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-1 Programmed ribosomal frameshifting in Class 2 umbravirus-like RNAs uses multiple long-distance interactions to shift between active and inactive structures and destabilize the frameshift stimulating element.

ABSTRACT: Plus-strand RNA viruses frequently employ -1 programmed ribosomal frameshifting (-1 PRF) to maximize their coding capacity. Ribosomes can frameshift at a slippery sequence if progression is impeded by a frameshift stimulating element (FSE), which is generally a stable, complex, dynamic structure with multiple conformations that contribute to the efficiency of -1 PRF. As FSE are usually analyzed separate from the viral genome, little is known about cis-acting long-distance interactions. Using full-length genomic RNA of umbravirus-like (ula)RNA citrus yellow vein associated virus (CY1) and translation in wheat germ extracts, six tertiary interactions were found associated with the CY1 FSE that span nearly three-quarters of the 2.7 kb genomic RNA. All six tertiary interactions are conserved in other Class 2 ulaRNAs and two are conserved in all ulaRNAs. Two sets of interactions comprise local and distal pseudoknots that involve overlapping FSE nucleotides and thus are structurally incompatible, suggesting that Class 2 FSEs assume multiple conformations. Importantly, two long-distance interactions connect with sequences on opposite sides of the critical FSE central stem, which would unzip the stem and destabilize the FSE. These latter interactions could allow a frameshifting ribosome to translate through a structurally disrupted upstream FSE that no longer blocks ribosome progression.

SUBMITTER: Mikkelsen AA

PROVIDER: S-EPMC10602861 | biostudies-literature | 2023 Oct

REPOSITORIES: biostudies-literature

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-1 Programmed ribosomal frameshifting in Class 2 umbravirus-like RNAs uses multiple long-distance interactions to shift between active and inactive structures and destabilize the frameshift stimulating element.

Mikkelsen Anna A AA Gao Feng F Carino Elizabeth E Bera Sayanta S Simon Anne E AE

Nucleic acids research 20231001 19

Plus-strand RNA viruses frequently employ -1 programmed ribosomal frameshifting (-1 PRF) to maximize their coding capacity. Ribosomes can frameshift at a slippery sequence if progression is impeded by a frameshift stimulating element (FSE), which is generally a stable, complex, dynamic structure with multiple conformations that contribute to the efficiency of -1 PRF. As FSE are usually analyzed separate from the viral genome, little is known about cis-acting long-distance interactions. Using ful ...[more]

PMID: 37742076

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Project description:The -2/-1 programmed ribosomal frameshifting (-2/-1 PRF) mechanism in porcine reproductive and respiratory syndrome virus (PRRSV) leads to the translation of two additional viral proteins, nonstructural protein 2TF (nsp2TF) and nsp2N. This -2/-1 PRF mechanism is transactivated by a viral protein, nsp1β, and cellular poly(rC) binding proteins (PCBPs). Critical elements for -2/-1 PRF, including a slippery sequence and a downstream C-rich motif, were also identified in 11 simarteriviruses. However, the slippery sequences (XXXUCUCU instead of XXXUUUUU) in seven simarteriviruses can only facilitate -2 PRF to generate nsp2TF. The nsp1β of simian hemorrhagic fever virus (SHFV) was identified as a key factor that transactivates both -2 and -1 PRF, and the universally conserved Tyr111 and Arg114 in nsp1β are essential for this activity. In vitro translation experiments demonstrated the involvement of PCBPs in simarterivirus -2/-1 PRF. Using SHFV reverse genetics, we confirmed critical roles of nsp1β, slippery sequence, and C-rich motif in -2/-1 PRF in SHFV-infected cells. Attenuated virus growth ability was observed in SHFV mutants with impaired expression of nsp2TF and nsp2N. Comparative genomic sequence analysis showed that key elements of -2/-1 PRF are highly conserved in all known arteriviruses except equine arteritis virus (EAV) and wobbly possum disease virus (WPDV). Furthermore, -2/-1 PRF with SHFV PRF signal RNA can be stimulated by heterotypic nsp1βs of all non-EAV arteriviruses tested. Taken together, these data suggest that -2/-1 PRF is an evolutionarily conserved mechanism employed in non-EAV/-WPDV arteriviruses for the expression of additional viral proteins that are important for viral replication.IMPORTANCE Simarteriviruses are a group of arteriviruses infecting nonhuman primates, and a number of new species have been established in recent years. Although these arteriviruses are widely distributed among African nonhuman primates of different species, and some of them cause lethal hemorrhagic fever disease, this group of viruses has been undercharacterized. Since wild nonhuman primates are historically important sources or reservoirs of human pathogens, there is concern that simarteriviruses may be preemergent zoonotic pathogens. Thus, molecular characterization of simarteriviruses is becoming a priority in arterivirology. In this study, we demonstrated that an evolutionarily conserved ribosomal frameshifting mechanism is used by simarteriviruses and other distantly related arteriviruses for the expression of additional viral proteins. This mechanism is unprecedented in eukaryotic systems. Given the crucial role of ribosome function in all living systems, the potential impact of the in-depth characterization of this novel mechanism reaches beyond the field of virology.

Dataset Information

-1 Programmed ribosomal frameshifting in Class 2 umbravirus-like RNAs uses multiple long-distance interactions to shift between active and inactive structures and destabilize the frameshift stimulating element.

Publications

-1 Programmed ribosomal frameshifting in Class 2 umbravirus-like RNAs uses multiple long-distance interactions to shift between active and inactive structures and destabilize the frameshift stimulating element.

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