RNA2 of grapevine fanleaf virus: sequence analysis and coat protein cistron location

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Abstract
The nucleotide sequence of the genomic RNA2 (3374 nucleotides) of grapevine fanleaf virus strain F13 was determined from overlapping cDNA clones and its genetic organization was deduced. Two rapid and efficient methods were used for cDNA cloning of the 5'' region of RNA2. The complete sequence contained only one long open reading frame of 3555 nucleotides (1184 codons, 131 K product). The analysis of the N-terminal sequence of purified coat protien (CP) and identification of its C-terminal residue have allowed the CP cistron to be precisely positioned within the polyprotein. The CP produced by proteolytic cleavage at the Arg/Gly site between residues 680 and 681 contains 504 amino acids (Mr 56019) and has hydrophobic properties. The Arg/Gly cleavage site deduced by N-terminal amino acid sequence analysis is the first for a nepovirus coat protein and for plant viruses expressing their genomic RNAs by polyprotien synthesis. Comparison of GFLV RNA2 with M RNA of cowpea mosaic comovirus and with RNA2 of two closely related neopoviruses, tomato black ring virus and Hungarian grapevine chrome mosaic virus, showed strong similarities among the 3'' non-coding regions but less similarity among the 5'' end non-coding sequences than reported among other nepovirus RNAs.