scholarly journals Alterations in the adenine-plus-thymine-rich region of CEN3 affect centromere function in Saccharomyces cerevisiae.

1987 ◽  
Vol 7 (1) ◽  
pp. 68-75 ◽  
Author(s):  
A Gaudet ◽  
M Fitzgerald-Hayes
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Base Composition ◽  
Consensus Sequence ◽  
Fold Increase ◽  
Base Pairs ◽  
Yeast Saccharomyces Cerevisiae ◽  
Core Element ◽  
Sequence Elements ◽  
Dna Elements ◽  
Chromosome Iii

Centromere DNA from 11 of the 16 chromosomes of the yeast Saccharomyces cerevisiae have been analyzed and reveal three sequence elements common to each centromere, referred to as conserved centromere DNA elements (CDE). The adenine-plus-thymine (A + T)-rich central core element, CDE II, is flanked by two short conserved sequences, CDE I (8 base pairs [bp]) and CDE III (25 bp). Although no consensus sequence exists among the different CDE II regions, they do have three common features of sequence organization. First, the CDE II regions are similar in length, ranging from 78 to 86 bp measured from CDE I to the left boundary of CDE III. Second, the base composition is always greater than 90% A + T. Finally, the A and T residues in these segments are often arranged in runs of A and runs of T residues, sometimes with six or seven bases in a stretch. We constructed insertion, deletion, and replacement mutations in the CDE II region of the centromere from chromosome III, CEN3, designed to investigate the length and sequence requirements for function of the CDE II region of the centromere. We analyzed the effect of these altered centromeres on plasmid and chromosome segregation in S. cerevisiae. Our results show that increasing the length of CDE II from 84 to 154 bp causes a 100-fold increase in chromosome nondisjunction. Deletion mutations removing segments of the A + T-rich CDE II DNA also cause aberrant segregation. In some cases partial function could be restored by replacing the deleted DNA with fragments whose primary sequence or base composition is very different from that of the wild-type CDE II DNA. In addition, we found that identical mutations introduced into different positions in CDE II have very similar effects.

Alterations in the adenine-plus-thymine-rich region of CEN3 affect centromere function in Saccharomyces cerevisiae

1987 ◽  
Vol 7 (1) ◽  
pp. 68-75
Author(s):  
A Gaudet ◽  
M Fitzgerald-Hayes
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Base Composition ◽  
Consensus Sequence ◽  
Fold Increase ◽  
Base Pairs ◽  
Yeast Saccharomyces Cerevisiae ◽  
Core Element ◽  
Sequence Elements ◽  
Dna Elements ◽  
Chromosome Iii

Centromere DNA from 11 of the 16 chromosomes of the yeast Saccharomyces cerevisiae have been analyzed and reveal three sequence elements common to each centromere, referred to as conserved centromere DNA elements (CDE). The adenine-plus-thymine (A + T)-rich central core element, CDE II, is flanked by two short conserved sequences, CDE I (8 base pairs [bp]) and CDE III (25 bp). Although no consensus sequence exists among the different CDE II regions, they do have three common features of sequence organization. First, the CDE II regions are similar in length, ranging from 78 to 86 bp measured from CDE I to the left boundary of CDE III. Second, the base composition is always greater than 90% A + T. Finally, the A and T residues in these segments are often arranged in runs of A and runs of T residues, sometimes with six or seven bases in a stretch. We constructed insertion, deletion, and replacement mutations in the CDE II region of the centromere from chromosome III, CEN3, designed to investigate the length and sequence requirements for function of the CDE II region of the centromere. We analyzed the effect of these altered centromeres on plasmid and chromosome segregation in S. cerevisiae. Our results show that increasing the length of CDE II from 84 to 154 bp causes a 100-fold increase in chromosome nondisjunction. Deletion mutations removing segments of the A + T-rich CDE II DNA also cause aberrant segregation. In some cases partial function could be restored by replacing the deleted DNA with fragments whose primary sequence or base composition is very different from that of the wild-type CDE II DNA. In addition, we found that identical mutations introduced into different positions in CDE II have very similar effects.

scholarly journals Nonrecombinant meiosis I nondisjunction in Saccharomyces cerevisiae induced by tRNA ochre suppressors.

Genetics ◽  
1989 ◽  
Vol 123 (1) ◽  
pp. 81-95 ◽  
Author(s):  
E J Louis ◽  
J E Haber
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Chromosome ◽  
Stop Codon ◽  
Fold Increase ◽  
Wild Type ◽  
Yeast Saccharomyces Cerevisiae ◽  
Nonsense Mutations ◽  
Chromosome Iii ◽  
Meiosis I ◽  
Chromosome Nondisjunction

Abstract The presence of the tRNA ochre suppressors SUP11 and SUP5 is found to induce meiosis I nondisjunction in the yeast Saccharomyces cerevisiae. The induction increases with increasing dosage of the suppressor and decreases in the presence of an antisuppressor. The effect is independent of the chromosomal location of SUP11. Each of five different chromosomes monitored exhibited nondisjunction at frequencies of 0.1%-1.1% of random spores, which is a 16-160-fold increase over wild-type levels. Increased nondisjunction is reflected by a marked increase in tetrads with two and zero viable spores. In the case of chromosome III, for which a 50-cM map interval was monitored, the resulting disomes are all in the parental nonrecombinant configuration. Recombination along chromosome III appears normal both in meioses that have no nondisjunction and in meioses for which there was nondisjunction of another chromosome. We propose that a proportion of one or more proteins involved in chromosome pairing, recombination or segregation are aberrant due to translational read-through of the normal ochre stop codon. Hygromycin B, an antibiotic that can suppress nonsense mutations via translational read-through, also induces nonrecombinant meiosis I nondisjunction. Increases in mistranslation, therefore, increase the production of aneuploids during meiosis. There was no observable effect of SUP11 on mitotic chromosome nondisjunction; however some disomes caused SUP11 ade2-ochre strains to appear white or red, instead of pink.

The Saccharomyces cerevisiae chromosome III left telomere has a type X, but not a type Y', ARS region

1986 ◽  
Vol 6 (4) ◽  
pp. 1352-1356
Author(s):  
L L Button ◽  
C R Astell
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Southern Hybridization ◽  
Repeat Sequence ◽  
Telomeric Region ◽  
Chromosome Walking ◽  
Base Pairs ◽  
Yeast Saccharomyces Cerevisiae ◽  
Simple Tandem Repeat ◽  
Sali Fragment ◽  
Chromosome Iii

A yeast Saccharomyces cerevisiae telomeric region was isolated by chromosome walking from HML alpha, the most distal known gene on the chromosome III left (IIIL) end. The terminal heterodisperse 3.3-kilobase (kb) SalI fragment on chromosome IIIL, 8.6 kb distal to HML alpha, was cloned in a circular vector to generate a telomeric probe. Southern hybridization and DNA sequencing analyses indicated that 0.6 kb (+/- 200 base pairs) of 5'-C1-3A-3' simple tandem repeat sequence, adjacent to a 1.2-kb type X ARS region, constitutes the telomere on the chromosome IIIL end, and no type Y' ARS region homologies exist between HML alpha and the IIIL terminus.

scholarly journals Nucleosome depletion alters the chromatin structure of Saccharomyces cerevisiae centromeres.

1990 ◽  
Vol 10 (11) ◽  
pp. 5721-5727 ◽  
Author(s):  
M J Saunders ◽  
E Yeh ◽  
M Grunstein ◽  
K Bloom
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Chromosome Segregation ◽  
Mutational Analysis ◽  
Base Pairs ◽  
Centromere Function ◽  
Nuclease Digestion ◽  
Dna Elements ◽  
Chromosome Iii ◽  
Sequence Requirements

Saccharomyces cerevisiae centromeric DNA is packaged into a highly nuclease-resistant chromatin core of approximately 200 base pairs of DNA. The structure of the centromere in chromosome III is somewhat larger than a 160-base-pair nucleosomal core and encompasses the conserved centromere DNA elements (CDE I, II, and III). Extensive mutational analysis has revealed the sequence requirements for centromere function. Mutations affecting the segregation properties of centromeres also exhibit altered chromatin structures in vivo. Thus the structure, as delineated by nuclease digestion, correlated with functional centromeres. We have determined the contribution of histone proteins to this unique structural organization. Nucleosome depletion by repression of either histone H2B or H4 rendered the cell incapable of chromosome segregation. Histone repression resulted in increased nuclease sensitivity of centromere DNA, with up to 40% of CEN3 DNA molecules becoming accessible to nucleolytic attack. Nucleosome depletion also resulted in an alteration in the distribution of nuclease cutting sites in the DNA surrounding CEN3. These data provide the first indication that authentic nucleosomal subunits flank the centromere and suggest that nucleosomes may be the central core of the centromere itself.

scholarly journals The Saccharomyces cerevisiae chromosome III left telomere has a type X, but not a type Y', ARS region.

1986 ◽  
Vol 6 (4) ◽  
pp. 1352-1356 ◽  
Author(s):  
L L Button ◽  
C R Astell
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Southern Hybridization ◽  
Repeat Sequence ◽  
Telomeric Region ◽  
Chromosome Walking ◽  
Base Pairs ◽  
Yeast Saccharomyces Cerevisiae ◽  
Simple Tandem Repeat ◽  
Sali Fragment ◽  
Chromosome Iii

A yeast Saccharomyces cerevisiae telomeric region was isolated by chromosome walking from HML alpha, the most distal known gene on the chromosome III left (IIIL) end. The terminal heterodisperse 3.3-kilobase (kb) SalI fragment on chromosome IIIL, 8.6 kb distal to HML alpha, was cloned in a circular vector to generate a telomeric probe. Southern hybridization and DNA sequencing analyses indicated that 0.6 kb (+/- 200 base pairs) of 5'-C1-3A-3' simple tandem repeat sequence, adjacent to a 1.2-kb type X ARS region, constitutes the telomere on the chromosome IIIL end, and no type Y' ARS region homologies exist between HML alpha and the IIIL terminus.

scholarly journals Nucleosome depletion alters the chromatin structure of Saccharomyces cerevisiae centromeres

1990 ◽  
Vol 10 (11) ◽  
pp. 5721-5727
Author(s):  
M J Saunders ◽  
E Yeh ◽  
M Grunstein ◽  
K Bloom
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Chromosome Segregation ◽  
Mutational Analysis ◽  
Base Pairs ◽  
Centromere Function ◽  
Nuclease Digestion ◽  
Dna Elements ◽  
Chromosome Iii ◽  
Sequence Requirements

Saccharomyces cerevisiae centromeric DNA is packaged into a highly nuclease-resistant chromatin core of approximately 200 base pairs of DNA. The structure of the centromere in chromosome III is somewhat larger than a 160-base-pair nucleosomal core and encompasses the conserved centromere DNA elements (CDE I, II, and III). Extensive mutational analysis has revealed the sequence requirements for centromere function. Mutations affecting the segregation properties of centromeres also exhibit altered chromatin structures in vivo. Thus the structure, as delineated by nuclease digestion, correlated with functional centromeres. We have determined the contribution of histone proteins to this unique structural organization. Nucleosome depletion by repression of either histone H2B or H4 rendered the cell incapable of chromosome segregation. Histone repression resulted in increased nuclease sensitivity of centromere DNA, with up to 40% of CEN3 DNA molecules becoming accessible to nucleolytic attack. Nucleosome depletion also resulted in an alteration in the distribution of nuclease cutting sites in the DNA surrounding CEN3. These data provide the first indication that authentic nucleosomal subunits flank the centromere and suggest that nucleosomes may be the central core of the centromere itself.

Expansions and contractions of the genetic map relative to the physical map of yeast chromosome III

1988 ◽  
Vol 8 (2) ◽  
pp. 595-604
Author(s):  
L S Symington ◽  
T D Petes
Keyword(s):  
Gene Conversion ◽  
Restriction Site ◽  
Physical Map ◽  
Minimum Length ◽  
Base Pairs ◽  
Yeast Saccharomyces Cerevisiae ◽  
Yeast Chromosome ◽  
Chromosome Maps ◽  
Chromosome Iii ◽  
The Relationship

To examine the relationship between genetic and physical chromosome maps, we constructed a diploid strain of the yeast Saccharomyces cerevisiae heterozygous for 12 restriction site mutations within a 23-kilobase (5-centimorgan) interval of chromosome III. Crossovers were not uniformly distributed along the chromosome, one interval containing significantly more and one interval significantly fewer crossovers than expected. One-third of these crossovers occurred within 6 kilobases of the centromere. Approximately half of the exchanges were associated with gene conversion events. The minimum length of gene conversion tracts varied from 4 base pairs to more than 12 kilobases, and these tracts were nonuniformly distributed along the chromosome. We conclude that the chromosomal sequence or structure has a dramatic effect on meiotic recombination.

scholarly journals DNA SEQUENCES OF FRAMESHIFT AND OTHER MUTATIONS INDUCED BY ICR-170 IN YEAST

Genetics ◽  
1985 ◽  
Vol 111 (2) ◽  
pp. 233-241
Author(s):  
Joachim F Ernst ◽  
D Michael Hampsey ◽  
Fred Sherman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Sequence Analysis ◽  
Genetic Analysis ◽  
Dna Sequences ◽  
Induced Mutations ◽  
Sequence Analyses ◽  
Base Pairs ◽  
Yeast Saccharomyces Cerevisiae ◽  
Frameshift Mutations

ABSTRACT ICR-170-induced mutations in the CYC1 gene of the yeast Saccharomyces cerevisiae were investigated by genetic and DNA sequence analyses. Genetic analysis of 33 cyc1 mutations induced by ICR-170 and sequence analysis of eight representatives demonstrated that over one-third were frameshift mutations that occurred at one site corresponding to amino acid positions 29-30, whereas the remaining mutations were distributed more-or-less randomly, and a few of these were not frameshift mutations. The sequence results indicate that ICR-170 primarily induces G·C additions at sites containing monotonous runs of three G·C base pairs. However, some (see PDF) sites within the CYC1 gene were not mutated by ICR-170. Thus, ICR-170 is a relatively specific mutagen that preferentially acts on certain sites with monotonous runs of G·C base pairs.

A synthetic silencer mediates SIR-dependent functions in Saccharomyces cerevisiae

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.

Isolation and characterization of an autonomously replicating sequence from Ustilago maydis

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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