Molecular Analysis of Replication and Packaging Mechanism of Satellite Tobacco Mosaic Virus
Satellite tobacco mosaic virus (STMV) as the name implies is the satellite virus of the well characterized helper Tobacco mosaic virus (TMV). This satellite virus is 18nm in diameter having a T1 symmetry encapsidating a positive sense genomic RNA of 1058nt in size whose 3' 150nt shares 65% homology with that of its Helper virus. Extensive Structural studies on STMV suggested that its coat protein interacts extensively with the stem-loops of the packaged genomic RNA and hence would be unstable to form virions without interacting with its genomic RNA internally. However invivo experiments to prove the same were not performed. Host range effects and the role of the positively charged amino acids at the N-terminal of its capsid protein on replication and packaging were not been studied yet on this Satellite virus (or in any other satellite virus). Chapter 1 of this dissertation describes T-DNA based agrotransformants of STMV to synthesize its genomic RNA in either (+) or (-) orientation when expressed ectopically in Nicotiana benthamiana plants. The biological activity of ectopically expressed agrotransformants of STMV (+) or (-) transcripts was confirmed when co-expressed with TMV. Furthermore, progeny virions of STMV recovered from agroinfiltrations were indistinguishable from those recovered from mechanically inoculated plants. However, STMV virions assembled in N. benthamiana, but not in tobacco, efficiently packaged a variant form of the genomic RNA. Chapter 2 describes Individual substitution of the positively charged amino acids at the N-terminal region of its Coat protein to a neutrally charged amino acid Alanine affecting dramatically the replication and packaging of these mutant viruses. Chapter 3 describes Agrobacterium mediated transient expression of the three genomic RNA of Tomato Aspermy virus in N.benthamiana and by utilizing this system we demonstrate that expressed genomic RNA were biologically active and also induced symptoms in N.benthamiana.