Plum pox potyvirus RNA replication in a crude membrane fraction from infected Nicotiana clevelandii leaves.

In vitro synthesis of plum pox potyvirus (PPV)-specific nucleic acid has been measured in a crude fraction prepared from leaves of PPV-infected Nicotiana clevelandii plants. Using alkali and DNase treatments, the synthesized nucleic acid was shown to be RNA. The electrophoretic mobility and the differing sensitivity to RNase at high and low salt concentrations allowed the identification of in vitro products probably corresponding to replicative form and replicative intermediate RNA, as well as to single-stranded RNA. Most of the PPV-specific RNA synthesized was shown to be of positive polarity. The in vitro RNA synthesis, performed in the presence of actinomycin D, required all four ribonucleoside triphosphates and Mg2+ ions. This enzyme extract contained about 6% of the leaf protein and most of the identified virus-encoded proteins. Altogether, the results presented in this paper suggest that in vitro RNA synthesis was carried out by the PPV replicase complex.