Indian Journal of Virology

Eukaryotes have evolved a defence system, termed RNA silencing, in which the production of short interfering (si) RNAs triggers the destruction of related complementary mRNAs. To counteract this system, both RNA and DNA viruses evolved suppressors of silencing, thereby allowing efficient viral replication and systemic infection. Suppressors have not been reported for members of the genus Mastrevirus in the family Geminiviridae which have single-stranded DNA genomes. Thus, we investigated whether the mastrevirus Bean yellow dwarf virus (BeYDV) induced silencing. BeYDV encodes two early proteins (Rep and RepA) required for virus replication and two late proteins (movement protein and capsid protein) required for virus movement. After determining that BeYDV could induce a silencing response in Nicotiana benthamiana, we used a coagroinfiltration assay in which a 35S-GFP construct is used to induce silencing to investigate whether any of the four BeYDV proteins could suppress siRNA accumulation. We found that expression of Rep and RepA diminished the level of GFP siRNAs but also caused necrosis of the infiltrated tissue. To determine the site of action of the proteins, we transiently expressed Rep::GFP or RepA::GFP in N. benthamiana leaves. Both proteins have a nucleocytoplasmic location, consistent with functions in viral DNA replication and RNA silencing. We have used RepA mutants to identify the regions determining subcellular localisation and those required for necrosis and suppression of silencing. Our studies emphasize the multifunctional nature of the mastrevirus early proteins and we will discuss the roles of Rep and RepA in the viral infection cycle and the likely mechanisms by which the functions are regulated.