Characterization of polyoma mutants with altered middle and large T-antigens.

The viable polyoma mutants dl1013, dl1014, and dl1015 produced shortened middle and large T-antigens. In mouse 3T3 cells, dl1013 and dl1014 grew at normal rates, and dl1015 grew at a reduced rate. dl1015 behaved phenotypically as a double mutant, with deficiencies both in the stimulation of the host cell and the replication of viral DNA. Only the former defect could be complemented by the ts-a mutant, which produced a normal middle T-antigen and a temperature-sensitive large T-antigen. This result suggests that middle T-antigen is involved in the induction of cellular DNA synthesis. Of the three mutants, dl1015 alone failed to transform rat fibroblasts to growth in semisolid medium. This defect could not be complemented by the ts-a mutant. Determination of the base sequences of the mutant DNAs showed that dl1013 and dl1014 had overlapping deletions of 21 and 9 base pairs, respectively, and that the dl1015 deletion of 30 base pairs was contiguous to the other mutations on their 3' sides. Analyses of the mutant t-antigens showed that all three mutants produced shortened middle T-antigens, whereas only dl1015 large T-antigen was detectably reduced in size.

DNA sequences of polyoma virus early deletion mutants.

The DNA sequences of four "early" viable deletion mutants of polyoma virus have been determined. Two of these (dl-8 and dl-23) are mutants with deletions in the region of the genome that codes for parts of both large and middle T-antigens, and two (dl-6 and dl-28) are mutants with deletions around the viral origin of replication. The former mutants have altered transformation properties relative to wild-type virus, and dl-8 appears to be replication deficient (B. E. Griffin and C. Maddock, J. Virol. 31:645-656, 1979). Sequences are discussed in terms of the altered phenotypes observed for the various mutants, the DNA structures and protein sequences that are affected by the deletions, and how these might affect the biological properties of the mutants.

Polyoma virus DNA: Sequence from the late region that specifies the leader sequence for late mRNA and codes for VP2, VP3, and the N-terminus of VP1.

The DNA sequence of part of the late region of the polyoma virus genome is presented. This sequence of 1,348 nucleotide pairs encompasses the leader region for late mRNA and the coding sequence for the two minor capsid proteins VP2 and VP3. The coding sequence for the N-terminus of the major capsid protein overlaps the C-terminus of VP2/VP3 by 32 nucleotide pairs. From the DNA sequence the sizes and sequences of VP2 and VP3 could be predicted. Potential splicing signals for the processing of late mRNA's could be identified. Comparisons are made between the sequence of polyoma virus DNA and corresponding regions of simian virus 40 DNA.

Coding potential and regulatory signals of the polyoma virus genome.

The complete DNA sequence of the A2 strain of polyoma virus has been determined. It consists of 5,292 base pairs. The sequence is analysed in terms of its coding potential and sites of possible functional significance or structural interest. The polyoma virus genome is compared with those of related tumour viruses, simian virus 40 and BK virus.

Sequence from early region of polyoma virus DNA containing viral replication origin and encoding small, middle and (part of) large T antigens.

The sequence of about one third of the polyoma virus genome is presented. This sequence covers the origin of replication of two large plaque strains (A2 and A3) of polyoma virus. The two strains differ by 11 bp in the origin region. A model for replication is suggested. The sequence probably also covers the entire coding region of two of the polyoma virus early proteins--small and middle T antigens--as well as part of the coding region for large T antigen. Over a small region of the DNA, all three coding frames contain termination codons, which argues a need for spliced early messenger RNAs. In another region of the DNA, two coding frames can be used. Correlation with protein data suggests that one frame codes for part of middle T antigen and the other for part of large T antigen.

Two transfer RNA (1-methylguanine) methylases from yeast.

Two distinct tRNA (m-1G) methylases have been found in the yeast Saccharomyces cerevisiae. They differ in their chromatographic properties on hydroxyapatite, in their response to spermine, and in their site specificity. Only one of the methylases is active against normal tRNA from Escherichia coli.

Transfer RNA methylating activity of yeast mitochondria.

Mitochondria isolated from Saccharomyces cerevisiae and purified in Urografin or sucrose gradient contain tRNA methylating activity with specificities different from those of the cytoplasm. The main reaction product, using E.coli tRNA as methyl group acceptor, is N(2),-N(2)-dimethylguanine. The corresponding mitochondrial methylase is coded by nuclear DNA. A DNA methylating activity is also associated with yeast mitochondria.