Host-dependent recombination of a Tomato bushy stunt virus coat protein mutant yields truncated capsid subunits that form virus-like complexes which benefit systemic spread

This study examined the contribution of the Tomato bushy stunt virus (TBSV) coat protein (CP) and its corresponding RNA to systemic infection of plants. Compared to results obtained with a mutant lacking the 5'-half of the CP gene, the presence of those CP-RNA sequences in another mutant benefited TBSV infection on Nicotiana benthamiana even though wild-type CP expression was eliminated by introduction of a small out-of-frame deletion. RT-PCR of viral RNA associated with rapid infections established by this CP frameshift deletion mutant revealed that in planta recombination had provided the progeny with the ability to express a truncated CP (tCP) with a block of N-proximal 30 residues deleted from the 66 amino acid RNA-binding domain. Subsequent biochemical characterizations revealed the presence of large ribonucleoprotein complexes that were shown to contain viral RNA as well as the approximately 38-kDa tCP. Electron microscopic examination of purified complexes showed particle-like structures that were nonuniform in size and shape compared to wild-type TBSV particles. Inoculation of pepper with the tCP-containing ribonucleoprotein complexes resulted in a rapid systemic infection similar to that caused by wild-type TBSV. In contrast, infections established in pepper by the original CP frameshift deletion mutant transcripts were restricted to inoculated leaves and did not yield recombinants capable of systemically infecting this host. In summary, TBSV possesses the flexibility to form alternative virion-like structures even if a substantial portion of the RNA-binding domain is deleted from the CP; mutants producing the tCP-containing particle-like structures are more effective for virus spread than those devoid of CP expression; and recombination events to produce the alternative tCP-RNA complexes are host-dependent.