scholarly journals Overexpression of the STE4 gene leads to mating response in haploid Saccharomyces cerevisiae.

1990 ◽  
Vol 10 (1) ◽  
pp. 217-222 ◽  
Author(s):  
M Whiteway ◽  
L Hougan ◽  
D Y Thomas
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
G Protein ◽  
Gene Product ◽  
Beta Subunit ◽  
Yeast Cells ◽  
Pheromone Response ◽  
Mating Pheromone ◽  
Cycle Arrest

The STE4 gene of Saccharomyces cerevisiae encodes the beta subunit of the yeast pheromone receptor-coupled G protein. Overexpression of the STE4 protein led to cell cycle arrest of haploid cells. This arrest was like the arrest mediated by mating pheromones in that it led to similar morphological changes in the arrested cells. The arrest occurred in haploid cells of either mating type but not in MATa/MAT alpha diploids, and it was suppressed by defects in genes such as STE12 that are needed for pheromone response. Overexpression of the STE4 gene product also suppressed the sterility of cells defective in the mating pheromone receptors encoded by the STE2 and STE3 genes. Cell cycle arrest mediated by STE4 overexpression was prevented in cells that either were overexpressing the SCG1 gene product (the alpha subunit of the G protein) or lacked the STE18 gene product (the gamma subunit of the G protein). This finding suggests that in yeast cells, the beta subunit is the limiting component of the active beta gamma element and that a proper balance in the levels of the G-protein subunits is critical to a normal mating pheromone response.

Overexpression of the STE4 gene leads to mating response in haploid Saccharomyces cerevisiae

1990 ◽  
Vol 10 (1) ◽  
pp. 217-222
Author(s):  
M Whiteway ◽  
L Hougan ◽  
D Y Thomas
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
G Protein ◽  
Gene Product ◽  
Beta Subunit ◽  
Yeast Cells ◽  
Pheromone Response ◽  
Mating Pheromone ◽  
Cycle Arrest

The STE4 gene of Saccharomyces cerevisiae encodes the beta subunit of the yeast pheromone receptor-coupled G protein. Overexpression of the STE4 protein led to cell cycle arrest of haploid cells. This arrest was like the arrest mediated by mating pheromones in that it led to similar morphological changes in the arrested cells. The arrest occurred in haploid cells of either mating type but not in MATa/MAT alpha diploids, and it was suppressed by defects in genes such as STE12 that are needed for pheromone response. Overexpression of the STE4 gene product also suppressed the sterility of cells defective in the mating pheromone receptors encoded by the STE2 and STE3 genes. Cell cycle arrest mediated by STE4 overexpression was prevented in cells that either were overexpressing the SCG1 gene product (the alpha subunit of the G protein) or lacked the STE18 gene product (the gamma subunit of the G protein). This finding suggests that in yeast cells, the beta subunit is the limiting component of the active beta gamma element and that a proper balance in the levels of the G-protein subunits is critical to a normal mating pheromone response.

scholarly journals FAR1 is required for posttranscriptional regulation of CLN2 gene expression in response to mating pheromone.

1993 ◽  
Vol 13 (2) ◽  
pp. 1013-1022 ◽  
Author(s):  
M H Valdivieso ◽  
K Sugimoto ◽  
K Y Jahng ◽  
P M Fernandes ◽  
C Wittenberg
Keyword(s):  
Cell Cycle ◽  
Posttranscriptional Regulation ◽  
Signal Transduction Pathway ◽  
Yeast Cells ◽  
Pheromone Response ◽  
Transcript Accumulation ◽  
Mating Pheromone ◽  
Cycle Arrest ◽  
Feedback Pathway ◽  
Posttranscriptional Level

Yeast cells arrest during the G1 interval of the cell cycle in response to peptide mating pheromones. The FAR1 gene is required for cell cycle arrest but not for a number of other aspects of the pheromone response. Genetic evidence suggests that FAR1 is required specifically for inactivation of the G1 cyclin CLN2. From these observations, the FAR1 gene has been proposed to encode an element of the interface between the mating pheromone signal transduction pathway and the cell cycle regulatory apparatus. We show here that FAR1 is necessary for the decrease in CLN1 and CLN2 transcript accumulation observed in response to mating pheromone but is unnecessary for regulation of the same transcripts during vegetative growth. However, the defect in regulation of CLN1 expression is dependent upon CLN2. We show that pheromone regulates the abundance of Cln2 at a posttranscriptional level and that FAR1 is required for that regulation. From these observations, we suggest that FAR1 function is limited to posttranscriptional regulation of CLN2 expression by mating pheromone. The failure of mating pheromone to repress CLN2 transcript levels in far1 mutants can be explained by the stimulatory effect of the persistent Cln2 protein on CLN2 transcription via the CLN/CDC28-dependent feedback pathway.

Mutations in cell division cycle genes CDC36 and CDC39 activate the Saccharomyces cerevisiae mating pheromone response pathway

1990 ◽  
Vol 10 (6) ◽  
pp. 2966-2972
Author(s):  
M de Barros Lopes ◽  
J Y Ho ◽  
S I Reed
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
G Protein ◽  
G1 Arrest ◽  
Restrictive Temperature ◽  
Gene Products ◽  
Pheromone Response ◽  
Mating Pheromone ◽  
Pheromone Response Pathway ◽  
Mutational Activation

Conditional mutations in the genes CDC36 and CDC39 cause arrest in the G1 phase of the Saccharomyces cerevisiae cell cycle at the restrictive temperature. We present evidence that this arrest is a consequence of a mutational activation of the mating pheromone response. cdc36 and cdc39 mutants expressed pheromone-inducible genes in the absence of pheromone and conjugated in the absence of a mating pheromone receptor. On the other hand, cells lacking the G beta subunit or overproducing the G alpha subunit of the transducing G protein that couples the receptor to the pheromone response pathway prevented constitutive activation of the pathway in cdc36 and cdc39 mutants. These epistasis relationships imply that the CDC36 and CDC39 gene products act at the level of the transducing G protein. The CDC36 and CDC39 gene products have a role in cellular processes other than the mating pheromone response. A mating-type heterozygous diploid cell, homozygous for either the cdc36 or cdc39 mutation, does not exhibit the G1 arrest phenotype but arrests asynchronously with respect to the cell cycle. A similar asynchronous arrest was observed in cdc36 and cdc39 cells where the pheromone response pathway had been inactivated by mutations in the transducing G protein. Furthermore, cdc36 and cdc39 mutants, when grown on carbon catabolite-derepressing medium, did not arrest in G1 and did not induce pheromone-specific genes at the restrictive temperature.

Saccharomyces cerevisiae mutants lacking a functional vacuole are defective for aspects of the pheromone response

1990 ◽  
Vol 97 (3) ◽  
pp. 517-525 ◽  
Author(s):  
V. Dulic ◽  
H. Riezman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Carbon Sources ◽  
Early Response ◽  
Pheromone Response ◽  
Optimal Response ◽  
Cycle Arrest ◽  
Alpha Factor ◽  
Vacuolar Protein

The end1 mutant belongs to a group of four vacuolar protein sorting mutants (class C vps) that lack a morphologically distinguishable and functional vacuole. These mutants share several other phenotypes, such as the inability to grow at 37 degrees C or on nonfermentable carbon sources. We show that, as in the case of the end1 mutant, vps16, vps18 and vps33 mutants all internalize but do not degrade alpha-factor. In addition, all four mutants are defective for alpha-factor-induced projection formation to the same extent. A more detailed investigation of pheromone response in the end1 mutant reveals that one aspect of the early response (induction of FUS1) is as defective as late responses (cell cycle arrest and projection formation). In contrast, another measure of the early response (induction of STE2) is normal. These data suggest that the biogenesis of a functional vacuole is necessary for optimal response to pheromone.

Characterization of RAD9 of Saccharomyces cerevisiae and evidence that its function acts posttranslationally in cell cycle arrest after DNA damage

1990 ◽  
Vol 10 (12) ◽  
pp. 6554-6564
Author(s):  
T A Weinert ◽  
L H Hartwell
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Dna Damage ◽  
Cell Cycle Arrest ◽  
Dna Sequence ◽  
Negative Regulator ◽  
Yeast Cells ◽  
Protein Synthesis Inhibitor ◽  
Cycle Arrest ◽  
Delta Cells

In eucaryotic cells, incompletely replicated or damaged chromosomes induce cell cycle arrest in G2 before mitosis, and in the yeast Saccharomyces cerevisiae the RAD9 gene is essential for the cell cycle arrest (T.A. Weinert and L. H. Hartwell, Science 241:317-322, 1988). In this report, we extend the analysis of RAD9-dependent cell cycle control. We found that both induction of RAD9-dependent arrest in G2 and recovery from arrest could occur in the presence of the protein synthesis inhibitor cycloheximide, showing that the mechanism of RAD9-dependent control involves a posttranslational mechanism(s). We have isolated and determined the DNA sequence of the RAD9 gene, confirming the DNA sequence reported previously (R. H. Schiestl, P. Reynolds, S. Prakash, and L. Prakash, Mol. Cell. Biol. 9:1882-1886, 1989). The predicted protein sequence for the Rad9 protein bears no similarity to sequences of known proteins. We also found that synthesis of the RAD9 transcript in the cell cycle was constitutive and not induced by X-irradiation. We constructed yeast cells containing a complete deletion of the RAD9 gene; the rad9 null mutants were viable, sensitive to X- and UV irradiation, and defective for cell cycle arrest after DNA damage. Although Rad+ and rad9 delta cells had similar growth rates and cell cycle kinetics in unirradiated cells, the spontaneous rate of chromosome loss (in unirradiated cells) was elevated 7- to 21-fold in rad9 delta cells. These studies show that in the presence of induced or endogenous DNA damage, RAD9 is a negative regulator that inhibits progression from G2 in order to preserve cell viability and to maintain the fidelity of chromosome transmission.

A RAD9-dependent checkpoint blocks meiosis of cdc13 yeast cells.

Genetics ◽  
1992 ◽  
Vol 131 (1) ◽  
pp. 55-63 ◽  
Author(s):  
L Weber ◽  
B Byers
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Mitotic Division ◽  
Yeast Cells ◽  
Mitotic Progression ◽  
Meiotic Progression ◽  
Cycle Arrest ◽  
Yeast Meiosis

Abstract Mutations in CDC13 have previously been found to cause cell cycle arrest of Saccharomyces cerevisiae at a stage in G2 immediately preceding the mitotic division. We show here that cdc13 blocks the meiotic pathway at a stage that follows DNA replication, but in this case the spindle has not yet formed nor have the chromosomes undergone synapsis or recombination. This arrest is alleviated by rad9, thus implicating the same checkpoint function that delays mitotic progression when chromosomal lesions are present. An assessment of the spores produced upon alleviation of the meiotic arrest by rad9 reveals that the absence of recombination in strains bearing cdc13 alone is attributable to the RAD9-mediated arrest rather than to other effects of cdc13 lesions. We have tested the possibility that this checkpoint function is important in regulating meiotic progression to permit resolution of recombinational intermediates during ongoing meiosis and have found no evidence that rad9 alters the execution of functions that might depend upon such regulation. We consider the possible role of other checkpoints in yeast meiosis.

scholarly journals Mutations in cell division cycle genes CDC36 and CDC39 activate the Saccharomyces cerevisiae mating pheromone response pathway.

1990 ◽  
Vol 10 (6) ◽  
pp. 2966-2972 ◽  
Author(s):  
M de Barros Lopes ◽  
J Y Ho ◽  
S I Reed
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
G Protein ◽  
G1 Arrest ◽  
Restrictive Temperature ◽  
Gene Products ◽  
Pheromone Response ◽  
Mating Pheromone ◽  
Pheromone Response Pathway ◽  
Mutational Activation

Conditional mutations in the genes CDC36 and CDC39 cause arrest in the G1 phase of the Saccharomyces cerevisiae cell cycle at the restrictive temperature. We present evidence that this arrest is a consequence of a mutational activation of the mating pheromone response. cdc36 and cdc39 mutants expressed pheromone-inducible genes in the absence of pheromone and conjugated in the absence of a mating pheromone receptor. On the other hand, cells lacking the G beta subunit or overproducing the G alpha subunit of the transducing G protein that couples the receptor to the pheromone response pathway prevented constitutive activation of the pathway in cdc36 and cdc39 mutants. These epistasis relationships imply that the CDC36 and CDC39 gene products act at the level of the transducing G protein. The CDC36 and CDC39 gene products have a role in cellular processes other than the mating pheromone response. A mating-type heterozygous diploid cell, homozygous for either the cdc36 or cdc39 mutation, does not exhibit the G1 arrest phenotype but arrests asynchronously with respect to the cell cycle. A similar asynchronous arrest was observed in cdc36 and cdc39 cells where the pheromone response pathway had been inactivated by mutations in the transducing G protein. Furthermore, cdc36 and cdc39 mutants, when grown on carbon catabolite-derepressing medium, did not arrest in G1 and did not induce pheromone-specific genes at the restrictive temperature.

FAR1 is required for posttranscriptional regulation of CLN2 gene expression in response to mating pheromone

1993 ◽  
Vol 13 (2) ◽  
pp. 1013-1022
Author(s):  
M H Valdivieso ◽  
K Sugimoto ◽  
K Y Jahng ◽  
P M Fernandes ◽  
C Wittenberg
Keyword(s):  
Cell Cycle ◽  
Posttranscriptional Regulation ◽  
Signal Transduction Pathway ◽  
Yeast Cells ◽  
Pheromone Response ◽  
Transcript Accumulation ◽  
Mating Pheromone ◽  
Cycle Arrest ◽  
Feedback Pathway ◽  
Posttranscriptional Level

Yeast cells arrest during the G1 interval of the cell cycle in response to peptide mating pheromones. The FAR1 gene is required for cell cycle arrest but not for a number of other aspects of the pheromone response. Genetic evidence suggests that FAR1 is required specifically for inactivation of the G1 cyclin CLN2. From these observations, the FAR1 gene has been proposed to encode an element of the interface between the mating pheromone signal transduction pathway and the cell cycle regulatory apparatus. We show here that FAR1 is necessary for the decrease in CLN1 and CLN2 transcript accumulation observed in response to mating pheromone but is unnecessary for regulation of the same transcripts during vegetative growth. However, the defect in regulation of CLN1 expression is dependent upon CLN2. We show that pheromone regulates the abundance of Cln2 at a posttranscriptional level and that FAR1 is required for that regulation. From these observations, we suggest that FAR1 function is limited to posttranscriptional regulation of CLN2 expression by mating pheromone. The failure of mating pheromone to repress CLN2 transcript levels in far1 mutants can be explained by the stimulatory effect of the persistent Cln2 protein on CLN2 transcription via the CLN/CDC28-dependent feedback pathway.

scholarly journals Characterization of RAD9 of Saccharomyces cerevisiae and evidence that its function acts posttranslationally in cell cycle arrest after DNA damage.

1990 ◽  
Vol 10 (12) ◽  
pp. 6554-6564 ◽  
Author(s):  
T A Weinert ◽  
L H Hartwell
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Dna Damage ◽  
Cell Cycle Arrest ◽  
Dna Sequence ◽  
Negative Regulator ◽  
Yeast Cells ◽  
Protein Synthesis Inhibitor ◽  
Cycle Arrest ◽  
Delta Cells

In eucaryotic cells, incompletely replicated or damaged chromosomes induce cell cycle arrest in G2 before mitosis, and in the yeast Saccharomyces cerevisiae the RAD9 gene is essential for the cell cycle arrest (T.A. Weinert and L. H. Hartwell, Science 241:317-322, 1988). In this report, we extend the analysis of RAD9-dependent cell cycle control. We found that both induction of RAD9-dependent arrest in G2 and recovery from arrest could occur in the presence of the protein synthesis inhibitor cycloheximide, showing that the mechanism of RAD9-dependent control involves a posttranslational mechanism(s). We have isolated and determined the DNA sequence of the RAD9 gene, confirming the DNA sequence reported previously (R. H. Schiestl, P. Reynolds, S. Prakash, and L. Prakash, Mol. Cell. Biol. 9:1882-1886, 1989). The predicted protein sequence for the Rad9 protein bears no similarity to sequences of known proteins. We also found that synthesis of the RAD9 transcript in the cell cycle was constitutive and not induced by X-irradiation. We constructed yeast cells containing a complete deletion of the RAD9 gene; the rad9 null mutants were viable, sensitive to X- and UV irradiation, and defective for cell cycle arrest after DNA damage. Although Rad+ and rad9 delta cells had similar growth rates and cell cycle kinetics in unirradiated cells, the spontaneous rate of chromosome loss (in unirradiated cells) was elevated 7- to 21-fold in rad9 delta cells. These studies show that in the presence of induced or endogenous DNA damage, RAD9 is a negative regulator that inhibits progression from G2 in order to preserve cell viability and to maintain the fidelity of chromosome transmission.

scholarly journals Mot3, a Zn Finger Transcription Factor That Modulates Gene Expression and Attenuates Mating Pheromone Signaling in Saccharomyces cerevisiae

Genetics ◽  
1998 ◽  
Vol 149 (2) ◽  
pp. 879-892 ◽  
Author(s):  
Anatoly V Grishin ◽  
Michael Rothenberg ◽  
Maureen A Downs ◽  
Kendall J Blumer
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Signaling Pathway ◽  
G Protein ◽  
Binding Sites ◽  
Dependent Manner ◽  
Pheromone Response ◽  
Mating Pheromone ◽  
Pheromone Signaling ◽  
Yeast Genes

Abstract In the yeast Saccharomyces cerevisiae, mating pheromone response is initiated by activation of a G protein- and mitogen-activated protein (MAP) kinase-dependent signaling pathway and attenuated by several mechanisms that promote adaptation or desensitization. To identify genes whose products negatively regulate pheromone signaling, we screened for mutations that suppress the hyperadaptive phenotype of wild-type cells overexpressing signaling-defective G protein β subunits. This identified recessive mutations in MOT3, which encodes a nuclear protein with two Cys2-His2 Zn fingers. MOT3 was found to be a dosage-dependent inhibitor of pheromone response and pheromone-induced gene expression and to require an intact signaling pathway to exert its effects. Several results suggested that Mot3 attenuates expression of pheromone-responsive genes by mechanisms distinct from those used by the negative transcriptional regulators Cdc36, Cdc39, and Mot2. First, a Mot3-lexA fusion functions as a transcriptional activator. Second, Mot3 is a dose-dependent activator of several genes unrelated to pheromone response, including CYC1, SUC2, and LEU2. Third, insertion of consensus Mot3 binding sites (C/A/T)AGG(T/C)A activates a promoter in a MOT3-dependent manner. These findings, and the fact that consensus binding sites are found in the 5′ flanking regions of many yeast genes, suggest that Mot3 is a globally acting transcriptional regulator. We hypothesize that Mot3 regulates expression of factors that attenuate signaling by the pheromone response pathway.

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