The Autonomously Replicating Sequence (ARS) of the Yeast Saccharomyces exiguus Yp74L-3

In Saccharomyces cerevisiae chromosomal DNA replication initiates at intervals of ∼40 kb and depends upon the activity of autonomously replicating sequence (ARS) elements. The identification of ARS elements and analysis of their function as chromosomal replication origins requires the use of functional assays because they are not sufficiently similar to identify by DNA sequence analysis. To complete the systematic identification of ARS elements onS. cerevisiae chromosome III, overlapping clones covering 140 kb of the right arm were tested for their ability to promote extrachromosomal maintenance of plasmids. Examination of chromosomal replication intermediates of each of the seven ARS elements identified revealed that their efficiencies of use as chromosomal replication origins varied widely, with four ARS elements active in ≤10% of cells in the population and two ARS elements active in ≥90% of the population. Together with our previous analysis of a 200-kb region of chromosome III, these data provide the first complete analysis of ARS elements and DNA replication origins on an entire eukaryotic chromosome.

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Domain B of ARS307 contains two functional elements and contributes to chromosomal replication origin function

Molecular and Cellular Biology ◽

10.1128/mcb.14.11.7652-7659.1994 ◽

1994 ◽

Vol 14 (11) ◽

pp. 7652-7659

Author(s):

J F Theis ◽

C S Newlon

Keyword(s):

Replication Origin ◽

Consensus Sequence ◽

Functional Elements ◽

Chromosomal Replication ◽

Autonomously Replicating Sequence ◽

Chromosomal Origin ◽

Highly Active ◽

Origin Function ◽

Chromosome Iii ◽

Replication Activity

ARS307 is highly active as a replication origin in its native location on chromosome III of Saccharomyces cerevisiae. Its ability to confer autonomous replication activity on plasmids requires the presence of an 11-bp autonomously replicating sequence (ARS) consensus sequence (ACS), which is also required for chromosomal origin function, as well as approximately 100 bp of sequence flanking the ACS called domain B. To further define the sequences required for ARS function, a linker substitution mutagenesis of domain B was carried out. The mutations defined two sequences, B1 and B2, that contribute to ARS activity. Therefore, like ARS1, domain B of ARS307 is composed of functional subdomains. Constructs carrying mutations in the B1 element were used to replace the chromosomal copy of ARS307. These mutations caused a reduction in chromosomal origin activity, demonstrating that the B1 element is required for efficient chromosomal origin function.

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Functional equivalency and diversity of cis-acting elements among yeast replication origins.

Molecular and Cellular Biology ◽

10.1128/mcb.17.9.5473 ◽

1997 ◽

Vol 17 (9) ◽

pp. 5473-5484 ◽

Cited By ~ 38

Author(s):

S Lin ◽

D Kowalski

Keyword(s):

Origin Recognition Complex ◽

Common Mechanism ◽

Dna Unwinding ◽

Replication Origins ◽

Yeast Saccharomyces Cerevisiae ◽

Autonomously Replicating Sequence ◽

Cis Acting ◽

Dna Elements ◽

Functional Dna ◽

Dna Replication Origins

The DNA replication origins of the yeast Saccharomyces cerevisiae require several short functional elements, most of which are not conserved in sequence. To better characterize ARS305, a replicator from a chromosomal origin, we swapped functional DNA elements of ARS305 with defined elements of ARS1. ARS305 contains elements that are functionally exchangeable with ARS1 A and B1 elements, which are known to bind the origin recognition complex; however, the ARS1 A element differs in that it does not require a 3' box adjacent to the essential autonomously replicating sequence consensus. At the position corresponding to ARS1 B3, ARS305 has a novel element, B4, that can functionally substitute for every type of short element (B1, B2, and B3) in the B domain. Unexpectedly, the replacement of element B4 by ARS1 B3, which binds ABF1p and is known as a replication enhancer, inhibited ARS305 function. ARS305 has no short functional element at or near positions corresponding to the B2 elements in ARS1 and ARS307 but contains an easily unwound region whose functional importance was supported by a broad G+C-rich substitution mutation. Surprisingly, the easily unwound region can functionally substitute for the ARS1 B2 element, even though ARS1 B2 was found to possess a distinct DNA sequence requirement. The functionally conserved B2 element in ARS307 contains a known sequence requirement, and helical stability analysis of linker and minilinker mutations suggested that B2 also contains a DNA unwinding element (DUE). Our findings suggest that yeast replication origins employ a B2 element or a DUE to mediate a common function, DNA unwinding during initiation, although not necessarily through a common mechanism.

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Evaluación de levaduras en la producción de etanol a partir de melaza de caña de azúcar

Revista ESPAMCIENCIA ◽

10.51260/revista_espamciencia.v11i2.193 ◽

2020 ◽

Vol 11 (2) ◽

pp. 115-119

Author(s):

Miguel Alejandro Tuárez Párraga ◽

◽

Mabel Leonela Laz Mero ◽

Stephany Judith Bermello Ochoa ◽

◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Candida Utilis ◽

Microsoft Excel ◽

C 30 ◽

Saccharomyces Exiguus

El objetivo de la investigación consistió en evaluar el efecto de la adición de tres tipos de levaduras comerciales Saccharomyces cerevisiae, Saccharomyces exiguus y Candida utilis, utilizando disolución de melaza, las etapas del experimento consistieron en la activación de levadura y fermentación para la producción de etanol mediante una destilación simple; determinando la producción de etanol más óptima del experimento, con diferentes concentraciones de sacarosa (19°Bx, 23°Bx y 27°Bx) y diferentes condiciones ambientales (27°C, 30°C y 33°C). Se aplicó un modelo estadístico de diseño cuadrado latino utilizando el programa Microsoft Excel, tomando como factor de interés tipos de levaduras y factores secundarios los sólidos solubles (°Bx) y temperaturas de almacenamiento (°C). Se formularon nueve tratamientos (T), de los cuales se observaron diferencias significativas (p<0,05) entre los sólidos solubles (°Bx) y (p>0,05) para los tipos de levaduras estudiadas y las temperaturas de almacenamiento. Durante la producción de etanol a partir de la disolución de melaza de caña, el mejor tratamiento fue el T8, utilizando levadura Saccharomyces cerevisiae a 27°Bx y con temperatura de 30°C.

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A synthetic silencer mediates SIR-dependent functions in Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.11.11.5648-5659.1991 ◽

1991 ◽

Vol 11 (11) ◽

pp. 5648-5659

Author(s):

F J McNally ◽

J Rine

Keyword(s):

Saccharomyces Cerevisiae ◽

Binding Site ◽

Binding Sites ◽

Consensus Sequence ◽

Protein A ◽

Replication Initiation ◽

Yeast Saccharomyces Cerevisiae ◽

Autonomously Replicating Sequence ◽

Functional Components

Copies of the mating-type genes are present at three loci on chromosome III of the yeast Saccharomyces cerevisiae. The genes at the MAT locus are transcribed, whereas the identical genes at the silent loci, HML and HMR, are not transcribed. Several genes, including the four SIR genes, and two sites, HMR-E and HMR-I, are required for repression of transcription at the HMR locus. Three elements have been implicated in the function of the HMR-E silencer: a binding site for the RAP1 protein, a binding site for the ABF1 protein, and an 11-bp consensus sequence common to nearly all autonomously replicating sequence (ARS) elements (putative origins of DNA replication). RAP1 and ABF1 binding sites of different sequence than those found at HMR-E were joined with an 11-bp ARS consensus sequence to form a synthetic silencer. The synthetic silencer was able to repress transcription of the HMRa1 gene, confirming that binding sites for RAP1 and ABF1 and the 11-bp ARS consensus sequence were the functional components of the silencer in vivo. Mutations in the ABF1 binding site or in the ARS consensus sequence of the synthetic silencer caused nearly complete derepression of transcription at HMR. The ARS consensus sequence mutation also eliminated the ARS activity of the synthetic silencer. These data suggested that replication initiation at the HMR-E silencer was required for establishment of the repressed state at the HMR locus.

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Isolation and characterization of an autonomously replicating sequence from Ustilago maydis

Molecular and Cellular Biology ◽

10.1128/mcb.8.9.3703-3709.1988 ◽

1988 ◽

Vol 8 (9) ◽

pp. 3703-3709

Author(s):

T Tsukuda ◽

S Carleton ◽

S Fotheringham ◽

W K Holloman

Keyword(s):

Saccharomyces Cerevisiae ◽

Consensus Sequence ◽

Ustilago Maydis ◽

Small Fragment ◽

Direct Repeats ◽

Base Pairs ◽

Autonomously Replicating Sequence ◽

Isolation And Characterization ◽

Extrachromosomal Elements

DNA fragments that function as autonomously replicating sequences (ARSs) have been isolated from Ustilago maydis. When inserted into an integrative transforming vector, the fragments increased the frequency of U. maydis transformation several-thousandfold. ARS-containing plasmids were transmitted in U. maydis as extrachromosomal elements through replication. They were maintained at a level of about 25 copies per cell but were mitotically unstable. One ARS characterized in detail, which we called UARS1, was localized to a 1.7-kilobase fragment. UARS1 contained a cluster of active sequences. This element could be reduced further into three separate subfragments, each of which retained ARS activity. The smallest one was 383 base pairs (bp) long. Although not active itself in yeast, this small fragment contained seven 8-bp direct repeats, two contiguous 30-bp direct repeats, and five 11-bp units in both orientations with sequences similar but not identical to the consensus sequence found to be crucial for ARS activity in Saccharomyces cerevisiae.

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Interaction of the H4 autonomously replicating sequence core consensus sequence and its 3'-flanking domain

Molecular and Cellular Biology ◽

10.1128/mcb.9.12.5464-5472.1989 ◽

1989 ◽

Vol 9 (12) ◽

pp. 5464-5472

Author(s):

S G Holmes ◽

M M Smith

Keyword(s):

Consensus Sequence ◽

Point Mutations ◽

Relative Orientation ◽

Autonomously Replicating Sequence ◽

The Core ◽

Multiple Copies ◽

Minimal Sequences ◽

Necessary And Sufficient ◽

The Relationship

Yeast autonomously replicating sequence (ARS) elements are composed of a conserved 11-base-pair (bp) core consensus sequence and a less well defined 3'-flanking region. We have investigated the relationship between the H4 ARS core consensus sequence and its 3'-flanking domain. The minimal sequences necessary and sufficient for function were determined by combining external 3' and 5' deletions to produce a nested set of ARS fragments. Sequences 5' of the core consensus were dispensable for function, but at least 66 bp of 3'-flanking domain DNA was required for full ARS function. The importance of the relative orientation of the core consensus element with respect to the 3'-flanking domain was tested by precisely inverting 14 bp of DNA including the core consensus sequence by oligonucleotide mutagenesis. This core inversion mutant was defective for all ARS function, showing that a fixed relative orientation of the core consensus and 3'-flanking domain is required for function. The 3'-flanking domain of the minimal functional H4 ARS fragment contains three sequences with a 9-of-11-bp match to the core consensus. The role of these near-match sequences was tested by directed mutagenesis. When all near-match sequences with an 8-of-11-bp match or better were simultaneously disrupted by point mutations, the resulting ARS construct retained full replication function. Therefore, multiple copies of a sequence closely related to the core consensus element are not required for H4 ARS function.

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Organization of replication of ribosomal DNA in Saccharomyces cerevisiae.

Molecular and Cellular Biology ◽

10.1128/mcb.8.11.4927 ◽

1988 ◽

Vol 8 (11) ◽

pp. 4927-4935 ◽

Cited By ~ 197

Author(s):

M H Linskens ◽

J A Huberman

Keyword(s):

Saccharomyces Cerevisiae ◽

Ribosomal Dna ◽

Repeat Unit ◽

Replication Forks ◽

Sequence Element ◽

Autonomously Replicating Sequence ◽

Nontranscribed Spacer ◽

Precursor Rna ◽

Mapping Techniques ◽

Region Ii

Using recently developed replicon mapping techniques, we have analyzed the replication of the ribosomal DNA in Saccharomyces cerevisiae. The results show that (i) the functional origin of replication colocalizes with an autonomously replicating sequence element previously mapped to the nontranscribed spacer region, (ii) only a fraction of the potential origins are utilized in a single S phase, and (iii) the replication forks moving counter to the direction of transcription of the 37S precursor RNA stop at or near the termination site of transcription. Consequently, most ribosomal DNA is replicated unidirectionally by forks moving in the direction of transcription and most replicons are larger than the repeat unit. The significance of this finding for the replication of abundantly transcribed genes is discussed.

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Participation of the yeast activator Abf1 in meiosis-specific expression of the HOP1 gene.

Molecular and Cellular Biology ◽

10.1128/mcb.16.6.2777 ◽

1996 ◽

Vol 16 (6) ◽

pp. 2777-2786 ◽

Cited By ~ 24

Author(s):

V Gailus-Durner ◽

J Xie ◽

C Chintamaneni ◽

A K Vershon

Keyword(s):

Transcriptional Activation ◽

Mutational Analysis ◽

Consensus Sequence ◽

Promoter Sequence ◽

Regulatory Elements ◽

Specific Gene ◽

Specific Expression ◽

Autonomously Replicating Sequence

The meiosis-specific gene HOP1, which encodes a component of the synaptonemal complex, is controlled through two regulatory elements, UASH and URS1H. Sites similar to URS1H have been identified in the promoter region of virtually every early meiosis-specific gene, as well as in many promoters of nonmeiotic genes, and it has been shown that the proteins that bind to this site function to regulate meiotic and nonmeiotic transcription. Sites similar to the UASH site have been found in a number of meiotic and nonmeiotic genes as well. Since it has been shown that UASH functions as an activator site in vegetative haploid cells, it seemed likely that the factors binding to this site regulate both meiotic and nonmeiotic transcription. We purified the factor binding to the UASH element of the HOP1 promoter. Sequence analysis identified the protein as Abf1 (autonomously replicating sequence-binding factor 1), a multifunctional protein involved in DNA replication, silencing, and transcriptional regulation. We show by mutational analysis of the UASH site, that positions outside of the proposed UASH consensus sequence (TNTGN[A/T]GT) are required for DNA binding in vitro and transcriptional activation in vivo. A new UASH consensus sequence derived from this mutational analysis closely matches a consensus Abf1 binding site. We also show that an Abf1 site from a nonmeiotic gene can replace the function of the UASH site in the HOP1 promoter. Taken together, these results show that Abf1 functions to regulate meiotic gene expression.

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