Physical monitoring of mating type switching in Saccharomyces cerevisiae

1988 ◽  
Vol 8 (6) ◽  
pp. 2342-2349 ◽  
Author(s):  
B Connolly ◽  
C I White ◽  
J E Haber
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Mating Type ◽  
G1 Phase ◽  
Opposite Mating Type ◽  
Synthesis Inhibitor ◽  
Mating Type Switching ◽  
G2 Phase ◽  
Kinetics Of ◽  
Ho Gene

The kinetics of mating type switching in Saccharomyces cerevisiae can be followed at the DNA level by using a galactose-inducible HO (GAL-HO) gene to initiate the event in synchronously growing cells. From the time that HO endonuclease cleaves MAT a until the detection of MAT alpha DNA took 60 min. When unbudded G1-phase cells were induced, switched to the opposite mating type in "pairs." In the presence of the DNA synthesis inhibitor hydroxyurea, HO-induced cleavage occurred but cells failed to complete switching. In these blocked cells, the HO-cut ends of MATa remained stable for at least 3 h. Upon removal of hydroxyurea, the cells completed the switch in approximately 1 h. The same kinetics of MAT switching were also seen in asynchronous cultures and when synchronously growing cells were induced at different times of the cell cycle. Thus, the only restriction that confined normal homothallic switching to the G1 phase of the cell cycle was the expression of HO endonuclease. Further evidence that galactose-induced cells can switch in the G2 phase of the cell cycle was the observation that these cells did not always switch in pairs. This suggests that two chromatids, both cleaved with HO endonuclease, can interact independently with the donors HML alpha and HMRa.

scholarly journals Physical monitoring of mating type switching in Saccharomyces cerevisiae.

1988 ◽  
Vol 8 (6) ◽  
pp. 2342-2349 ◽  
Author(s):  
B Connolly ◽  
C I White ◽  
J E Haber
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Mating Type ◽  
G1 Phase ◽  
Opposite Mating Type ◽  
Synthesis Inhibitor ◽  
Mating Type Switching ◽  
G2 Phase ◽  
Kinetics Of ◽  
Ho Gene

The kinetics of mating type switching in Saccharomyces cerevisiae can be followed at the DNA level by using a galactose-inducible HO (GAL-HO) gene to initiate the event in synchronously growing cells. From the time that HO endonuclease cleaves MAT a until the detection of MAT alpha DNA took 60 min. When unbudded G1-phase cells were induced, switched to the opposite mating type in "pairs." In the presence of the DNA synthesis inhibitor hydroxyurea, HO-induced cleavage occurred but cells failed to complete switching. In these blocked cells, the HO-cut ends of MATa remained stable for at least 3 h. Upon removal of hydroxyurea, the cells completed the switch in approximately 1 h. The same kinetics of MAT switching were also seen in asynchronous cultures and when synchronously growing cells were induced at different times of the cell cycle. Thus, the only restriction that confined normal homothallic switching to the G1 phase of the cell cycle was the expression of HO endonuclease. Further evidence that galactose-induced cells can switch in the G2 phase of the cell cycle was the observation that these cells did not always switch in pairs. This suggests that two chromatids, both cleaved with HO endonuclease, can interact independently with the donors HML alpha and HMRa.

scholarly journals DAF1, a mutant gene affecting size control, pheromone arrest, and cell cycle kinetics of Saccharomyces cerevisiae.

1988 ◽  
Vol 8 (11) ◽  
pp. 4675-4684 ◽  
Author(s):  
F R Cross
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Critical Size ◽  
Size Control ◽  
Mutant Gene ◽  
G1 Phase ◽  
Wild Type ◽  
Alpha Factor ◽  
Multiple Copies ◽  
Kinetics Of

The mating pheromone alpha-factor arrests Saccharomyces cerevisiae MATa cells in the G1 phase of the cell cycle. Size control is also exerted in G1, since cells do not exit G1 until they have attained a critical size. A dominant mutation (DAF1-1) which causes both alpha-factor resistance and small cell size (volume about 0.6-fold that of the wild type) has been isolated and characterized genetically and by molecular cloning. Several alpha-factor-induced mRNAs were induced equivalently in daf1+ and DAF1-1 cells. The DAF1-1 mutation consisted of a termination codon two-thirds of the way through the daf1+ coding sequence. A chromosomal deletion of DAF1 produced by gene transplacement increased cell volume about 1.5-fold; thus, DAF1-1 may be a hyperactive or deregulated allele of a nonessential gene involved in G1 size control. Multiple copies of DAF1-1 also greatly reduced the duration of the G1 phase of the cell cycle.

DAF1, a mutant gene affecting size control, pheromone arrest, and cell cycle kinetics of Saccharomyces cerevisiae

1988 ◽  
Vol 8 (11) ◽  
pp. 4675-4684
Author(s):  
F R Cross
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Critical Size ◽  
Size Control ◽  
Mutant Gene ◽  
G1 Phase ◽  
Wild Type ◽  
Alpha Factor ◽  
Multiple Copies ◽  
Kinetics Of

The mating pheromone alpha-factor arrests Saccharomyces cerevisiae MATa cells in the G1 phase of the cell cycle. Size control is also exerted in G1, since cells do not exit G1 until they have attained a critical size. A dominant mutation (DAF1-1) which causes both alpha-factor resistance and small cell size (volume about 0.6-fold that of the wild type) has been isolated and characterized genetically and by molecular cloning. Several alpha-factor-induced mRNAs were induced equivalently in daf1+ and DAF1-1 cells. The DAF1-1 mutation consisted of a termination codon two-thirds of the way through the daf1+ coding sequence. A chromosomal deletion of DAF1 produced by gene transplacement increased cell volume about 1.5-fold; thus, DAF1-1 may be a hyperactive or deregulated allele of a nonessential gene involved in G1 size control. Multiple copies of DAF1-1 also greatly reduced the duration of the G1 phase of the cell cycle.

A CIS-ACTING MUTATION WITHIN THE MAT  a LOCUS OF SACCHAROMYCES CEREVISIAE THAT PREVENTS EFFICIENT HOMOTHALLIC MATING-TYPE SWITCHING

Genetics ◽  
1980 ◽  
Vol 94 (2) ◽  
pp. 341-360
Author(s):  
Deborah Wygal Mascioli ◽  
James E Haber
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mating Type ◽  
Pleiotropic Effect ◽  
Wild Type ◽  
Dominant Mutation ◽  
Cis Acting ◽  
Silent Genes ◽  
Factor Expression ◽  
Mating Type Switching ◽  
Ho Gene

ABSTRACT Homothallic strains of Saccharomyces cerevisiae are able to switch from one mating-type to the other as frequently as every cell division. We have identified a cis-dominant mutation of the MATa locus, designated MATa-inc, that can be converted to MATα at only about 5% of the normal efficiency. In homothallic MATa-inc/mata* diploids, the MATa-inc locus switched to MATα in only one of 30 cases, while the mata* locus switched to MATα in all 30 cases. The MATa-inc mutation can be "healed" by a series of switches, first to MATα and then to a normal allele of MATa. These data are consistent with the "cassette" model of HICKS, STRATHERN and HERSKOWITZ (1977), in which mating conversions involve the transposition of wild-type copies of a or α information from silent genes elsewhere in the genome. The MATa-inc mutation appears to alter a DNA sequence necessary for the replacement of MATa by MATα. The MATa-inc mutation has no other effect on MATa functions. In heterothallic backgrounds, the mutation has no effect on the sensitivity to α-factor, synthesis of a-factor, expression of barrier phenotype or ability to mate or sporulate.—The MATa-inc allele does, however, exhibit one pleiotropic effect. About 1% of homothallic MATa-inc cells become completely unable to switch mating type because ofmutations at HMa, the locus proposed to carry the silent copy of α information.—In addition, we have isolated a less efficient allele of the HO gene.

scholarly journals The Ubp15 deubiquitinase promotes timely entry into S phase in Saccharomyces cerevisiae

2015 ◽  
Vol 26 (12) ◽  
pp. 2205-2216 ◽  
Author(s):  
Denis Ostapenko ◽  
Janet L. Burton ◽  
Mark J. Solomon
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Physiological Role ◽  
S Phase ◽  
G1 Phase ◽  
Deubiquitinating Enzyme ◽  
Anaphase Promoting Complex ◽  
Synthesis Inhibitor ◽  
Yeast Saccharomyces Cerevisiae ◽  
Increased Sensitivity

The anaphase-promoting complex in partnership with its activator, Cdh1, is an E3 ubiquitin ligase responsible for targeting cell cycle proteins during G1 phase. In the budding yeast Saccharomyces cerevisiae, Cdh1 associates with the deubiquitinating enzyme Ubp15, but the significance of this interaction is unclear. To better understand the physiological role(s) of Ubp15, we examined cell cycle phenotypes of cells lacking Ubp15. We found that ubp15∆ cells exhibited delayed progression from G1 into S phase and increased sensitivity to the DNA synthesis inhibitor hydroxyurea. Both phenotypes of ubp15∆ cells were rescued by additional copies of the S-phase cyclin gene CLB5. Clb5 is an unstable protein targeted for proteasome-mediated degradation by several pathways. We found that during G1 phase, the APCCdh1-mediated degradation of Clb5 was accelerated in ubp15∆ cells. Ubp15 interacted with Clb5 independent of Cdh1 and deubiquitinated Clb5 in a reconstituted system. Thus deubiquitination by Ubp15 counteracts APC activity toward cyclin Clb5 to allow Clb5 accumulation and a timely entry into S phase.

scholarly journals INTERCONVERSION OF YEAST MATING TYPES III. ACTION OF THE HOMOTHALLISM (HO) GENE IN CELLS HOMOZYGOUS FOR THE MATING TYPE LOCUS

Genetics ◽  
1977 ◽  
Vol 85 (3) ◽  
pp. 395-405 ◽  
Author(s):  
James B Hicks ◽  
Jeffrey N Strathern ◽  
Ira Herskowitz
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Life Cycle ◽  
Gene Product ◽  
Direct Observation ◽  
Mating Type ◽  
Mating Types ◽  
Type Locus ◽  
Mating Type Locus ◽  
Mating Type Switching ◽  
Ho Gene

ABSTRACT Mating type interconversion in homothallic Saccharomyces cerevisiae has been studied in diploids homozygous for the mating type locus produced by sporulation of a/a/a/α and a/a/α/α tetraploid strains. Mating type switches have been analyzed by techniques including direct observation of cells for changes in α-factor sensitivity. Another method of following mating type switching exploits the observation that a/α cells exhibit polar budding and a/a and α/α cells exhibit medial budding.—These studies indicate the following: (1) The allele conferring the homothallic life cycle (HO) is dominant to the allele conferring the heterothallic life cycle (ho). (2) The action of the HO gene is controlled by the mating type locus—active in a/a and α/α cells but not in a/α cells. (3) The HO (or HO-controlled) gene product can act independently on two mating type alleles located on separate chromosomes in the same nucleus. (4) A switch in mating type is observed in pairs of cells, each of which has the same change.

Yeast dom34 Mutants Are Defective in Multiple Developmental Pathways and Exhibit Decreased Levels of Polyribosomes

Genetics ◽  
1998 ◽  
Vol 149 (1) ◽  
pp. 45-56
Author(s):  
Luther Davis ◽  
JoAnne Engebrecht
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Heterologous Expression ◽  
Protein Translation ◽  
Developmental Pathways ◽  
G1 Phase ◽  
Growth Defect ◽  
Wild Type ◽  
Drosophila Homolog ◽  
Mutant Cells

Abstract The DOM34 gene of Saccharomyces cerevisiae is similar togenes found in diverse eukaryotes and archaebacteria. Analysis of dom34 strains shows that progression through the G1 phase of the cell cycle is delayed, mutant cells enter meiosis aberrantly, and their ability to form pseudohyphae is significantly diminished. RPS30A, which encodes ribosomal protein S30, was identified in a screen for high-copy suppressors of the dom34Δ growth defect. dom34Δ mutants display an altered polyribosome profile that is rescued by expression of RPS30A. Taken together, these data indicate that Dom34p functions in protein translation to promote G1 progression and differentiation. A Drosophila homolog of Dom34p, pelota, is required for the proper coordination of meiosis and spermatogenesis. Heterologous expression of pelota in dom34Δ mutants restores wild-type growth and differentiation, suggesting conservation of function between the eukaryotic members of the gene family.

Homothallic mating type switching generates lethal chromosome breaks in rad52 strains of Saccharomyces cerevisiae

1981 ◽  
Vol 1 (6) ◽  
pp. 522-534
Author(s):  
B Weiffenbach ◽  
J E Haber
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mating Type ◽  
Deoxyribonucleic Acid ◽  
Type Locus ◽  
Mating Type Switching ◽  
Lethal Event ◽  
Bona Fide ◽  
Chromosome Iii ◽  
Mat Locus ◽  
Type Information

In homothallic cells of Saccharomyces cerevisiae, a or alpha mating type information at the mating type locus (MAT) is replaced by the transposition of the opposite mating type allele from HML alpha or HMRa. The rad52-1 mutation, which reduces mitotic and abolishes meiotic recombination, also affects homothallic switching (Malone and Esposito, Proc. Natl. Acad. Sci. U.S.A. 77:503-507, 1980). We have found that both HO rad52 MATa and HO rad52 MAT alpha cells die. This lethality is suppressed by mutations that substantially reduce but do not eliminate homothallic conversions. These mutations map at or near the MAT locus (MAT alpha inc, MATa-inc, MATa stk1) or are unlinked to MAT (HO-1 and swi1). These results suggest that the switching event itself is involved in the lethality. With the exception of swi1, HO rad52 strains carrying one of the above mutations cannot convert mating type at all. MAT alpha rad52 HO swi1 strains apparently can switch MAT alpha to MATa. However, when we analyzed these a maters, we found that few, if any, of them were bona fide MATa cells. These a-like cells were instead either deleted for part of chromosome III distal to and including MAT or had lost the entire third chromosome. Approximately 30% of the time, an a-like cell could be repaired to a normal MATa genotype if the cell was mated to a RAD52 MAT alpha-inc strain. The effects of rad52 were also studied in mata/MAT alpha-inc rad52/rad52 ho/HO diploids. When this diploid attempted to switch mata to MATa, an unstable broken chromosome was generated in nearly every cell. These studies suggest that homothallic switching involves the formation of a double-stranded deoxyribonucleic acid break or a structure which is labile in rad52 cells and results in a broken chromosome. We propose that the production of a double-stranded deoxyribonucleic acid break is the lethal event in rad52 HO cells.

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