Experimental test of a TMV replication model. Attempts to identify molecular precursors of TMV-RNA and determine the time required to synthesize a TMV-RNA molecule

Abstract

A kinetic model of TMV replication based on the observed virus accumulation and known models of RNA virus replication has been tested experimentally by applying it to the identification of molecular precursors of TMV-RNA and the time required to synthesize a TMV-RNA molecule. We have kinetically characterized a MW 4.0 × 106 RNA species (component “Y”) from TMV-infected leaves which accumulates 32P in a manner consistent with the predictions of our model for a double-stranded replicative form (RF) intermediate in TMV-RNA replication. Another high molecular-weight RNA component (3 × 106) was studied with respect to its kinetics of 32P incorporation. This RNA species (component “X”) is shown to accumulate at late stages of infection and to incorporate 32P at rates which indicate that it is not a precursor of TMV-RNA.Estimates from analysis of early (exponential) stages of TMV infection suggest that the time required to synthesize a TMV-RNA molecule is on the order of 20 min. This result assumes a replicative intermediate which synthesizes only one TMV-RNA molecule at a time. Data obtained at late (linear) stages of infection suggest that, relative to the early stage, there are proportionately more nascent TMV-RNA plus strands per replicative intermediate and that metabolic turnover of minus-strand templates occurs at a significant rate.