Genome Organization of Retroviruses V. In Vitro-Synthesized Moloney Murine Leukemia Viral DNA Has Long Terminal Redundancy

Abstract

Purified virions of Moloney murine leukemia virus can synthesize genome-length double-stranded DNA in vitro. Two predominant species of long DNA transcripts, with average sizes of 9.1 and 8.5 kilobases (kb) can be identified. Both species of DNA contain the negative (complementary to viral RNA) and positive (same polarity as viral RNA) strands. However, only the negative strand of the 8.5-kb species can be identified if the synthesis of DNA is carried out in the presence of the drug actinomycin D. The 9.1-kb species appears to be slightly larger than the genomic RNA. If the linear double-stranded 9.1-kb species is treated with Escherichia coli exonuclease III and allowed to anneal, circular DNA molecules can be observed. Furthermore, polyadenylate-containing short genomic RNA fragments (0.5 to 1.0 kb) can anneal to both the 5′ and the 3′ termini of 9.1-kb complementary DNA. The polyadenylate moiety of the RNA fragments can be identified by tagging it with circular polyoma DNA containing polydeoxybromouridylic acid tails. Thus, the 9.1-kb complementary DNA transcript with two circular polyoma DNA molecules at its termini can be observed. However, when similar annealings are performed with 8.5-kb complementary DNA species, only one end of the resulting molecule has circular polyoma DNA. We conclude that the 9.1-kb complementary DNA species has a large terminal redundancy. The sequences involved in terminal redundancy appear to be derived from the 3′ end of the genomic RNA.