Characterization and detection of sc4: a sixth gene encoded by sonchus yellow net virus

The nucleotide sequence of a sixth gene (sc4) of the plant rhabdovirus sonchus yellow net virus (SYNV) was determined from viral genomic and poly(A)+ cDNA clones. The sc4 gene is 1196 nucleotides (nt) and has an open reading frame of 972 nt that is capable of encoding a protein of 324 amino acids. Primer extension analyses of poly(A)+ RNA isolated from infected plants indicate that the 5' end of the sc4 mRNA corresponds to nucleotide 2840, relative to the 3' end of the minus-sense genomic RNA and extends to nucleotide 4035. A 43-nt untranslated leader sequence precedes the predicted first AUG codon and a 181-nt untranslated sequence follows the translational stop codon. This gene is similar to the other SYNV genes in that it is flanked on each side by a conserved gene junction sequence. Polyclonal antibodies raised to an sc4 fusion protein react with a 37-kDa protein in virus-infected plants that is close to the predicted size of the sc4 protein. Western blot analyses of cellular fractionations from infected plants show that sc4 is membrane associated and sucrose density gradient analyses demonstrate that sc4 sediments in the same fractions as SYNV virions. Analysis of the sc4 open reading frame reveals that 16% of the amino acids are serine or threonine residues and that the protein has four potential consensus casein kinase II phosphorylation sites. The deduced amino acid sequence of sc4 also contains a motif related to alpha amylases and aspartic proteases. This completes the sequence determination of the 13,720-nt SYNV genome.