Abstract
The 3' termini of the three genomic RNAs of BMV contain cis-acting elements essential for the promotion of (-)-strand synthesis. To understand the biological significance of the maintenance of specific base differences in the 3' 200 nucleotides of the three genomic RNAs, all possible permutations were used to exchange these regions among the genomic RNAs. When all RNAs bore the 3' end of RNA-1, the total RNA accumulation was only 15% of wild-type; when the 3' end of RNAs -2 or -3 was present on all three RNAs, the total RNA accumulation was reduced to 30 or 35% of wild-type. Two major processes were found to be involved in these dramatic differences. The first reflects the distinct and competitive strengths of the (-)-strand promoters in these sequences. The second is the importance of the context of upstream sequences in which the 3' promoter is placed. The transfer of previously constructed mutations to the 3' end of RNA-1 helped in understanding the role in BMV RNA replication of RNA-1 and the la protein encoded by it. The debilitating effect on replication of aminoacylation defective mutations when present on RNAs -1 and -2 suggests a role for aminoacylation in the replication of RNAs encoding replication-essential proteins. To understand the reason(s) behind the persistence of base differences among the three genomic RNAs and the impairing effect on replication of mutations in the 3' end, RNA-protein binding techniques were developed. Contaminating coat protein in the BMV replicase extract was found to interact specifically with two domains of RNA-1. A mutant of coat protein (lacking the first 25 amino acids) that was previously found to be defective in encapsidation was found to be incapable of binding RNAs in vitro. On the other hand, the wild-type coat protein was found to bind RNAs in a cooperative manner, which is believed to be the mechanism responsible for rapid encapsidation in vivo.
Duggal, Rohit (1993). Modulation of replication and RNA-protein interactions in brome mosaic virus. Texas A&M University. Texas A&M University. Libraries. Available electronically from
https : / /hdl .handle .net /1969 .1 /DISSERTATIONS -1525827.