In vitro- and in vivo-generated defective RNAs of satellite panicum mosaic virus define cis-acting RNA elements required for replication and movement.
Satellite panicum mosaic virus (SPMV) depends on its helper virus, panicum mosaic virus (PMV), to provide trans-acting proteins for replication and movement. The 824-nucleotide (nt) genome of SPMV possesses an open reading frame encoding a 17.5-kDa capsid protein (CP), which is shown to be dispensable for SPMV replication. To localize cis-acting RNA elements required for replication and movement, a comprehensive set of SPMV cDNA deletion mutants was generated. The results showed that the 263-nt 3' untranslated region (UTR) plus 73 nt upstream of the CP stop codon and the first 16 nt in the 5' UTR are required for SPMV RNA amplification and/or systemic spread. A region from nt 17 to 67 within the 5' UTR may have an accessory role in RNA accumulation, and a fragment bracketing nt 68 to 104 appears to be involved in the systemic movement of SPMV RNA in a host-dependent manner. Unexpectedly, defective RNAs (D-RNAs) accumulated de novo in millet plants coinfected with PMV and either of two SPMV mutants: SPMV-91, which is incapable of expressing the 17.5-kDa CP, and SPMV-GUG, which expresses low levels of the 17.5-kDa CP. The D-RNA derived from SPMV-91 was isolated from infected plants and used as a template to generate a cDNA clone. RNA transcripts derived from this 399-nt cDNA replicated and moved in millet plants coinoculated with PMV. The characterization of this D-RNA provided a biological confirmation that the critical RNA domains identified by the reverse genetic strategy are essential for SPMV replication and movement. The results additionally suggest that a potential "trigger" for spontaneous D-RNA accumulation may be associated with the absence or reduced accumulation of the 17.5-kDa SPMV CP. This represents the first report of a D-RNA associated with a satellite virus.
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Department of Plant Pathology and Microbiology, Texas A&M University, College Station, Texas 77843, USA.
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D020413:3' Untranslated Regions / Q000502:physiology; D020121:5' Untranslated Regions / Q000502:physiology; D001483:Base Sequence; D002213:Capsid / Q000032:analysis / Q000172:deficiency / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D017353:Gene Deletion; D016679:Genome, Viral; D008969:Molecular Sequence Data; D009029:Mosaic Viruses / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D009693:Nucleic Acid Hybridization; D016366:Open Reading Frames; D008897:Panicum / Q000821:virology; D012333:RNA, Messenger / Q000235:genetics; D019244:RNA, Satellite / Q000737:chemistry / Q000235:genetics*; D012526:Satellite Viruses / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment; D014779:Virus Replication
D020413:3' Untranslated Regions; D020121:5' Untranslated Regions; D018076:DNA, Complementary; D012333:RNA, Messenger; D019244:RNA, Satellite