On the mechanism of premeiotic DNA synthesis in the yeast Saccharomyces cerevisiae

Cultures of the yeast Saccharomyces cerevisiae that are heterozygous for the mating type (MATa/MAT alpha) undergo synchronous meiosis and spore formation when starved for nitrogen and supplied with a nonfermentable carbon source such as acetate. Haploid and homozygous MAT alpha/MAT alpha and MATa/MATa diploid cells incubated under the same conditions fail to undergo meiosis and are asporogenous. It has not yet been firmly established that gene expression during sporulation is controlled at the level of transcript accumulation. To examine this question, we used cloned genes that encode a variety of "housekeeping" functions to probe Northern blots to assay the appearance of specific transcripts in both sporulating and asporogenous S. cerevisiae. In sporulating cells, each transcript showed a characteristic pattern of accumulation, reaching a maximum relative abundance at one of several different periods. In contrast, in both asporogenous haploid MATa and diploid MAT alpha/MAT alpha cells, all transcripts accumulated with similar kinetics. These results suggest a sporulation-specific pattern for transcript appearance. During these studies, high levels of several different transcripts were observed at unexpected times in sporulating cells. Histone (H)2A and (H)2B1 transcripts, although most abundant during premeiotic DNA synthesis, remained at one-third to one-half maximal levels after its end and were found in mature ascospores. Their appearance at this time is in sharp contrast to vegetative cells in which these histone transcripts are only found just before and during the period of DNA synthesis. Furthermore, transcripts from GAL10 and CDC10 genes, which are believed to be dispensable for sporulation, were much more abundant in sporulating cells than in asporogenous cells and vegetative cells grown on glucose or acetate. The presence of these transcripts did not appear to be due to a general activation of transcription because each accumulated with different kinetics. In addition, the transcript for at least one gene, HO, that is also dispensable for sporulation was not detected. The increased abundance of transcripts from some genes not required for sporulation leads us to propose that genes preferentially expressed during sporulation need not be essential for this differentiation.

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Interaction of replication factor Sld3 and histone acetyl transferase Esa1 alleviates gene silencing and promotes the activation of late and dormant replication origins

Genetics ◽

10.1093/genetics/iyaa001 ◽

2020 ◽

Vol 217 (1) ◽

pp. 1-11

Author(s):

Seiji Tanaka

Keyword(s):

Saccharomyces Cerevisiae ◽

Gene Silencing ◽

Dna Synthesis ◽

Histone Acetyltransferase ◽

Replication Origins ◽

Histone Acetyl Transferase ◽

Yeast Saccharomyces Cerevisiae ◽

Step Process ◽

Helicase Loading ◽

Rate Limiting

Abstract DNA replication in eukaryotes is a multi-step process that consists of three main reactions: helicase loading (licensing), helicase activation (firing), and nascent DNA synthesis (elongation). Although the contributions of some chromatin regulatory factors in the licensing and elongation reaction have been determined, their functions in the firing reaction remain elusive. In the budding yeast Saccharomyces cerevisiae, Sld3, Sld7, and Cdc45 (3–7–45) are rate-limiting in the firing reaction and simultaneous overexpression of 3–7–45 causes untimely activation of late and dormant replication origins. Here, we found that 3–7–45 overexpression not only activated dormant origins in the silenced locus, HMLα, but also exerted an anti-silencing effect at this locus. For these, interaction between Sld3 and Esa1, a conserved histone acetyltransferase, was responsible. Moreover, the Sld3–Esa1 interaction was required for the untimely activation of late origins. These results reveal the Sld3–Esa1 interaction as a novel level of regulation in the firing reaction.

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Saccharomyces cerevisiae exhibits a sporulation-specific temporal pattern of transcript accumulation

Molecular and Cellular Biology ◽

10.1128/mcb.5.4.751-761.1985 ◽

1985 ◽

Vol 5 (4) ◽

pp. 751-761

Author(s):

D B Kaback ◽

L R Feldberg

Keyword(s):

Saccharomyces Cerevisiae ◽

Dna Synthesis ◽

Specific Pattern ◽

Transcript Accumulation ◽

Yeast Saccharomyces Cerevisiae ◽

Vegetative Cells ◽

Activation Of Transcription ◽

Nonfermentable Carbon Source ◽

General Activation ◽

Diploid Cells

Cultures of the yeast Saccharomyces cerevisiae that are heterozygous for the mating type (MATa/MAT alpha) undergo synchronous meiosis and spore formation when starved for nitrogen and supplied with a nonfermentable carbon source such as acetate. Haploid and homozygous MAT alpha/MAT alpha and MATa/MATa diploid cells incubated under the same conditions fail to undergo meiosis and are asporogenous. It has not yet been firmly established that gene expression during sporulation is controlled at the level of transcript accumulation. To examine this question, we used cloned genes that encode a variety of "housekeeping" functions to probe Northern blots to assay the appearance of specific transcripts in both sporulating and asporogenous S. cerevisiae. In sporulating cells, each transcript showed a characteristic pattern of accumulation, reaching a maximum relative abundance at one of several different periods. In contrast, in both asporogenous haploid MATa and diploid MAT alpha/MAT alpha cells, all transcripts accumulated with similar kinetics. These results suggest a sporulation-specific pattern for transcript appearance. During these studies, high levels of several different transcripts were observed at unexpected times in sporulating cells. Histone (H)2A and (H)2B1 transcripts, although most abundant during premeiotic DNA synthesis, remained at one-third to one-half maximal levels after its end and were found in mature ascospores. Their appearance at this time is in sharp contrast to vegetative cells in which these histone transcripts are only found just before and during the period of DNA synthesis. Furthermore, transcripts from GAL10 and CDC10 genes, which are believed to be dispensable for sporulation, were much more abundant in sporulating cells than in asporogenous cells and vegetative cells grown on glucose or acetate. The presence of these transcripts did not appear to be due to a general activation of transcription because each accumulated with different kinetics. In addition, the transcript for at least one gene, HO, that is also dispensable for sporulation was not detected. The increased abundance of transcripts from some genes not required for sporulation leads us to propose that genes preferentially expressed during sporulation need not be essential for this differentiation.

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ANALYSIS OF MEIOSIS-DEFECTIVE MUTATIONS IN YEAST BY PHYSICAL MONITORING OF RECOMBINATION

Genetics ◽

10.1093/genetics/113.3.551 ◽

1986 ◽

Vol 113 (3) ◽

pp. 551-567

Author(s):

Rhona H Borts ◽

Michael Lichten ◽

James E Haber

Keyword(s):

Saccharomyces Cerevisiae ◽

Dna Synthesis ◽

Vegetative Growth ◽

Growth Conditions ◽

Intragenic Recombination ◽

Dna Molecules ◽

Wild Type ◽

Yeast Saccharomyces Cerevisiae ◽

Restriction Fragments ◽

High Level

ABSTRACT We have developed a method by which the extent of physical exchange of DNA molecules can be determined throughout meiosis in the yeast Saccharomyces cerevisiae. We have used this technique to analyze the effect of five meiosis-defective mutations (rad6, rad50, rad52, rad57 and spo11) on the physical exchange of DNA molecules. In the same experiments, we have also measured other meiotic parameters, such as premeiotic DNA synthesis, commitment to intragenic recombination, haploidization, ascus formation, and viability. rad50 and spo11 diploids make an undetectable amount of physically recombined DNA and <1% of wild-type levels of viable intragenic recombinants. In contrast, diploids homozygous for rad52, rad6 or rad57 all yield significant amounts of novel restriction fragments which arise by recombination. rad57 diploids make nearly wild-type levels of the recombined restriction fragments, although they produce <10% of the wild-type levels of viable intragenic recombinants. rad52 strains are also capable of a significant (33%) amount of exchange of DNA molecules, but make <1% of wild-type levels of viable intragenic recombinants. rad6 diploids are also capable of undergoing a high level of exchange, as measured by the appearance of the recombined restriction fragment. In addition, rad6 diploids show an unusual allele- or locus-specific variability in the level of viable intragenic recombinants produced. Although rad6 diploids produce no viable spores, they are able to complete a significant amount of haploidization upon return to vegetative growth conditions.

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The Prevention of Repeat-Associated Deletions in Saccharomyces cerevisiae by Mismatch Repair Depends on Size and Origin of Deletions

Genetics ◽

10.1093/genetics/143.4.1579 ◽

1996 ◽

Vol 143 (4) ◽

pp. 1579-1587 ◽

Cited By ~ 6

Author(s):

Hiep T Tran ◽

Dmitry A Gordenin ◽

Michael A Resnick

Keyword(s):

Saccharomyces Cerevisiae ◽

Dna Synthesis ◽

Mismatch Repair ◽

Dna Polymerase ◽

Recombinational Repair ◽

Null Mutant ◽

Yeast Saccharomyces Cerevisiae ◽

Dna Polymerase Δ ◽

Polymerase Slippage

Abstract We have investigated the effects of mismatch repair on 1- to 61-bp deletions in the yeast Saccharomyces cerevisiae. The deletions are likely to involve unpaired loop intermediates resulting from DNA polymerase slippage. The mutator effects of mutations in the DNA polymerase δ (POL3) gene and the recombinational repair RAD52 gene were studied in combination with mismatch repair defects. The pol3-t mutation increased up to 1000-fold the rate of extended (7-61 bp) but not of 1-bp deletions. In a rad52 null mutant only the 1-bp deletions were increased (12-fold). The mismatch repair mutations pmsl, msh2 and msh3 did not affect 31- and 61-bp deletions in the pol3-t but increased the rates of 7- and 1-bp deletions. We propose that loops less than or equal to seven bases generated during replication are subject to mismatch repair by the PMSI, MSH2, MSH3 system and that it cannot act on loops ≤31 bases. In contrast to the pol3-t, the enhancement of 1-bp deletions in a rad52 mutant is not altered by a pmsl mutation. Thus, mismatch repair appears to be specific to errors of DNA synthesis generated during semiconservative replication.

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