Bioinformatic and Physical Characterizations of Genome-Scale Ordered RNA Structure in Mammalian RNA Viruses

Abstract

). Genome-scale ordered RNA structure (GORS) was widely distributed in many animal and plant viruses, much greater in extent than RNA structures required for viral translation or replication, but in mammalian viruses was associated with host persistence. To substantiate the existence of large-scale RNA structure differences between viruses, a large set of alignments of mammalian RNA viruses and rRNA sequences as controls were examined by thermodynamic methods (to calculate minimum free energy differences) and by algorithmically independent RNAz and Pfold methods. These methods produced generally concordant results and identified substantial differences in the degrees of evolutionarily conserved, sequence order-dependent RNA secondary structure between virus genera and groups. A probe hybridization accessibility assay was used to investigate the physical nature of GORS. Transcripts of hepatitis C virus (HCV), hepatitis G virus/GB virus-C (HGV/GBV-C), and murine norovirus, which are predicted to be structured, were largely inaccessible to hybridization in solution, in contrast to the almost universal binding of probes to a range of unstructured virus transcripts irrespective of G؉C content. Using atomic force microscopy, HCV and HGV/GBV-C RNA was visualized as tightly compacted prolate spheroids, while under the same experimental conditions the predicted unstructured poliovirus and rubella virus RNA were pleomorphic and had extensively single-stranded RNA on deposition. Bioinformatic and physical characterization methods both identified fundamental differences in the configurations of viral genomic RNA that may modify their interactions with host cell defenses and their ability to persist.The genetic material of the three domains of life is doublestranded DNA. In contrast, many of the viruses that parasitize these organisms have RNA-based genomes. In addition to their capacity to directly encode proteins, RNA genomes have the ability to form secondary and higher-order RNA structures that contribute to their stability and can form sequence motifs in a structural context that may function during the virus life cycle. For example, several virus groups have evolved internal ribosome entry sites that function in ribosome recruitment and the initiation of translation. A range of different types of internal ribosome entry site elements are recognized, all characterized by extensive (200-to 500-nucleotide [nt]) internal RNA-RNA base pairing (5). Similarly, genome replication per se or the packaging of the genome into capsid precursors can involve the interaction of the virus polymerase and/or other viral proteins with defined RNA structural elements (9,16,18,28,36,46). Typically, RNA structures involved in these replication functions take the form of discrete stem loops of various sizes, frequently with evolutionarily highly conserved sequences in terminal loops that are involved in RNA-protein or RNA-RNA interactions.Of the many methods devised to predict RNA secondary structures, the algorithm used in MFOLD and RNAFold ...

Genomic features and evolutionary constraints in Saffold-like cardioviruses

“…Secondly, RNA secondary structure in coding regions could restrict variation in cardioviruses. Many RNA viruses from different families have been suggested, by means of mathematic modelling, to contain a genome-scale ordered RNA structure (GORS) (Simmonds et al, 2004), which was later confirmed by experimental evidence (Davis et al, 2008). It was suggested that such structures could protect the virus from RNases or shield it from recognition by intracellular defence systems.…”

Functionally conserved architecture of hepatitis C virus RNA genomes

“…1 and 2C). These regions may be evolving under a complex mix of pressures related to maintaining highly compacted physical configurations (18,19) while evading immune recognition. HCV genomes replicate in the presence of innate immune response sensors that detect single-stranded (RNase L) and double-stranded (PKR and others) viral RNAs.…”

“…The HCV genomic RNA is physically compact (18) and highly structured (19). These features likely facilitate persistent HCV infections in humans by protecting the genome from degradation by innate antiviral defenses (20,21).…”

Large-scale screening and characterization of enteroviruses and kobuviruses infecting pigs in Vietnam

“…Although frequencies of persistence in PKV and other kobuviruses are poorly documented, genomes of all members of this genus possess structured RNA genomes (Davis et al, 2008;Simmonds et al, 2004) -a generic property associated with persistence in several other picornavirus genera and amongst other families of positive-stranded RNA viruses. High rates of detection in older pigs and …”