DNA Integration by Ty Integrase in <i>yku70</i>Mutant <i>Saccharomyces cerevisiae</i> Cells

Kiechle, Markus; Friedl, Anna A.; Manivasakam, Palaniyandi; Eckardt‐Schupp, Friederike; Schiestl, Robert H.

doi:10.1128/mcb.20.23.8836-8844.2000

Cited by 11 publications

(13 citation statements)

References 45 publications

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“…In Ku-deficient yeast cells, increased 5′–3′ single-strand degradation is known to occur (Lee et al 1998; Tomita et al 2003), and indeed end-processing of the plasmid was frequently seen in Ku-deficient cells (Kiechle et al 2000). In some of the composite elements, extensive microhomology was present at the plasmid-Ty junctions, and it seems reasonable to assume that exposure of single-stranded regions in the course of end-processing was instrumental in the formation of the junctions.…”

Section: Discussionmentioning

confidence: 99%

Ty1 integrase overexpression leads to integration of non-Ty1 DNA fragments into the genome of Saccharomyces cerevisiae

Friedl

Kiechle

Maxeiner³

et al. 2010

Mol Genet Genomics

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The integrase of the Saccharomyces cerevisiae retrotransposon Ty1 integrates Ty1 cDNA into genomic DNA likely via a transesterification reaction. Little is known about the mechanisms ensuring that integrase does not integrate non-Ty DNA fragments. In an effort to elucidate the conditions under which Ty1 integrase accepts non-Ty DNA as substrate, PCR fragments encompassing a selectable marker gene were transformed into yeast strains overexpressing Ty1 integrase. These fragments do not exhibit similarity to Ty1 cDNA except for the presence of the conserved terminal dinucleotide 5′-TG-CA-3′. The frequency of fragment insertion events increased upon integrase overexpression. Characterization of insertion events by genomic sequencing revealed that most insertion events exhibited clear hallmarks of integrase-mediated reactions, such as 5 bp target site duplication and target site preferences. Alteration of the terminal dinucleotide abolished the suitability of the PCR fragments to serve as substrates. We hypothesize that substrate specificity under normal conditions is mainly due to compartmentalization of integrase and Ty cDNA, which meet in virus-like particles. In contrast, recombinant integrase, which is not confined to virus-like particles, is able to accept non-Ty DNA, provided that it terminates in the proper dinucleotide sequence.

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Section: Discussionmentioning

confidence: 99%

Ty1 integrase overexpression leads to integration of non-Ty1 DNA fragments into the genome of Saccharomyces cerevisiae

Friedl

Kiechle

Maxeiner³

et al. 2010

Mol Genet Genomics

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“…Most insertion events present hallmarks of IN-mediated events, such as 5-bp TSDs and Ty1 target site preferences. IN-mediated insertion of non-Ty1 DNA fragments has also been observed in cells lacking the Ku70 protein, suggesting that a Ku70-dependent mechanism restricts endogenous IN activity to Ty1 cDNA [177]. …”

Section: Post-translational Steps In Retrotranspositionmentioning

confidence: 99%

The Ty1 LTR-Retrotransposon of Budding Yeast,Saccharomyces cerevisiae

2015

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Summary Long-terminal repeat (LTR)-retrotransposons generate a copy of their DNA (cDNA) by reverse transcription of their RNA genome in cytoplasmic nucleocapsids. They are widespread in the eukaryotic kingdom and are the evolutionary progenitors of retroviruses [1]. The Ty1 element of the budding yeast Saccharomyces cerevisiae was the first LTR-retrotransposon demonstrated to mobilize through an RNA intermediate, and not surprisingly, is the best studied. The depth of our knowledge of Ty1 biology stems not only from the predominance of active Ty1 elements in the S. cerevisiae genome but also the ease and breadth of genomic, biochemical and cell biology approaches available to study cellular processes in yeast. This review describes the basic structure of Ty1 and its gene products, the replication cycle, the rapidly expanding compendium of host co-factors known to influence retrotransposition and the nature of Ty1's elaborate symbiosis with its host. Our goal is to illuminate the value of Ty1 as a paradigm to explore the biology of LTR-retrotransposons in multicellular organisms, where the low frequency of retrotransposition events presents a formidable barrier to investigations of retrotransposon biology.

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“…One possibility is that Ku might facilitate the integration process by stabilizing breaks created by the integrase. Alternatively, Ku might ensure Ty1 integrase substrate-specificity since it was reported that Ty1 integrase mediates non-homologous integration of foreign DNA in the absence of Ku (Kiechle et al, 2000). How Ku might prevent integrase from accepting non-Ty DNA as a substrate has not yet been elucidated.…”

Section: Post-transcriptional Regulations Of Ty1 Retrotransposition Imentioning

confidence: 99%

Happy together: the life and times of Ty retrotransposons and their hosts

Lesage¹,

Todeschini²

2005

Cytogenet Genome Res

118

125

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The aim of this review is to describe the level of intimacy between Ty retrotransposons (Ty1–Ty5) and their host the yeast Saccharomyces cerevisiae. The effects of Ty location in the genome and of host proteins on the expression and mobility of Ty elements are highlighted. After a brief overview of Ty diversity and evolution, we describe the factors that dictate Ty target-site preference and the impact of targeting on Ty and adjacent gene expression. Studies on Ty3 and Ty5 have been especially informative in unraveling the role of host factors (Pol III machinery and silencing proteins, respectively) and integrase in controlling the specificity of integration. In contrast, not much is known regarding Ty1, Ty2 and Ty4, except that their insertion depends on the transcriptional competence of the adjacent Pol III gene and might be influenced by some chromatin components. This review also brings together recent findings on the regulation of Ty1 retrotransposition. A large number of host proteins (over 30) involved in a wide range of cellular processes controls either directly or indirectly Ty1 mobility, primarily at post-transcriptional steps. We focus on several genes for which more detailed analyses have permitted the elaboration of regulatory models. In addition, this review describes new data revealing that repression of Ty1 mobility also involves two forms of copy number control that act at both the trancriptional and post-transcriptional levels. Since S. cerevisiae lacks the conserved pathways for copy number control via transcriptional and post-transcriptional gene silencing found in other eukaryotes, Ty1 copy number control must be via another mechanism whose features are outlined. Ty1 response to stress also implicates activation at both transcriptional and postranscriptional steps of Ty1. Finally, we provide several insights in the role of Ty elements in chromosome evolution and yeast adaptation and discuss the factors that might limit Ty ectopic recombination.

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DNA Integration by Ty Integrase in yku70Mutant Saccharomyces cerevisiae Cells

Cited by 11 publications

References 45 publications

Ty1 integrase overexpression leads to integration of non-Ty1 DNA fragments into the genome of Saccharomyces cerevisiae

Ty1 integrase overexpression leads to integration of non-Ty1 DNA fragments into the genome of Saccharomyces cerevisiae

The Ty1 LTR-Retrotransposon of Budding Yeast,Saccharomyces cerevisiae

Happy together: the life and times of Ty retrotransposons and their hosts

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