Access to DNA establishes a secondary target site bias for the yeast retrotransposon Ty5

Abstract

Integration sites for many retrotransposons and retroviruses are determined by interactions between retroelement-encoded integrases and specific DNA-bound proteins. The Saccharomyces retrotransposon Ty5 preferentially integrates into heterochromatin because of interactions between Ty5 integrase and the heterochromatin protein silent information regulator 4. We mapped over 14,000 Ty5 insertions onto the S. cerevisiae genome, 76% of which occurred in heterochromatin, which is consistent with the known target site bias of Ty5. Using logistic regression, associations were assessed between Ty5 insertions and various chromosomal features such as genome-wide distributions of nucleosomes and histone modifications. Sites of Ty5 insertion, regardless of whether they occurred in heterochromatin or euchromatin, were strongly associated with DNase hypersensitive, nucleosome-free regions flanking genes. Our data support a model wherein silent information regulator 4 tethers the Ty5 integration machinery to domains of heterochromatin, and then, specific target sites are selected based on DNA access, resulting in a secondary target site bias. For insertions in euchromatin, DNA access is the primary determinant of target site choice. One consequence of the secondary target site bias of Ty5 is that insertions in coding sequences occur infrequently, which may preserve genome integrity.T he insertion of mobile genetic elements into new chromosomal sites profoundly impacts genome structure and evolution. For many mobile elements, integration sites are not chosen randomly. Target site biases are particularly well-documented for the LTR retrotransposons and retroviruses (1-3). These retroelements replicate by reverse-transcribing mRNA into cDNA and then inserting the cDNA into their host's genome using an element-encoded integrase (IN). Retrotransposons are among the most abundant interspersed repeats in eukaryotic genomes, and retroviruses are often used as vectors for gene therapy. Understanding mechanisms of retroelement target site choice, therefore, has value for both basic and applied research.In the best studied cases, retroelement target site choice is dictated by interactions between IN and specific DNA-bound proteins. HIV IN, for example, interacts with the transcription coactivator lens epithelial-derived growth factor (4), and sites of HIV integration are influenced by sites of this protein's chromosomal occupancy (5). The role of chromatin in target site choice is also well-established for model yeast retrotransposons. The Schizosaccharomyces pombe Tf1 element inserts preferentially into regions upstream of some genes transcribed by RNA polymerase (pol) II (6). Tf1 IN interacts with the transcription factor Atf1p (7), and at the fbp1 promoter, Atf1p alone mediates target site choice (8). The Saccharomyces cerevisiae Ty1 and Ty3 retrotransposons prefer to integrate upstream of genes transcribed by RNA pol III, likely because of interactions between IN and components of the pol III machinery or associated chromatin (9, 10)....

A nucleosomal surface defines an integration hotspot for the Saccharomyces cerevisiae Ty1 retrotransposon

“…Biases in insertion-site patterns may result if HIS3 is not expressed in certain chromosomal environments. However, in previous work with the related yeast retrotransposon Ty5, we found HIS3 to be a very robust reporter for recovering insertions in heterochromatin-the preferred sites of Ty5 integration (Baller et al 2011). Additionally, the experimental approach for recovering Ty1 insertions in the companion study by Mularoni et al (2011) did not select cells harboring Ty1 integration events, and yet produced a similar genome-wide pattern of insertions.…”

“…Like Ty1, however, insertions of HIV and gammaretroviruses were both associated with epigenetic modifications correlated with transcription. Ty1's preference for nucleosomal DNA stands in contrast to the related S. cerevisiae retrotransposon Ty5, which prefers nucleosome-free DNA for integration (Baller et al 2011). Nucleosomes are also avoided by the DNA transposon Hermes when it transposes in yeast (Gangadharan et al 2010).…”

“…The use of high-throughput DNA sequencing to map large numbers of transposable element insertions is increasingly employed to understand how mobile elements interface with their host genome (Gangadharan et al 2010;Guo and Levin 2010;Baller et al 2011). In species such as Saccharomyces cerevisiae, the availability of genome-wide data sets for a large number of chromosomal features and functions (e.g., histone modifications or sites of DNA replication) makes it possible to relate insertion sites to diverse aspects of genome biology.…”

Telomerase and retrotransposons: Reverse transcriptases that shaped genomes

“…Ty5 preferentially targets heterochromatic regions of the yeast genome via an interaction between Ty5 integrase and the silencing protein Sir4. As reported in the accompanying article (35), high-throughput DNA sequencing of a library of Ty5 integration sites in the Saccharomyces cerevisiae genome yielded a surprising finding: nearly one-quarter of Ty5 retrotransposition events are euchromatic. Further analysis of both heterochromatic and euchromatic insertions uncovered a secondary Ty5 target site, in DNase I-sensitive, nucleosome-free regions.…”

“…Further analysis of both heterochromatic and euchromatic insertions uncovered a secondary Ty5 target site, in DNase I-sensitive, nucleosome-free regions. In the accompanying article (35), Voytas speculates that Sir4 brings the cDNA-integrase complex to the vicinity of heterochromatin, but specific sites of insertion are governed by the accessibility of the DNA. DNA is most accessible in the nucleosome-free regions flanking ORFs; hence both the primary and this newly revealed secondary target site bias give rise to Ty5 retrotransposition events that rarely disrupt essential coding sequences.…”