Department of Molecular Biology and Biochemistry, Rutgers University, CABM 306, 679 Hoes Lane, Piscataway, New Jersey 08855, USA
Ty1, the genetically tractable retrotransposable element found
in the yeast
Saccharomyces cerevisiae, closely resembles vertebrate
retroviruses both in structure and in mechanism of replication.
By direct sequence analysis, we examined the rate and spectrum
of new mutations appearing during a single cycle of Ty1 replication.
The rate of new mutations was comparable to those seen for replicating
retroviruses. All observed changes were base substitutions,
and their location suggested that template ends may be hot spots
for generating these mutations. To test this, we developed methods
to examine, at the nucleotide level, the end structure of the
expected Ty1 replication intermediates. Our results demonstrate
that Ty1 reverse transcriptase can add terminal non-templated
bases
in vivo during each step in replication. Furthermore,
Ty1 RNAse H creates multiple template ends by imprecisely cleaving
RNA. This expands the range of sites of subsequent non-templated
base addition. Finally, on reaching template ends, Ty1 reverse
transcriptase can strand transfer to inappropriate templates.
Taken together, these mutagenic mechanisms may influence the
evolution of particular regions of the Ty1 genome and serve
as a mechanism to regulate the overall level of Ty1 transposition
in its host cell.
