Influence of the transduced 3'UTR of the c-src oncogene on tumour growth induced by the v-src gene of avian sarcoma virus PR2257.

Avian sarcoma virus PR2257 contains 952 bp transduced from the left part of the 3'UTR of the chicken c-src oncogene. Deletion mutants were constructed to determine the effect of the 3'UTR on tumorigenicity in vivo and in vitro. In the presence of the 3'UTR, tumours were 3.4 times larger in vivo, and tumorigenicity was increased 2.5-fold in vitro. Several regulatory submotifs were also found within the 3'UTR. Parts of the 3'UTR were cloned into the LTR CAT plasmid and analysed for CAT expression. A 170 bp element was found to be responsible for the enhanced expression of the CAT gene. These results demonstrate the effect of the transduced 3'UTR sequence during long-term interaction between PR2257 virus and the chicken genome, and suggest a novel regulatory mechanism of the src oncogene.

Origin and evolution of the c-src-transducing avian sarcoma virus PR2257.

Avian sarcoma virus PR2257 transduced de novo the c-src gene and about 900 bp of 3' non-coding sequences belonging to the src locus. This virus contains only one mutation in the c-src coding sequence causing a reading frame shift after Pro-525. The molecular clone studied was derived from a cell line of transformed quail fibroblasts, C7. It contains endogenous virus (ev) derived sequences in the U5 and 3' non-coding regions, indicating that multiple recombination occurred with endogenous virus. Here we investigated the possible evolution of PR2257 when the original tumour was repeatedly passaged in vivo. After 16 passages a new virus, designated PR2257/16, appeared with a tenfold higher titre. The sequence of PR2257/16 was determined and showed that PR2257/16 resulted from recombination of PR2257 with the env gene of the helper virus (td daPR-C). This recombination expanded the env gene content in PR2257/16 and, in addition, five point mutations occurred in its genome. Because we thought that an endogenous virus might be involved in the mechanism of c-src transduction, we also reinvestigated the presence of ev sequences in PR2257 proviruses from several early passages of the original tumour. We found that in contrast with the first isolate from the C7 cell line, the provirus in these tumours did not contain such sequences. These results do not support the hypothesis that endogenous sequences were involved in the transduction process.

Microtubular dynamics of Saccharomyces cerevisiae temperature-sensitive secretory mutants: the sec 1 product and polymerization of microtubules.

The relation of cytoplasmic microtubules to intracellular transport was studied in temperature-sensitive (ts) secretory mutants of Saccharomyces cerevisiae at permissive and nonpermissive temperature using indirect immunofluorescence with monoclonal antibody TU-01 against alpha-tubulin. It was found that in the sec 1 mutant, which at 37 degrees C accumulated secretory vesicles and in which therefore transport of secretory material from secretory vesicles to cytoplasmic membrane was inhibited, cytoplasmic and in some cases nuclear microtubules were impaired. After 4 h of postcultivation at 24 degrees C the altered phenotype reverted to the original state, the cells began to divide, and were comparable with control. Use of the sec 1 mutant protoplasts suggested that the product, whose gene is mutated, is probably involved in microtubular polymerization. The sec 7 mutant, which accumulates the Golgi complex under nonpermissive conditions and in which the transfer of secretory material from the Golgi complex to the secretory vesicles is thus inhibited, showed no significant changes in the length or number of cytoplasmic microtubules. As a result of secretory product accumulation, the cytoplasmic microtubules were displaced towards the periphery in some cells.