COMPLETE INHIBITION OF VIRION ASSEMBLY IN-VIVO WITH MUTANT PROCAPSID RNA ESSENTIAL FOR PHAGE PHI-29 DNA PACKAGING

A highly efficient method for the inhibition of bacteriophage phi 29 a ssembly was developed with the use of mutant forms of the viral procap sid (or packaging) RNA (pRNA) indispensable for phi 29 DNA packaging, Phage phi 29 assembly was severely reduced in vitro in the presence of mutant pRNA and completely blocked in vivo when the host cell express ed mutant pRNA, Addition of 45% mutant pRNA resulted in a reduction of infectious virion production by 4 orders of magnitude, indicating tha t factors involved in viral assembly can be targets for efficient and specific antiviral treatment, The mechanism leading to the high effici ency of inhibition was attributed to two pivotal features, First, the pRNA contains two separate, essential functional domains, one for proc apsid binding and the other for a DNA-packaging role other than procap sid binding, Mutation of the DNA-packaging domain resulted in a pRNA w ith no DNA-packaging activity but intact procapsid binding competence, Second, multiple copies of the pRNA were involved in the packaging of one genome, This higher-order dependence of pRNA in viral replication concomitantly resulted in its higher-order inhibitory effect, This fi nding suggested that the collective DNA-packaging activity of multiple copies of pRNA could be disrupted by the incorporation of perhaps an individual mutant pRNA into the group, Although this mutant pRNA could not be used for the inhibition of the replication of other viruses di rectly, the principle of using molecules with two functional domains a nd multiple-copy involvement as targets for antiviral agents could be applied to certain viral structural proteins, enzymes, and other facto rs or RNAs involved in the viral life cycle, This principle also impli es a strategy for gene therapy, intracellular immunization, or constru ction of transgenic plants resistant to viral infection.