COAT PROTEIN STIMULATES REPLICATION COMPLEXES OF ALFALFA MOSAIC-VIRUSTO PRODUCE VIRION RNAS IN-VITRO

Viral replication complexes (RCs) were gradient-purified from cowpea m esophyll protoplasts 21 h after inoculation with alfalfa mosaic virus. These membranous structures incorporate [P-32]UMP into double- and si ngle-stranded RNAs in the absence of added template. When coat protein is added prior to the reaction the incorporation in both RNA fraction s is stimulated several times. Part of the single-stranded product RNA s are released from the RCs. The stimulation of incorporation in high molecular mass RNAs by coat protein can be mimicked only to a certain extent by addition of a ribonuclease inhibitor or of an excess of vira l RNA prior to the reaction. This shows that the coat protein is not o nly protecting the product of the RCs against degradation by ribonucle ase, but that it is stimulating the synthesis and release of viral RNA s from RCs as well. This leads to the hypothesis that with alfalfa mos aic virus some coat protein is necessary for the release of messenger RNA from the RC. The hypodiesis explains why the viral genome RNAs, al though they are of messenger polarity, cannot start a replication cycl e in the absence of coat protein: RCs containing the parental RNAs cou ld be formed but no amplification of them could take place since no me ssenger RNAs needed for the production of viral polymerase proteins wo uld be released into the cytoplasm.