Genome analysis of the 3'-terminal part of the little cherry disease associated dsRNA reveals a monopartite clostero-like virus

The high molecular weight dsRNA associated with little cherry disease (LCD) was extracted from infected plant tissue and cloned as cDNA. The sequence of the 3' 8337 nucleotides was determined. Computer assisted translation of the sequence portion identified six open reading frames potentially encoding proteins (from 5' to 3') with molecular masses of 70 kDa, 61 kDa, 46 kDa, 76 kDa, 21 kDa and 27 kDa respectively. A 3'-terminal non-translated region of 210 nucleotides was present. The 70 kDa protein represents a homolog of the cellular HSP70 heat shock proteins, and the 61 kDa protein showed homology to the similarly encoded products of beet yellows (BYV), citrus tristeza (CTV) and lettuce infectious yellows (LIYV) closteroviruses. The putative coat protein (CP) was found to be of 46 kDa and its diverged copy of 76 kDa. The potential coding capacity of these notably large closterovirus proteins was confirmed by their expression in vitro and immunoblotting. No proteins with significant similarity to the two C-terminal proteins were identified, but they are related in molecular mass and location to BYV. The gene arrangement as well as the alignments of the closteroviruses CPs and their diverged copies suggest that the mealybug transmissible virus associated with LCD takes an intermediate evolutionary position between the aphid- and whitefly transmissible closteroviruses.