Date of Award
Doctor of Philosophy
Genetics, Development and Cell Biology
Daniel F. Voytas
The retrotransposons and retroviruses display a strong preference in selecting integration sites. The molecular mechanisms underlying this process, however, are largely unknown. I have characterized a new retrotransposon, called Ty5, in diverse strains of Saccharomyces cerevisiae and related species. A functional element, Ty5-6p, was identified from S. paradoxus. To investigate Ty5 target preference, the locations of 13 native insertions were determined in S. cerevisiae and S. paradoxus. Twelve were found near the telomeres and the mating locus HMR. To determine if this distribution is a consequence of targeted integration, a Ty5 transposition assay was developed in S. cerevisiae using Ty5-6p. The locations of 19 elements on chromosome III and six on other chromosomes were mapped. Twenty-two were near the telomeres and transcriptional silencers flanking HML and HMR, which share a common chromatin structure that represses transcription of adjacent genes (silent chromatin). These observations indicate that Ty5 preferentially transposes to silent chromatin. This target preference has likely had consequences for the genomic organization of chromosome ends. Recombination between elements is evidenced by the lack of target site duplications among native Ty5 insertions. In addition, native elements mark telomeric regions that have been duplicated and rearranged in S. cerevisiae and S. paradoxus. Molecular mechanisms of Ty5 targeting were investigated in various mutants that affect silent chromatin. HMR-E is the cis-sequence required for assembly of silent chromatin at HMR and accounts for approximately 2% of de novo transpositions. Mutations at HMR-E, which disrupt silent chromatin, abolish Ty5 transposition to this region. Deletion of SIR3, a gene that encodes a protein component of silent chromatin, resulted in an approximately six fold decrease in transposition. By analogy to Ty3 and HIV, we hypothesize that Ty5 target preference is due to interactions between its integration complex and protein components of silent chromatin. Ty5 target preference extends the link between telomere structure and reverse transcription as carried out by telomerase and Drosophila retrotransposons.
Digital Repository @ Iowa State University, http://lib.dr.iastate.edu/
Zou, Sige, "Integration specificity of the retrovirus-like transposable element Ty5 of Saccharomyces " (1996). Retrospective Theses and Dissertations. 11353.