scholarly journals Copy number and partition of the Saccharomyces cerevisiae 2 micron plasmid controlled by transcription regulators.

1988 ◽  
Vol 8 (11) ◽  
pp. 4949-4957 ◽  
Author(s):  
B E Veit ◽  
W L Fangman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Copy Number ◽  
Gene Products ◽  
Plasmid Copy ◽  
Cis Acting ◽  
Master Regulators ◽  
Transcription Regulators ◽  
2 Micron Plasmid ◽  
Mother Nucleus ◽  
Specific Sequences

The 2 micron plasmid of Saccharomyces cerevisiae is maintained by the action of plasmid-encoded gene products that control copy number and promote equipartition of plasmid copies at cell division. We show that the REP1 and REP2 plasmid-encoded gene products are master regulators that act in concert to autoregulate the level of their own transcripts and to regulate transcript levels of the FLP gene that promotes plasmid copy amplification. REP1 and REP2 are also shown to repress transcription at REP3, the cis-acting site essential for plasmid equipartitioning. We propose a model in which REP3 acts by dislodging transcription apparatuses that otherwise cause plasmid molecules to adhere to the mother nucleus and segregate asymmetrically. On the basis of their ability to generate specific chromatin structures, we also propose that the REP1 and REP2 gene products interact with different specific sequences found iterated in the 2 micron plasmid.

Copy number and partition of the Saccharomyces cerevisiae 2 micron plasmid controlled by transcription regulators

1988 ◽  
Vol 8 (11) ◽  
pp. 4949-4957
Author(s):  
B E Veit ◽  
W L Fangman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Copy Number ◽  
Gene Products ◽  
Plasmid Copy ◽  
Cis Acting ◽  
Master Regulators ◽  
Transcription Regulators ◽  
2 Micron Plasmid ◽  
Mother Nucleus ◽  
Specific Sequences

The 2 micron plasmid of Saccharomyces cerevisiae is maintained by the action of plasmid-encoded gene products that control copy number and promote equipartition of plasmid copies at cell division. We show that the REP1 and REP2 plasmid-encoded gene products are master regulators that act in concert to autoregulate the level of their own transcripts and to regulate transcript levels of the FLP gene that promotes plasmid copy amplification. REP1 and REP2 are also shown to repress transcription at REP3, the cis-acting site essential for plasmid equipartitioning. We propose a model in which REP3 acts by dislodging transcription apparatuses that otherwise cause plasmid molecules to adhere to the mother nucleus and segregate asymmetrically. On the basis of their ability to generate specific chromatin structures, we also propose that the REP1 and REP2 gene products interact with different specific sequences found iterated in the 2 micron plasmid.

scholarly journals Chromatin organization of the Saccharomyces cerevisiae 2 microns plasmid depends on plasmid-encoded products.

1985 ◽  
Vol 5 (9) ◽  
pp. 2190-2196 ◽  
Author(s):  
B E Veit ◽  
W L Fangman
Keyword(s):  
Copy Number ◽  
Plasmid Copy Number ◽  
Chromatin Organization ◽  
Open Reading Frames ◽  
Plasmid Copy ◽  
Cis Acting ◽  
Stable Plasmid ◽  
2 Micron Plasmid ◽  
Plasmid Genes ◽  
A Site

We have used gene disruptions and nuclease probes to assess the roles of yeast 2 micron plasmid genes in plasmid chromatin organization. The chromatin structure at the replication origin is not dependent on any of the four major open reading frames, A, B, C, or D. While stable plasmid maintenance is known to depend on a cis-acting locus STB and genes B and C, we find that only gene B influences STB chromatin. Other interactions between plasmid gene products and sequences may reflect gene regulation: the chromatin organization at the 5' end of gene A, which codes for a site-specific recombinase, depends on both gene B and gene C. Since disruption of gene C results in an increase in plasmid copy number that is dependent on gene A, we propose that gene C (and probably gene B) control copy number by regulating the level of the gene A recombinase.

scholarly journals Replication and segregation of plasmids containing cis-acting regulatory sites of silent mating-type genes in Saccharomyces cerevisiae are controlled by the SIR genes.

1987 ◽  
Vol 7 (12) ◽  
pp. 4225-4237 ◽  
Author(s):  
W J Kimmerly ◽  
J Rine
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mating Type ◽  
Copy Number ◽  
Consensus Sequence ◽  
Stability Characteristic ◽  
Mitotic Stability ◽  
Wild Type ◽  
Plasmid Copy ◽  
Cis Acting ◽  
Regulatory Sites

In Saccharomyces cerevisiae, two cis-acting regulatory sites called E and I flank the silent mating-type gene, HMRa, and mediate SIR-dependent transcriptional repression of the a1-a2 promoters. It has been shown previously that the E and I sites have plasmid replicator (ARS) activity. We show in this report that the ARS activity of the E and I sites is governed by the SIR genotype of the cell. In wild-type cells, a plasmid carrying the E site from HMRa (HMR E) in the vector YIp5 exhibited very high mitotic stability at a copy number of approximately 25 per cell. However, in sir2, sir3, or sir4 mutants, plasmids with HMR E had the low mitotic stability characteristic of plasmids containing ARS1, a SIR-independent replicator. Elevated mitotic stability of plasmids that carry HMR E is due to a segregation mechanism provided by SIR and HMR E. In sir2 and sir4 mutants, the plasmid copy number was significantly lowered, suggesting that these gene products also participate in the replication of plasmids carrying HMR E. The phenotype of point mutations introduced at an 11-base-pair ARS consensus sequence present at HMR E indicated that this sequence is functional but not absolutely required for autonomous replication of the plasmid and that it is not required for SIR-dependent mitotic stabilization. A plasmid carrying both a centromere and HMR E exhibited reduced mitotic stability in wild-type cells. This destabilization appeared to be due to antagonism between the segregation functions provided by the centromere and by HMR E.

Replication and segregation of plasmids containing cis-acting regulatory sites of silent mating-type genes in Saccharomyces cerevisiae are controlled by the SIR genes

1987 ◽  
Vol 7 (12) ◽  
pp. 4225-4237
Author(s):  
W J Kimmerly ◽  
J Rine
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mating Type ◽  
Copy Number ◽  
Consensus Sequence ◽  
Stability Characteristic ◽  
Mitotic Stability ◽  
Wild Type ◽  
Plasmid Copy ◽  
Cis Acting ◽  
Regulatory Sites

In Saccharomyces cerevisiae, two cis-acting regulatory sites called E and I flank the silent mating-type gene, HMRa, and mediate SIR-dependent transcriptional repression of the a1-a2 promoters. It has been shown previously that the E and I sites have plasmid replicator (ARS) activity. We show in this report that the ARS activity of the E and I sites is governed by the SIR genotype of the cell. In wild-type cells, a plasmid carrying the E site from HMRa (HMR E) in the vector YIp5 exhibited very high mitotic stability at a copy number of approximately 25 per cell. However, in sir2, sir3, or sir4 mutants, plasmids with HMR E had the low mitotic stability characteristic of plasmids containing ARS1, a SIR-independent replicator. Elevated mitotic stability of plasmids that carry HMR E is due to a segregation mechanism provided by SIR and HMR E. In sir2 and sir4 mutants, the plasmid copy number was significantly lowered, suggesting that these gene products also participate in the replication of plasmids carrying HMR E. The phenotype of point mutations introduced at an 11-base-pair ARS consensus sequence present at HMR E indicated that this sequence is functional but not absolutely required for autonomous replication of the plasmid and that it is not required for SIR-dependent mitotic stabilization. A plasmid carrying both a centromere and HMR E exhibited reduced mitotic stability in wild-type cells. This destabilization appeared to be due to antagonism between the segregation functions provided by the centromere and by HMR E.

Chromatin organization of the Saccharomyces cerevisiae 2 microns plasmid depends on plasmid-encoded products

1985 ◽  
Vol 5 (9) ◽  
pp. 2190-2196
Author(s):  
B E Veit ◽  
W L Fangman
Keyword(s):  
Copy Number ◽  
Plasmid Copy Number ◽  
Chromatin Organization ◽  
Open Reading Frames ◽  
Plasmid Copy ◽  
Cis Acting ◽  
Stable Plasmid ◽  
2 Micron Plasmid ◽  
Plasmid Genes ◽  
A Site

We have used gene disruptions and nuclease probes to assess the roles of yeast 2 micron plasmid genes in plasmid chromatin organization. The chromatin structure at the replication origin is not dependent on any of the four major open reading frames, A, B, C, or D. While stable plasmid maintenance is known to depend on a cis-acting locus STB and genes B and C, we find that only gene B influences STB chromatin. Other interactions between plasmid gene products and sequences may reflect gene regulation: the chromatin organization at the 5' end of gene A, which codes for a site-specific recombinase, depends on both gene B and gene C. Since disruption of gene C results in an increase in plasmid copy number that is dependent on gene A, we propose that gene C (and probably gene B) control copy number by regulating the level of the gene A recombinase.

The HML mating-type cassette of Saccharomyces cerevisiae is regulated by two separate but functionally equivalent silencers

1989 ◽  
Vol 9 (11) ◽  
pp. 4621-4630
Author(s):  
D J Mahoney ◽  
J R Broach
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mating Type ◽  
Binding Sites ◽  
The Other ◽  
Functional Domains ◽  
Gene Products ◽  
Cis Acting ◽  
Full Expression ◽  
Mating Type Genes ◽  
Chromosome Iii

Mating-type genes resident in the silent cassette HML at the left arm of chromosome III are repressed by the action of four SIR gene products, most likely mediated through two cis-acting sites located on opposite sides of the locus. We showed that deletion of either of these two cis-acting sites from the chromosome did not yield any detectable derepression of HML, while deletion of both sites yielded full expression of the locus. In addition, each of these sites was capable of exerting repression of heterologous genes inserted in their vicinity. Thus, HML expression is regulated by two independent silencers, each fully competent for maintaining repression. This situation was distinct from the organization of the other silent locus, HMR, at which a single silencer served as the predominant repressor of expression. Examination of identifiable domains and binding sites within the HML silencers suggested that silencing activity can be achieved by a variety of combinations of various functional domains.

scholarly journals Introduction of extra telomeric DNA sequences into Saccharomyces cerevisiae results in telomere elongation.

1989 ◽  
Vol 9 (4) ◽  
pp. 1488-1497 ◽  
Author(s):  
K W Runge ◽  
V A Zakian
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Sequences ◽  
Copy Number ◽  
Plasmid Copy Number ◽  
High Copy Number ◽  
Limiting Factor ◽  
Telomeric Dna ◽  
Base Pairs ◽  
Plasmid Copy ◽  
Telomere Elongation

The termini of Saccharomyces cerevisiae chromosomes consist of tracts of C1-3A (one to three cytosine and one adenine residue) sequences of approximately 450 base pairs in length. To gain insights into trans-acting factors at telomeres, high-copy-number linear and circular plasmids containing tracts of C1-3A sequences were introduced into S. cerevisiae. We devised a novel system to distinguish by color colonies that maintained the vector at 1 to 5, 20 to 50, and 100 to 400 copies per cell and used it to change the amount of telomeric DNA sequences per cell. An increase in the number of C1-3A sequences caused an increase in the length of telomeric C1-3A repeats that was proportional to plasmid copy number. Our data suggest that telomere growth is inhibited by a limiting factor(s) that specifically recognizes C1-3A sequences and that this factor can be effectively competed for by long tracts of C1-3A sequences at telomeres or on circular plasmids. Telomeres without this factor are exposed to processes that serve to lengthen chromosome ends.

Selection of yeast cells with a higher plasmid copy number in a Saccharomyces cerevisiae autoselection system

Yeast ◽  
1996 ◽  
Vol 12 (3) ◽  
pp. 199-205 ◽  
Author(s):  
Concetta Compagno ◽  
Danilo Porro ◽  
Stefania Radice ◽  
Enzo Martegani ◽  
Bianca Maria Ranzi
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Copy Number ◽  
Plasmid Copy Number ◽  
Yeast Cells ◽  
Plasmid Copy ◽  
Selection Of

scholarly journals Roles of the 2 microns gene products in stable maintenance of the 2 microns plasmid of Saccharomyces cerevisiae.

1987 ◽  
Vol 7 (10) ◽  
pp. 3566-3573 ◽  
Author(s):  
A E Reynolds ◽  
A W Murray ◽  
J W Szostak
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Copy Number ◽  
Gene Products ◽  
Site Specific ◽  
Cell Cycles ◽  
Daughter Cells ◽  
Repeat Sequences ◽  
Copy Numbers ◽  
Mother And Daughter ◽  
Stable Maintenance

We have examined the replication and segregation of the Saccharomyces cerevisiae 2 microns circle. The amplification of the plasmid at low copy numbers requires site-specific recombination between the 2 microns inverted repeat sequences catalyzed by the plasmid-encoded FLP gene. No other 2 microns gene products are required. The overexpression of FLP in a strain carrying endogenous 2 microns leads to uncontrolled plasmid replication, longer cell cycles, and cell death. Two different assays show that the level of Flp activity decreases with increasing 2 microns copy number. This regulation requires the products of the REP1 and REP2 genes. These gene products also act together to ensure that 2 microns molecules are randomly segregated between mother and daughter cells at cell division.

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