Two efficient ribosomal frameshifting events are required for synthesis of mouse mammary tumor virus gag-related polyproteins.

Abstract

The primary translation products of retroviral pol genes are polyproteins initiated in an upstream gene (gag). To investigate the manner in which the gag-initiated polyproteins of techniques mammary tumor virus are produced, we determined in the nucleotide sequence of a 1.8-kilobase DNA fragment that spans the region between gag and pol in the C3H strain of mouse mammary tumor virus. The sequence reveals three overlapping open reading frames: the first encodes products of gag (p27gag and p14gag); the second encodes a protein domain of unknown function (termed X) that is highly related to a similarly positioned sequence in simian type D retroviruses and the viral protease (pro); and the third encodes the reverse transcriptase. The reading frames are organized to permit uninterrupted readthrough from gag to pol if ribosomal frameshifts occur in the -1 direction within each of the two overlapping regions, one of which is 16 nucleotides in length and the other 13 nucleotides. Cell free translation of RNA containing these overlap regions shows that fusion of the reading frames by ribosomal frameshifting occurs efficiently; about one-fourth of the ribosomes traversing the gag-X-/pro overlap and one-tenth traversing the X/pro-pol overlap shift frames, generating gap-related polyproteins in ratios similar to those observed in vivo. Synthetic oligonucleotides containing either of the overlap regions inserted into novel context do not induce frameshifting; hence the overlapping portions of the reading frames are not sufficient to induce a frameshift event, and a larger sequence context or secondary structure may be implicated.