RNA-guided mechanisms offer precise and programmable means to introduce modifications at specific RNA sites. Recently, emerging evidence has shed light on RNA-guided RNA modifications as a captivating area of research, providing precise and programmable control over RNA sequences and functions.In this Account, we focus on RNA modifications synthesized in an RNA-guided manner, including 2'-O-methylated nucleotides (Nm), pseudouridine (Ψ), N⁴-acetylcytidine (ac⁴C), and inosine (I). This Account sheds light on the intricate processes of biogenesis and elucidates the regulatory roles of these modifications in RNA metabolism. These roles include pivotal functions such as RNA stability, translation, and splicing, where each modification contributes to the diverse and finely tuned regulatory landscape of RNA biology. In addition to elucidating the biogenesis and functions of these modifications, we also provide an overview of high-throughput methods and their underlying biochemical principles used for the transcriptome-wide investigation of these modifications and their fundamental interactions in RNA-guided systems. This includes exploring RNA-protein interactions and RNA-RNA interactions, which play crucial roles in the dynamic regulatory networks of RNA-guided modifications. The ever-advancing methodologies have greatly enhanced our understanding of the dynamic and widespread nature of RNA-guided RNA modifications and their regulatory functions. Furthermore, the applications of RNA-guided RNA modifications are discussed, illuminating their potential in diverse fields. From basic research to gene therapy, the programmable nature of RNA-guided modifications presents exciting opportunities for manipulating gene expression and developing innovative therapeutic strategies.