A Circular Plasmid from the YeastTorulaspora delbrueckii

Blaisonneau, Joël; Sor, Frédéric; Chéret, Geneviève; Yarrow, D.; Fukuhara, Hiroshi

doi:10.1006/plas.1997.1315

Cited by 31 publications

(29 citation statements)

References 21 publications

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“…Three additional yeast recombinases have been described; we did not try TD1 (46), and our initial attempts to use SM (34) and KW (34) were unsuccessful. In addition to the yeast recombinases, we also tested the ability of Dre, a recombinase closely related to Cre, to work in Drosophila.…”

Section: Results Andmentioning

confidence: 99%

Multiple new site-specific recombinases for use in manipulating animal genomes

Nern

Pfeiffer

Svoboda

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

159

171

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Site-specific recombinases have been used for two decades to manipulate the structure of animal genomes in highly predictable ways and have become major research tools. However, the small number of recombinases demonstrated to have distinct specificities, low toxicity, and sufficient activity to drive reactions to completion in animals has been a limitation. In this report we show that four recombinases derived from yeast-KD, B2, B3, and R-are highly active and nontoxic in Drosophila and that KD, B2, B3, and the widely used FLP recombinase have distinct target specificities. We also show that the KD and B3 recombinases are active in mice.gene expression | genetic engineering S ite-specific DNA recombinases are widely used in multicellular organisms to manipulate the structure of genomes and, in turn, to control gene expression (for reviews see refs. 1-4). These enzymes, derived from bacteria and fungi, catalyze directionally sensitive DNA exchange reactions between short (30-40 nucleotides) target site sequences that are specific to each recombinase (5). These reactions enable four basic functional modules-excision/insertion, inversion, translocation and cassette exchange-that have been used individually or combined in a wide range of configurations to control gene expression (Fig. 1A).The use of site-specific recombination to manipulate genomes has been limited by the availability of recombinases with high activity, distinct site specificity, and low toxicity. In Drosophila, the most widely used recombinase is FLP, encoded by the Saccharomyces cerevisiae 2-μm plasmid (6). FLP was first shown to work in a heterologous, multicellular organism by Golic and Lindquist in 1989 (7) who demonstrated the excision reaction on chromosomally inserted target sites (FRTs). Since that time FLP/FRT recombination has been widely used in Drosophila in applications based on excision (8) and translocation (9-11).Complex manipulations of genome structure can require the use of more than one of the modules diagrammed in Fig. 1A, or parallel independent implementations of the same module, in a single individual. To accomplish such manipulations, the modules must be implemented with different recombinases that do not recognize each other's target sites. Similarly, a number of powerful methods have been developed for using the excision and inversion reactions to control expression of a transgene specifically in cells where two independent gene expression patterns overlap (2,3,12,13). Such intersectional methods rely on pairs of orthogonal recombinases; see, for example, Fig. 1B. For these reasons, we sought to discover additional recombinases with distinct site-specificity.FLP recombinase has been mutated to recognize altered FRT sites, but some cross-reaction still remains (14, 15). Cre, encoded by the bacteriophage P1, is the most widely used recombinase in mammalian cells (16)(17)(18). Cre functions in Drosophila (19), but exhibits obvious toxicity (20), a problem also observed in mammalian cells (21, 22; reviewed in ref. 23) an...

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Section: Results Andmentioning

confidence: 99%

Multiple new site-specific recombinases for use in manipulating animal genomes

Nern

Pfeiffer

Svoboda

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

159

171

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“…The switch to point centromeres appears to have occurred contemporaneously with the loss of most or all of the machinery required for heterochromatin assembly and RNA interference (1). Quite remarkably, the presence of stably propagating 2m circle-related nuclear plasmids appears to be unique to the Saccharomycetaceae lineage as well (3). It is reasonable, therefore, that the partitioning locus of an ancestral plasmid, following chromosomal integration, could have served as the progenitor of the point centromere (29).…”

Section: Discussionmentioning

confidence: 99%

Cse4 (CenH3) Association with the Saccharomyces cerevisiae Plasmid Partitioning Locus in Its Native and Chromosomally Integrated States: Implications in Centromere Evolution

Huang

Hajra

Ghosh

et al. 2011

Molecular and Cellular Biology

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The histone H3 variant Cse4 specifies centromere identity in Saccharomyces cerevisiae by its incorporation into a special nucleosome positioned at CEN DNA and promotes the assembly of the kinetochore complex, which is required for faithful chromosome segregation. Our previous work showed that Cse4 is also associated with the partitioning locus STB of the 2m circle-a multicopy plasmid that resides in the yeast nucleus and propagates itself stably. Cse4 is essential for the functional assembly of the plasmid partitioning complex, including the recruitment of the yeast cohesin complex at STB. We have located Cse4 association strictly at the origin-proximal subregion of STB. Three of the five directly repeated tandem copies of a 62-bp consensus sequence element constituting this region are necessary and sufficient for the recruitment of Cse4. The association of Cse4 with STB is dependent on Scm3, the loading factor responsible for the incorporation of Cse4 into the CEN nucleosome. A chromosomally integrated copy of STB confers on the integration site the capacity for Cse4 association as well as cohesin assembly. The localization of Cse4 in chromatin digested by micrococcal nuclease is consistent with the potential assembly of one Cse4-containing nucleosome, but not more than two, at STB. The remarkable ability of STB to acquire a very specialized, and strictly regulated, chromosome segregation factor suggests its plausible evolutionary kinship with CEN.The 2m plasmid of Saccharomyces cerevisiae is an example of a highly optimized, circular, multicopy extrachromosomal selfish DNA element (25,27,49). The plasmid does not seem to contribute to the host's fitness under standard laboratory growth conditions. However, at its steady-state copy number of 40 to 60 molecules per cell, any growth disadvantage imposed by the plasmid is rather small (16,30). The most remarkable attribute of this selectively almost neutral entity is its ability to propagate with nearly chromosome-like stability at its steadystate copy number. The entire genetic makeup of the 2m circle is devoted to three functions: efficient replication by the host machinery, equal segregation, and maintenance of the copy number. The plasmid accomplishes these goals with minimal metabolic encumbrance to its host.Direct visualization of fluorescence-tagged reporter plasmids in live cells suggests that 2m circle molecules are organized in the nucleus as 3 to 5 dynamic foci that form a closeknit cluster (47). The plasmid also segregates as a clustered entity; sister clusters part from each other and move away at the anaphase stage of the cell cycle. Population analysis and time lapse assays have revealed close similarities between the 2m circle and the yeast chromosomes or a centromere plasmid (minichromosome) in their dynamics and kinetics of segregation (17, 47). The relevant inference from a variety of experiments is that plasmid segregation is tightly coupled to chromosome segregation, perhaps by attachment of duplicated plasmid clusters to a pair of sister chro...

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“…Furthermore, plasmids similar in organization to the 2μm plasmid have been detected only among a limited number of fungal species belonging to this lineage [41]. These circumstantial pieces of evidence suggest the possibility that the loss of the capacity to establish a functional regional centromere was compensated for by the domestication of the plasmid partitioning locus as the central DNA element for directing chromosome segregation as well [26].…”

Section: Dna Topology Within the Cen And Stb Chromatinmentioning

confidence: 97%

Topological similarity between the 2μm plasmid partitioning locus and the budding yeast centromere: evidence for a common evolutionary origin?

Jayaram

Chang

Hui

et al. 2013

Biochemical Society Transactions

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The partitioning locus STB of the selfish plasmid, the 2μm circle, of Saccharomyces cerevisiae is essential for the propagation of this multi-copy extra-chromosomal DNA element with nearly chromosome-like stability. The functional competence of STB requires the plasmid-coded partitioning proteins Rep1 and Rep2 as well as host-coded proteins. Host factors that associate with STB in a Rep1- and Rep2-dependent manner also interact with centromeres, and play important roles in chromosome segregation. They include the cohesin complex and the centromere-specific histone H3 variant Cse4. The genetically defined point centromere of S. cerevisiae differs starkly from the much more widespread epigenetically specified regional centromeres of eukaryotes. The particularly small size of the S. cerevisiae centromere and the association of chromosome segregation factors with STB raise the possibility of an evolutionary link between these two partitioning loci. The unusual positive supercoiling harboured by the S. cerevisiae centromere and STB in vivo in their functional states, unveiled by recent experiments, bolsters the notion of their potential descent from an ancestral plasmid partitioning locus.

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A Circular Plasmid from the YeastTorulaspora delbrueckii

Cited by 31 publications

References 21 publications

Multiple new site-specific recombinases for use in manipulating animal genomes

Multiple new site-specific recombinases for use in manipulating animal genomes

Cse4 (CenH3) Association with the Saccharomyces cerevisiae Plasmid Partitioning Locus in Its Native and Chromosomally Integrated States: Implications in Centromere Evolution

Topological similarity between the 2μm plasmid partitioning locus and the budding yeast centromere: evidence for a common evolutionary origin?

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