Simian virus 40 mutants carrying extensive deletions in the 72-base-pair repeat region

The 5' ends of the simian virus 40 (SV40) late RNAs are heterogeneous in location, spanning a 300-nucleotide region from residues 28 to 325. To examine whether upstream or downstream measuring functions analogous to the TATA box play roles in positioning the 5' ends of these RNAs, we determined by S1 and primer extension mapping the locations of the 5' ends of the late viral RNAs made in monkey cells infected with: (i) three wild-type strains of SV40 that contain tandem duplications of the enhancer region that are 64, 85, and 91, rather than 72, base pairs in length; (ii) four viable mutants that contain alterations in the 21-base-pair tandem repeats; and (iii) four viable mutants that possess small deletions or insertions at or near the major cap site at residue 325. Most of the 5' ends of the RNAs were identical in location to those seen with wild-type strain 776. The only exceptions were the absence of RNAs whose 5' ends mapped to within three bases upstream or downstream of a sequence alteration. In addition, the sequences within residues 251 to 277 that function as transcriptional initiation sites in wild-type strain 776 also did so in their second locations in the wild-type strains in which these sequences are duplicated. Differences were noted in the relative abundances of the numerous 5' ends of the late RNAs, even among the wild-type strains. These findings indicate that many (and likely all) of the approximately two dozen locations of 5' ends of SV40 late RNAs are each determined largely by sequences within their immediate vicinity. However, sequences somewhat removed from these transcriptional initiation sites may modulate the efficiencies with which they are utilized.

Section: Kesultssupporting

confidence: 91%

Sequences involved in determining the locations of the 5' ends of the late RNAs of simian virus 40

Somasekhar¹,

Mertz²

1985

“…3C; Table 1). In addition to differences in the ratios of the various spliced RNA species, the precise sequences of the leader regions of these RNAs differ because the deletions both remove specific sequences and shift upstream the locations of the 5' ends of the 19S RNAs (13,14,45,55).…”

Section: Resultsmentioning

confidence: 99%

Both VP2 and VP3 are synthesized from each of the alternative spliced late 19S RNA species of simian virus 40

Good

Welch

Barkan

et al. 1988

The late 19S RNAs of simian virus 40 consist of a family of alternatively spliced RNAs, each of which contains open reading frames corresponding to all three of the virion proteins. Two approaches were used to test the hypothesis that each alternatively spliced 19S RNA species is translated to synthesize preferentially only one of the virion proteins. First, we analyzed the synthesis of virion proteins in simian virus 40 mutant-infected monkey cells that accumulate predominantly either only one spliced 19S RNA species or only the 19S RNAs. Second, we determined the virion proteins synthesized in a rabbit reticulocyte lysate programmed with specific, in vitro-transcribed 19S RNA species. These results indicated that VP2 and VP3, but not VP1, are synthesized from all 19S RNA species. Quantitative analysis of these data indicated that individual 19S RNA species containing a translation initiation signal upstream of the VP2 AUG codon were translated in a cell extract three- to fivefold less efficiently than were 19S RNA species lacking this signal and that the precise rate of synthesis of VP2 relative to VP3 varied somewhat with the sequence of the leader region. These data are consistent with the synthesis of VP2 and VP3 occurring by a leaky scanning mechanism in which initiation of translation at a specific AUG codon is affected by both (i) the intrinsic efficiency of ribosomes recognizing the sequences surrounding the AUG codon as an initiation signal and (ii) partial interference from 5'-proximal initiation signals and their corresponding open reading frames.

“…It has been shown recently that the 216-bp BKV Dunlop fragment containing three 68-bp repeats can act as an enhancer for the chloramphenicol acetyltransferase gene expression from the SV40 promoter (15). The SV40 72-bp enhancer is essential for in vivo viability (5,7) and affects the host range of this virus (2,11,12,22), and the structure of the similar region of mouse polyomavirus determines whether this virus can grow in teratocarcinoma cells (6,9,10,17 Fig. 1B, C, D, and E. The explanation for boxes, small triangles, and small circles is given in the legend to Fig.…”

Section: Resultsmentioning

confidence: 99%

DNA rearrangement affecting expression of the BK virus transforming gene

Watanabe¹,

Soeda²,

Uchida³

et al. 1984

BK virus mutant pm-522, forming small turbid plaques on human cells, can transform rat or hamster cells much more efficiently than the wild-type BK virus (wt-501) does. We compared the nucleotide sequence of wt-501 Hindlll C segment with that of pm-522 HindIII-C, which contains the mutation responsible for the altered plaque type and transforming capacity. The difference between the two BK viruses was the local DNA rearrangement (deletions and duplications) that had occurred in the putative control region for early transcription in pm-522 DNA. Whereas wt-501 had three sets of 68-base pair repeats (the central set had a deletion of 18 base pairs) in this region, pm-522 had one set of 68-base-pair units and two sets of shorter 37base-pair repeats. Three BK virus mutants, forming clear large plaques like those of wt-501 but capable of transforming rat cells, were derived from the recombinant virus carrying the Hindlll C segment of pm-522. These mutants had further duplications of short segments originating from the pm-522 sequence in the putative early control region.