Structural Dynamics of Knot-Like Viral RNAs Studied at the Single-Molecule Level: From Mechanisms to Drug Targets

Many viruses exploit special structures encoded in their RNA to carry out functions important for viral replication or defeat of host immune defenses, making them potential targets for novel types of antiviral drugs. I will discuss work on two such structures: knot-like folds that resist degradation by host RNases (known as xrRNAs) to make RNA fragments that interfere with immune responses, and pseudoknots that stimulate programmed ribosomal frameshifting (PRF) to produce essential viral proteins. Studying the structural dynamics of Zika virus xrRNAs using optical tweezers, we find that it is not the unusual knot-like fold topology that prevents digestion, but rather the extreme mechanical rigidity of the xrRNA. Using mutations and anti-sense oligomers, we identify the key load-bearing contacts essential for RNase resistance, as targets for therapeutic disruption. In contrast, studying the structural dynamics of a series of viral frameshift-stimulatory pseudoknots, we find that their mechanical rigidity is not responsible for their efficiency at stimulating PRF. Instead, conformational heterogeneity of the stimulatory structure appears to play a key role. Focusing on frameshifting in coronaviruses, we characterize the structures of pseudoknots as drugs targets and identify many new potent inhibitors of PRF, some inhibiting PRF across a broad-spectrum of coronaviruses, with potential as antiviral drugs.