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Viruses commonly use specifically folded RNA elements that interact with both host and viral proteins to perform functions important for diverse viral processes. Examples are found at the 3′ termini of certain positive-sense ssRNA virus genomes where they partially mimic tRNAs, including being aminoacylated by host cell enzymes. Valine-accepting tRNA-like structures (TLSVal) are an example that share some clear homology with canonical tRNAs but have several important structural differences. Although many examples of TLSVal have been identified, we lacked a full understanding of their structural diversity and phylogenetic distribution. To address this, we undertook an in-depth bioinformatic and biochemical investigation of these RNAs, guided by recent high-resolution structures of a TLSVal. We cataloged many new examples in plant-infecting viruses but also in unrelated insect-specific viruses. Using biochemical and structural approaches, we verified the secondary structure of representative TLSVal substrates and tested their ability to be valylated, confirming previous observations of structural heterogeneity within this class. In a few cases, large stem–loop structures are inserted within variable regions located in an area of the TLS distal to known host cell factor binding sites. In addition, we identified one virus whose TLS has switched its anticodon away from valine, causing a loss of valylation activity; the implications of this remain unclear. These results refine our understanding of the structural and functional mechanistic details of tRNA mimicry and how this may be used in viral infection.

Polycomb Repressive Complex 2 (PRC2) contributes to initiation of heterochromatin through catalysis of histone H3 lysine 27 (H3K27) methylation. Human PRC2 can work productively with specific long noncoding RNAs (lncRNAs) at certain target loci, yet the underlying mechanism is unclear. In apparent contradiction to lncRNA-mediated PRC2 activity, PRC2 binds to RNA in a low-specificity, high-affinity manner, effectively competing with chromatin and making RNA a potent catalytic inhibitor of H3K27 methylation activity. We address this contradiction by testing the model that intermolecular RNA-RNA interactions between the lncRNA HOTAIR and its target genes can generate a context that is favorable for PRC2 activity. This RNA matchmaking activity is promoted by the heterogenous nuclear ribonucleoprotein (hnRNP) B1, which uses multiple protein domains to bind to specific regions of HOTAIR via multi-valent protein-RNA interactions. Using chemical probing, we find that RNA-RNA interaction and B1 binding facilitate RNA structural changes in HOTAIR. Finally, we demonstrate that RNA-RNA matches of HOTAIR with target gene RNAs can relieve the inhibitory effect of a single lncRNA for PRC2 activity. By dissecting molecular details of RNA matchmaking within the HOTAIR mechanism, we highlight a switch that can occur as PRC2 samples RNA interactions broadly in the nucleus, being activated on chromatin by specific RNA matches. This concept may generalize to many contexts where RNA plays a role in potentiating PRC2 activity.