title: Deciphering the mechanism of defective interfering RNA (DI RNA) biogenesis reveals that a viral protein and the DI RNA act antagonistically in virus infection.
abstract: Potato mop-top virus (PMTV) produces a defective RNA (D RNA) encompassing the 5'-terminal 479 nucleotides (nt) and 3'-terminal 372 nt of RNA-TGB (where TGB is triple gene block). The mechanism that controls D RNA biogenesis and the role of D RNA in virus accumulation was investigated by introducing deletions, insertions, and point mutations into the sequences of the open reading frames (ORFs) of TGB1 and the 8-kilodalton (8K) protein that were identified as required for efficient production of the D RNA. Transient expression of RNA-TGB in the absence of RNA-Rep (which encodes the replicase) did not result in accumulation of D RNA, indicating that its production is dependent on PMTV replication. The D RNA could be eliminated by disrupting a predicted minus-strand stem-loop structure comprising complementary sequences of the 5' TGB1 ORF and the 3' 8K ORF, suggesting intramolecular template switching during positive-strand synthesis as a mechanism for the D RNA biogenesis. Virus accumulation was reduced when the 8K ORF was disrupted but D RNA was produced. Conversely, the virus accumulated at higher titers when the 8K ORF was intact and D RNA production was blocked. These data demonstrate that the D RNA interferes with virus infection and therefore should be referred to as a defective interfering RNA (DI RNA). The 8K protein was shown to be a weak silencing suppressor. This study provides an example of the interplay between a pathogen and its molecular parasite where virus accumulation was differentially regulated by the 8K protein and DI RNA, indicating that they play antagonistic roles and suggesting a mechanism by which the virus can attenuate replication, decreasing viral load and thereby enhancing its efficiency as a parasite.
authors.affiliation: Department of Plant Biology and Forest Genetics, Uppsala BioCenter, Swedish University of Agricultural Sciences and Linnean Center for Plant Biology, Uppsala, Sweden.
authors.forename: Nina I
authors.initials: NI
authors.lastname: Lukhovitskaya
authors.forename: Srinivas
authors.initials: S
authors.lastname: Thaduri
authors.forename: Sonya K
authors.initials: SK
authors.lastname: Garushyants
authors.forename: Lesley
authors.initials: L
authors.lastname: Torrance
authors.forename: Eugene I
authors.initials: EI
authors.lastname: Savenkov
mesh_terms: D001483:Base Sequence; D003673:Defective Viruses / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D055029:Inverted Repeat Sequences; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010935:Plant Diseases / Q000821:virology*; D034622:RNA Interference; D012328:RNA Viruses / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D012367:RNA, Viral / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D014026:Nicotiana / Q000821:virology*; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D012367:RNA, Viral; D014764:Viral Proteins