Bacteriophage phi6 contains three pieces of double-stranded RNA (S, M and L) in its nucleocapsid. The pieces possess different genetic information but have similar termini. The packaging of the genomic segments is precise in that virions contain one each of the three normal segments.
The aim of this proposal is to determine how the packaging system is able to select one of each of the chromosomes and how the system acts to replicate the genome. Procapsids produced from cDNA copies of segment L in Escherichia coli are capable of packaging single stranded viral RNA, synthesizing the complementary minus strand to form double stranded RNA and then transcribing this dsRNA to form plus ssRNA. We have developed a novel and unique model for genomic packaging in phi6 that involves specific binding of genomic segments to sites on the outside of the procapsid that appear and disappear in a program that is determined by the amount of RNA within the procapsid. The model is supported by previously unexplained observations as well as findings based upon new model based predictions. We will narrow down the RNA packaging sequences to identify the nucleotides binding directly to the procapsid sites. Then we will identify the procapsid binding sites and show how their exposure changes with the packaging program. The nature of the mechanisms of selective packaging in viruses of eukaryotes having segmented genomes (reoviridae) is currently unknown. The elucidation of the mechanism for phi6 has relevance to the packaging of these viruses. Several members of the reoviridae, e.g. rotavirus and blue tongue virus, are the etiologic agents of important human and animal diseases.
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