Retrotransposons are mobile genetic elements that replicate by reverse transcription. Retrotransposons are important vehicles for genome change and are frequently the causal agents of mutation, gene duplication and chromosome rearrangements. Retrotransposons have been found in all eukaryotic genomes analyzed to date, and they often constitute a substantial fraction of the nuclear DNA. In maize, for example, over 50% of the genome is retrotransposon in origin. The ubiquity and abundance of these sequences speaks to their effectiveness as genetic parasites. The research in the Voytas laboratory focuses on understanding the retrotransposon/host relationship and how retrotransposons have adapted to life in their host’s chromosomes.
Target specificity of the Saccharomyces cerevisiae Ty5 retrotransposons
The Ty5 retrotransposons of Saccharomyces preferentially integrate near the telomeres or the silent mating loci. These regions are bound in silent chromatin, which is analogous to heterochromatin of higher eukaryotes. Because the telomeres are largely devoid of coding information, target specificity may represent a compromise between Ty5 and its host: the telomeres are a safe haven wherein retrotransposons can integrate without causing deleterious genetic effects.
To understand the mechanism of Ty5 target specificity, the Voytas lab generated several Ty5 mutants that no longer integrate into silent chromatin (Mol. Cell 1:1051; see also minireview in Cell 93:1087). The mutations cluster within a span of six amino acids (called the targeting domain) near the C-terminus of integrase, a poorly defined region of the protein in both the retrotransposons and retroviruses. The targeting domain interacts with Sir4p, a well-studied protein component of silent chromatin. The interaction between the Ty5 targeting domain and Sir4p is necessary and sufficient for targeted integration (PNAS 100:5891; see also minireviews in PNAS 100:5586 and Cell 115:135). The targeting determinants of Ty5 integrase can be replaced with other peptides that have known interacting partners. Ty5 elements with these modified integrases now have altered target specificity: they insert at DNA locations to which the protein partner is bound. These experiments demonstrate that retroelement target specificity can be manipulated, opening up the possibility to engineer retroviruses with altered target site choice and offering promise for improved vectors for gene therapy.
How widespread is retrotransposon targeting specificity?
Current work in the Voytas lab continues to dissect the mechanism of Ty5 target specificity. In addition, other retrotransposons are being analyzed to determine how widely targeting mechanisms are utilized. A particular emphasis is being placed on retrotransposons that encode chromodomains within their integrase C-termini. Chromodomains interact with modified residues on histones, and the hypothesis being tested is whether integrase chromodomains direct integration to specific chromatin domains through recognition of the histone code.