Context-dependent structure formation of RNA hairpin motifs

Abstract

Many functions of ribonucleic acid (RNA) rely on its ability to assume specific sequence-structure motifs. Packaging signals found in certain RNA viruses are one such prominent example of functional RNA motifs. These signals are short hairpin loops that interact with coat proteins and drive viral self-assembly. As they are found in different positions along the much longer genomic RNA, the formation of their correct structure occurs as a part of a larger context. Any changes to this context can consequently lead to changes in the structure of the motifs themselves. In fact, previous studies have shown that structure and function of RNA motifs can be highly context-sensitive to the flanking sequence surrounding them. However, in what ways different flanking sequences influence the structure of an RNA motif they surround has yet to be studied in detail. We focus on a hairpin-rich region of the RNA genome of bacteriophage MS2--a well-studied RNA virus with a wide potential for use in biotechnology--and systematically examine context-dependent structural stability of 14 previously identified hairpin motifs, which include putative and confirmed packaging signals. Combining secondary and tertiary RNA structure prediction of the hairpin motifs placed in different contexts, ranging from the native genomic sequence to random RNA sequences and unstructured poly-U sequences, we determine different measures of motif structural stability. In this way, we show that while some motif structures can be stable in any context, others require specific context provided by the genome. Our results demonstrate the importance of context in RNA structure formation and how changes in the flanking sequence of an RNA motif sometimes lead to drastic changes in its structure. Structural stability of a motif in different contexts could provide additional insights into its functionality as well as assist in determining whether it remains functional when intentionally placed in other contexts. STATEMENT OF SIGNIFICANCERNA motifs are groups of related RNAs that possess similar sequence and/or structure and consequently assume similar functions. Despite their similarities, these motifs are often only a small part of larger RNA molecules, situated in various contexts provided by the surrounding (flanking) sequences. How the nature of the flanking sequence influences the structure of a motif it surrounds is a fundamental yet underexplored question. We systematically study context dependence of several hairpin motifs in the genomic RNA of bacteriophage MS2 which act as packaging signals, indispensable for virus assembly. We show that while some motifs fold into the correct structure no matter the nature of their context, others require the specific context provided by the genomic RNA.