SPT3 is required for normal levels of a-factor and alpha-factor expression in Saccharomyces cerevisiae

1988 ◽  
Vol 8 (2) ◽  
pp. 822-827
Author(s):  
J N Hirschhorn ◽  
F Winston
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mating Type ◽  
Transcription Initiation ◽  
Nuclear Fusion ◽  
Mating Pheromone ◽  
Noncoding Regions ◽  
Factor Expression ◽  
Alpha Factor ◽  
Cellular Fusion ◽  
Insertion Mutations

Mutations in the Saccharomyces cerevisiae SPT3 gene were previously found to cause suppression of Ty and delta insertion mutations in 5'-noncoding regions of genes. This suppression likely results from the fact that SPT3 is required for transcription initiation in delta sequences. Other additional phenotypes of spt3 mutants, including a mating defect, suggest that SPT3 is required for normal levels of expression of other genes. We analyzed the mating defect in spt3 mutants and showed that the levels of transcripts of the three major mating pheromone genes, MF alpha 1, MFa1, MFa2, were all reduced. The reduction in expression of these genes in spt3 mutants was not due to expression of a silent mating type cassette. Furthermore, we showed that the spt3 mating defect was manifest at the levels of both cellular fusion and nuclear fusion.

scholarly journals SPT3 is required for normal levels of a-factor and alpha-factor expression in Saccharomyces cerevisiae.

1988 ◽  
Vol 8 (2) ◽  
pp. 822-827 ◽  
Author(s):  
J N Hirschhorn ◽  
F Winston
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mating Type ◽  
Transcription Initiation ◽  
Nuclear Fusion ◽  
Mating Pheromone ◽  
Noncoding Regions ◽  
Factor Expression ◽  
Alpha Factor ◽  
Cellular Fusion ◽  
Insertion Mutations

Mutations in the Saccharomyces cerevisiae SPT3 gene were previously found to cause suppression of Ty and delta insertion mutations in 5'-noncoding regions of genes. This suppression likely results from the fact that SPT3 is required for transcription initiation in delta sequences. Other additional phenotypes of spt3 mutants, including a mating defect, suggest that SPT3 is required for normal levels of expression of other genes. We analyzed the mating defect in spt3 mutants and showed that the levels of transcripts of the three major mating pheromone genes, MF alpha 1, MFa1, MFa2, were all reduced. The reduction in expression of these genes in spt3 mutants was not due to expression of a silent mating type cassette. Furthermore, we showed that the spt3 mating defect was manifest at the levels of both cellular fusion and nuclear fusion.

scholarly journals Isolation and genetic analysis of Saccharomyces cerevisiae mutants supersensitive to G1 arrest by a factor and alpha factor pheromones.

1982 ◽  
Vol 2 (1) ◽  
pp. 11-20 ◽  
Author(s):  
R K Chan ◽  
C A Otte
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mating Type ◽  
Solid Medium ◽  
G1 Arrest ◽  
Alpha Cells ◽  
Opposite Mating Type ◽  
Alpha Factor ◽  
Complementation Groups ◽  
Chromosome Iii ◽  
The Right

Eight independently isolated mutants which are supersensitive (Sst-) to the G1 arrest induced by the tridecapeptide pheromone alpha factor were identified by screening mutagenized Saccharomyces cerevisiae MATa cells on solid medium for increased growth inhibition by alpha factor. These mutants carried lesions in two complementation groups, sst1 and sst2. Mutations at the sst1 locus were mating type specific: MATa sst1 cells were supersensitive to alpha factor, but MAT alpha sst1 cells were not supersensitive to a factor. In contrast, mutations at the sst2 locus conferred supersensitivity to the pheromones of the opposite mating type on both MATa and MAT alpha cells. Even in the absence of added alpha pheromone, about 10% of the cells in exponentially growing cultures of MATa strains carrying any of three different alleles of sst2 (including the ochre mutation sst2-4) had the aberrant morphology ("shmoo" shape) that normally develops only after MATa cells are exposed to alpha factor. This "self-shmooing" phenotype was genetically linked to the sst2 mutations, although the leakiest allele isolated (sst2-3) did not display this characteristic. Normal MATa/MAT alpha diploids do not respond to pheromones; diploids homozygous for an sst2 mutation (MATa/MAT alpha sst2-1/sst2-1) were still insensitive to alpha factor. The sst1 gene was mapped to within 6.9 centimorgans of his6 on chromosome IX. The sst2 gene was unlinked to sst1, was not centromere linked, and was shown to be neither linked to nor centromere distal to MAT on the right arm of chromosome III.

Isolation and genetic analysis of Saccharomyces cerevisiae mutants supersensitive to G1 arrest by a factor and alpha factor pheromones

1982 ◽  
Vol 2 (1) ◽  
pp. 11-20
Author(s):  
R K Chan ◽  
C A Otte
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mating Type ◽  
Solid Medium ◽  
G1 Arrest ◽  
Alpha Cells ◽  
Opposite Mating Type ◽  
Alpha Factor ◽  
Complementation Groups ◽  
Chromosome Iii ◽  
The Right

Eight independently isolated mutants which are supersensitive (Sst-) to the G1 arrest induced by the tridecapeptide pheromone alpha factor were identified by screening mutagenized Saccharomyces cerevisiae MATa cells on solid medium for increased growth inhibition by alpha factor. These mutants carried lesions in two complementation groups, sst1 and sst2. Mutations at the sst1 locus were mating type specific: MATa sst1 cells were supersensitive to alpha factor, but MAT alpha sst1 cells were not supersensitive to a factor. In contrast, mutations at the sst2 locus conferred supersensitivity to the pheromones of the opposite mating type on both MATa and MAT alpha cells. Even in the absence of added alpha pheromone, about 10% of the cells in exponentially growing cultures of MATa strains carrying any of three different alleles of sst2 (including the ochre mutation sst2-4) had the aberrant morphology ("shmoo" shape) that normally develops only after MATa cells are exposed to alpha factor. This "self-shmooing" phenotype was genetically linked to the sst2 mutations, although the leakiest allele isolated (sst2-3) did not display this characteristic. Normal MATa/MAT alpha diploids do not respond to pheromones; diploids homozygous for an sst2 mutation (MATa/MAT alpha sst2-1/sst2-1) were still insensitive to alpha factor. The sst1 gene was mapped to within 6.9 centimorgans of his6 on chromosome IX. The sst2 gene was unlinked to sst1, was not centromere linked, and was shown to be neither linked to nor centromere distal to MAT on the right arm of chromosome III.

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.

Multiple regulation of STE2, a mating-type-specific gene of Saccharomyces cerevisiae

1986 ◽  
Vol 6 (6) ◽  
pp. 2106-2114
Author(s):  
A Hartig ◽  
J Holly ◽  
G Saari ◽  
V L MacKay
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Regulation ◽  
Gene Product ◽  
Mating Type ◽  
Type A ◽  
Specific Gene ◽  
Alpha Cells ◽  
Pheromone Response ◽  
Mating Pheromone ◽  
A Cells

The Saccharomyces cerevisiae STE2 gene, which is required for pheromone response and conjugation specifically in mating-type a cells, was cloned by complementation of the ste2 mutation. Transcription of STE2 is repressed by the MAT alpha 2 gene product, so that the 1.4-kilobase STE2 RNA is detected only in a or mat alpha 2 strains, not in alpha or a/alpha cells. However, STE2 RNA levels are also increased by the mating pheromone alpha-factor and decreased in strains bearing mutations in the nonspecific STE4 gene. Regulation of STE2 expression in a cells is therefore achieved by several mechanisms.

scholarly journals Regulation of alpha-factor production in Saccharomyces cerevisiae: a-factor pheromone-induced expression of the MF alpha 1 and STE13 genes.

1989 ◽  
Vol 9 (10) ◽  
pp. 4507-4514 ◽  
Author(s):  
T Achstetter
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Fusion Gene ◽  
Invertase Activity ◽  
Transcriptional Level ◽  
Alpha Cells ◽  
Mating Pheromone ◽  
Induced Expression ◽  
Alpha Factor ◽  
Suc2 Gene ◽  
Proteolytic Activities

Production of the mating pheromone alpha-factor was examined in Saccharomyces cerevisiae MAT alpha cells that had been exposed to the mating pheromone a-factor. A 2-h treatment with a-factor caused a significant increase in alpha-factor concentration in the medium as demonstrated by a halo assay. MF alpha 1 is one of the two genes coding for a precursor of alpha-factor. A Northern (RNA) analysis of total RNA from a-factor-treated MAT alpha cells revealed a rapid two- to threefold increase in MF alpha 1 transcript levels, reaching maximum within 60 min of exposure to the pheromone. Pheromone induction did not require ongoing protein synthesis. a-Factor-induced MF alpha 1 expression was quantitated by analysis of an MF alpha 1::SUC2 fusion gene whose product was assayed for invertase activity. Expression of the MF alpha 1::SUC2 gene in MAT alpha cells responded to the a-factor signal like the chromosomal version of MF alpha 1. Maturation of the alpha-factor precursor involves three proteolytic activities which are encoded by the KEX1, KEX2, and STE13 genes, respectively. Two of these genes, namely, KEX2 and STE13, were examined for pheromone-induced expression. Only the STE13 gene exhibited pheromone induction at the transcriptional level.

scholarly journals Saccharomyces cerevisiae nuclear fusion requires prior activation by alpha factor.

1986 ◽  
Vol 6 (10) ◽  
pp. 3490-3497 ◽  
Author(s):  
M D Rose ◽  
B R Price ◽  
G R Fink
Keyword(s):  
Mating Type ◽  
Nuclear Fusion ◽  
Cell System ◽  
Fusion System ◽  
Intact Cells ◽  
Type Locus ◽  
Constitutive Function ◽  
Alpha Factor ◽  
Prior Arrest ◽  
Spheroplast Fusion

We have developed a protocol for efficient fusion of spheroplasts of the same mating type. Nuclear fusion in this whole-cell system is also efficient and closely parallels nuclear fusion in heterosexual mating of intact cells. In the spheroplast fusion system, nuclear fusion is dependent on both the KAR1 gene and prior exposure to alpha factor. The major products of nuclear fusion in the spheroplast fusion assay were true diploids that were homozygous at the mating-type locus. An additional 10% of the products were cells of ploidy greater than diploid. The dependence of nuclear fusion on alpha factor treatment could not be replaced by synchronization in G1 by mutations in CDC28 and CDC35 or by prior arrest in stationary phase. These data suggest that nuclear fusion is not a constitutive function of the nucleus, but rather is specifically induced by mating hormone.

scholarly journals The Saccharomyces cerevisiae SPT7 gene encodes a very acidic protein important for transcription in vivo.

Genetics ◽  
1995 ◽  
Vol 139 (2) ◽  
pp. 523-536 ◽  
Author(s):  
L J Gansheroff ◽  
C Dollard ◽  
P Tan ◽  
F Winston
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transcription Initiation ◽  
Tata Binding Protein ◽  
Acidic Protein ◽  
Null Mutation ◽  
Start Site ◽  
Transcription Start ◽  
Mutant Phenotypes ◽  
Insertion Mutations

Abstract Mutations in the SPT7 gene of Saccharomyces cerevisiae originally were identified as suppressors of Ty and delta insertion mutations in the 5' regions of the HIS4 and LYS2 genes. Other genes that have been identified in mutant hunts of this type have been shown to play a role in transcription. In this work we show that SPT7 is also important for proper transcription in vivo. We have cloned and sequenced the SPT7 gene and have shown that it encodes a large, acidic protein that is localized to the nucleus. The SPT7 protein contains a bromodomain sequence; a deletion that removes the bromodomain from the SPT7 protein causes no detectable mutant phenotype. Strains that contain an spt7 null mutation are viable but grow very slowly and have transcriptional defects at many loci including insertion mutations, Ty elements, the INO1 gene and the MFA1 gene. These transcriptional defects and other mutant phenotypes are similar to those caused by certain mutations in SPT15, which encodes the TATA binding protein (TBP). The similarity of the phenotypes of spt7 and spt15 mutants, including effects of spt7 mutations on the transcription start site of certain genes, suggests that SPT7 plays an important role in transcription initiation in vivo.

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