Deciphering the RNA secondary structure landscape of human genes

Abstract

Ribonucleic acids (RNAs) are key macromolecules that play essential roles in almost every aspect of biology. One of the most critical roles it plays is in post-transcriptional regulation of gene expression, which is primarily influenced by folding of RNA into secondary and tertiary structure. As this folding influences many of the functional roles in biology, it is crucial to accurately predict secondary structures and detect those with the highest likelihood of function. With technological advancements and the increased interest in RNA as a therapeutic agent and target, mapping RNA secondary structural landscapes and determining functional regions within human genes and transcripts is of utmost importance to drive forward treatment of “undruggable” diseases. The work presented in this dissertation provides a large step towards this goal through advancements in our lab’s functional structure prediction tool, ScanFold; generation of large RNA structure databases; and defining best practices for identification of functional and targetable RNA secondary structures. Here, ScanFold and other state of the art RNA modeling programs were used in combination with experimental RNA structure probing to characterize the RNA folding landscape of both human and viral genes and transcripts. The following chapters describe the many improvements and validations of ScanFold; used of different biochemical probing methods; implementation of covariation and genomic annotation data; the experimental validation of functional structures; and the use of ScanFold in the design of RNA-based therapeutics. By applying these techniques to all human genes and transcripts and performing in-depth studies on select “undruggable” gene targets we have generated a map of the human RNA structurome and experimentally validated many functional and targetable structures. This work provides new insights into RNA biology and offers a valuable foundation for the development of RNA-based therapeutics.