OUP accepted manuscript

Abstract

Positive-sense single-stranded RNA viruses form the largest and most diverse group of eukaryote-infecting viruses. Their genomes comprise one or more segments of coding-sense RNA that function directly as messenger RNAs upon release into the cytoplasm of infected cells. Positive-sense RNA viruses are generally accepted to encode proteins solely on the positive strand. However, we previously identified a surprisingly long ($1,000-codon) open reading frame (ORF) on the negative strand of some members of the family Narnaviridae which, together with RNA bacteriophages of the family Leviviridae, form a sister group to all other positive-sense RNA viruses. Here, we completed the genomes of three mosquito-associated narnaviruses, all of which have the long reverse-frame ORF. We systematically identified narnaviral sequences in public data sets from a wide range of sources, including arthropod, fungal, and plant transcriptomic data sets. Long reverse-frame ORFs are widespread in one clade of narnaviruses, where they frequently occupy >95 per cent of the genome. The reverseframe ORFs correspond to a specific avoidance of CUA, UUA, and UCA codons (i.e. stop codon reverse complements) in the forward-frame RNA-dependent RNA polymerase ORF. However, absence of these codons cannot be explained by other factors such as inability to decode these codons or GC3 bias. Together with other analyses, we provide the strongest evidence yet of coding capacity on the negative strand of a positive-sense RNA virus. As these ORFs comprise some of the longest known overlapping genes, their study may be of broad relevance to understanding overlapping gene evolution and de novo origin of genes.

Virome Analysis of Aphid Populations That Infest the Barley Field: The Discovery of Two Novel Groups of Nege/Kita-Like Viruses and Other Novel RNA Viruses

“…that potentially encodes a reverse-ORF, as previous studies have proposed for some narna-like viruses, such as Hubei narna-like virus 21 and Zhejiang mosquito virus 3 (Cook et al, 2013;DeRisi et al, 2019;Dinan et al, 2020; Figure 6A). In the phylogenetic tree, BARV-6 and -7 grouped together with yeast narnaviruses (family Narnaviridae) and many invertebrate-associated narnalike viruses (Figure 6B).…”

Characterization of a Novel Mitovirus of the Sand Fly Lutzomyia longipalpis Using Genomic and Virus–Host Interaction Signatures

“…Conversely, the Narnavirus genus shows an average genome size of 2500-3000 nt and an RdRp ORF with a maximum length of up to 1200 aa residues. Some narnaviruses also present ambigrammatic sequences, characterized by an additional large ORF coded in the reverse strand of the genome [78,79]. To characterize and propose a probable host of Lul-MV-1, we analyzed some intrinsic features of the viral genome.…”

Shadow ORFs illuminated: long overlapping genes inPseudomonas aeruginosaare translated and under purifying selection

“…The latter contains an unusual exception constituting an antisense ORF of ~1000 AA in the family Narnaviridae 84 . This ORF was hypothesized to be protein-coding 85 , but experimental evidence is lacking.…”