Structure of the Saccharomyces cerevisiae HO gene and analysis of its upstream regulatory region.

The HO gene product of Saccharomyces cerevisiae is a site-specific endonuclease that initiates mating type interconversion. We have determined the nucleotide sequence of a 3,129-base-pair (bp) segment containing HO. The segment contains a single long open reading frame encoding a polypeptide of 586 amino acids, which has unusual (unbiased) codon usage and is preceded by 762 bp of upstream region. The predicted HO protein is basic (16% lysine and arginine) and is calculated to have a secondary structure that is 30% helical. The corresponding transcript is initiated approximately 50 nucleotides prior to the presumed initiation codon. Insertion of an Escherichia coli lacZ gene fragment into the putative HO coding segment inactivated HO and formed a hybrid HO-lacZ gene whose beta-galactosidase activity was regulated by the mating type locus in the same manner as HO (repressed by a 1-alpha 2). Upstream regions of 1,360 and 762 bp conferred strong repression; 436 bp led to partial constitutivity and 301 bp to full constitutivity. Thus, DNA sequences that confer repression of HO by a1-alpha 2 are at least 250 nucleotides upstream of the transcription start point and are within 436 nucleotides of the HO initiation codon. The progressive loss of repression suggests that both the -762 to -436 and the -436 to -301 intervals contain sites for regulation by a1-alpha 2. The HO gene contains two distinct regions that promote autonomous replication of plasmids in S. cerevisiae. These regions contain sequences that are homologous to the two conserved sequences that are associated with ARS activity.

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INTERCONVERSION OF YEAST MATING TYPES II. RESTORATION OF MATING ABILITY TO STERILE MUTANTS IN HOMOTHALLIC AND HETEROTHALLIC STRAINS

Genetics ◽

10.1093/genetics/85.3.373 ◽

1977 ◽

Vol 85 (3) ◽

pp. 373A-393

Author(s):

James B Hicks ◽

Ira Herskowitz

Keyword(s):

Saccharomyces Cerevisiae ◽

Mating Type ◽

Yeast Cells ◽

Mating Types ◽

Yeast Saccharomyces Cerevisiae ◽

Type Locus ◽

Mating Ability ◽

Mating Type Locus ◽

Regulatory Information ◽

Ho Gene

ABSTRACT The two mating types of the yeast Saccharomyces cerevisiae can be interconverted in both homothallic and heterothallic strains. Previous work indicates that all yeast cells contain the information to be both a and α and that the HO gene (in homothallic strains) promotes a change in mating type by causing a change at the mating type locus itself. In both heterothallic and homothallic strains, a defective α mating type locus can be converted to a functional a locus and subsequently to a functional α locus. In contrast, action of the HO gene does not restore mating ability to a strain defective in another gene for mating which is not at the mating type locus. These observations indicate that a yeast cell contains an additional copy (or copies) of α information, and lead to the "cassette" model for mating type interconversion. In this model, HM a and hmα loci are blocs of unexpressed α regulatory information, and HMα and hm a loci are blocs of unexpressed a regulatory information. These blocs are silent because they lack an essential site for expression, and become active upon insertion of this information (or a copy of the information) into the mating type locus by action of the HO gene.

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DEPENDENCE ON MATING TYPE FOR THE OVERPRODUCTION OF ISO-2-CYTOCHROME c IN THE YEAST MUTANT CYC7–H2

Genetics ◽

10.1093/genetics/94.4.891 ◽

1980 ◽

Vol 94 (4) ◽

pp. 891-898

Author(s):

Rodney J Rothstein ◽

Fred Sherman

Keyword(s):

Saccharomyces Cerevisiae ◽

Cytochrome C ◽

Mating Type ◽

Regulatory Region ◽

The Other ◽

Yeast Mutant ◽

Yeast Saccharomyces Cerevisiae ◽

Type Locus ◽

Active Expression ◽

Structural Region

ABSTRACT The CYC7-H2 mutation causes an approximately 20-fold overproduction of iso-2-cytochromo c in a and α haploid strains of the yeast Saccharomyces cerevisiae due to an alteration in the nontranslated regulatory region that is presumably contiguous with the structural region. In this investigation, we demonstrated that heterozygosity at the mating type locus, a /α or a/a/α/α, prevents expression of the overproduction, while homozygosity, a/a and α/α and hemizygosity, a/O and α/O, allow full expression of the CYC7-H2 mutation, equivalent to the expression observed in a and α haploid strains. There is no decrease in the overproduction of iso-2-cytochrome c in a/α diploid strains containing either of the other two similar mutations, CYC7-H1 and CYC7-H3. It appears as if active expression of one or another of the mating-type alleles is required for the overproduction of iso-2-cytochrome c in CYC7-H2 mutants.

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Isolation and functional analysis of sporulation-induced transcribed sequences from Saccharomyces cerevisiae.

Molecular and Cellular Biology ◽

10.1128/mcb.6.6.2185 ◽

1986 ◽

Vol 6 (6) ◽

pp. 2185-2197 ◽

Cited By ~ 17

Author(s):

E Gottlin-Ninfa ◽

D B Kaback

Keyword(s):

Saccharomyces Cerevisiae ◽

Mating Type ◽

Dna Sequences ◽

Recombinant Dna ◽

Spore Formation ◽

Yeast Genome ◽

In Vitro Mutagenesis ◽

Sporulation Medium ◽

Type Locus ◽

Mating Type Locus

Strains of the yeast Saccharomyces cerevisiae that are heterozygous for the mating-type locus (MATa/MAT alpha) undergo meiosis and spore formation when they are starved for nitrogen and are provided with a nonfermentable carbon source such as potassium acetate. Haploids and diploids homozygous for the mating-type locus (MAT alpha/MAT alpha or MATa/MATa) are asporogenous and undergo neither meiosis nor spore formation when incubated under the same conditions. A small number of genes produce transcripts that appear to be induced specifically in sporulating cells. These transcripts either are not found or are present at much lower levels both in vegetatively growing cells and in cells from asporogenous strains that have been incubated in sporulation medium. Several genes complementary to these MATa/MAT alpha-dependent sporulation-induced transcripts were isolated from a gene-size insert yeast-lambda recombinant DNA library, by differential-plaque filter hybridization. An attempt was made to determine the function of three of these genes by mutating them in the yeast genome with in vitro mutagenesis and one-step gene replacement techniques. One gene was extensively disrupted by both a 0.3-kilobase deletion and the insertion of two large DNA sequences at different sites within the gene. Surprisingly, this compound mutation did not appear to affect meiosis or the production of viable ascospores, indicating that this gene was dispensable for differentiation. The other two genes were disrupted by simple insertion mutations at a site where it was possible that they might still possess some gene activity. These mutations also did not appear to affect sporulation. These results suggest that not all sporulation-induced genes are essential for meiosis and the production of viable ascospores under the conditions examined.

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

Genetics ◽

10.1093/genetics/85.3.395 ◽

1977 ◽

Vol 85 (3) ◽

pp. 395-405 ◽

Cited By ~ 7

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.

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Isolation and functional analysis of sporulation-induced transcribed sequences from Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.6.6.2185-2197.1986 ◽

1986 ◽

Vol 6 (6) ◽

pp. 2185-2197

Author(s):

E Gottlin-Ninfa ◽

D B Kaback

Keyword(s):

Saccharomyces Cerevisiae ◽

Mating Type ◽

Dna Sequences ◽

Recombinant Dna ◽

Spore Formation ◽

Yeast Genome ◽

In Vitro Mutagenesis ◽

Sporulation Medium ◽

Type Locus ◽

Mating Type Locus

Strains of the yeast Saccharomyces cerevisiae that are heterozygous for the mating-type locus (MATa/MAT alpha) undergo meiosis and spore formation when they are starved for nitrogen and are provided with a nonfermentable carbon source such as potassium acetate. Haploids and diploids homozygous for the mating-type locus (MAT alpha/MAT alpha or MATa/MATa) are asporogenous and undergo neither meiosis nor spore formation when incubated under the same conditions. A small number of genes produce transcripts that appear to be induced specifically in sporulating cells. These transcripts either are not found or are present at much lower levels both in vegetatively growing cells and in cells from asporogenous strains that have been incubated in sporulation medium. Several genes complementary to these MATa/MAT alpha-dependent sporulation-induced transcripts were isolated from a gene-size insert yeast-lambda recombinant DNA library, by differential-plaque filter hybridization. An attempt was made to determine the function of three of these genes by mutating them in the yeast genome with in vitro mutagenesis and one-step gene replacement techniques. One gene was extensively disrupted by both a 0.3-kilobase deletion and the insertion of two large DNA sequences at different sites within the gene. Surprisingly, this compound mutation did not appear to affect meiosis or the production of viable ascospores, indicating that this gene was dispensable for differentiation. The other two genes were disrupted by simple insertion mutations at a site where it was possible that they might still possess some gene activity. These mutations also did not appear to affect sporulation. These results suggest that not all sporulation-induced genes are essential for meiosis and the production of viable ascospores under the conditions examined.

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rme1 Mutation of Saccharomyces cerevisiae: map position and bypass of mating type locus control of sporulation.

Molecular and Cellular Biology ◽

10.1128/mcb.1.10.958 ◽

1981 ◽

Vol 1 (10) ◽

pp. 958-960 ◽

Cited By ~ 35

Author(s):

J Rine ◽

G F Sprague ◽

I Herskowitz

Keyword(s):

Saccharomyces Cerevisiae ◽

Mating Type ◽

Type Locus ◽

Mating Type Locus ◽

Type Information

Sporulation in Saccharomyces cerevisiae normally occurs only in MATa/MAT alpha diploids. We show that mutations in RME1 bypassed the requirements for both a and alpha mating type information in sporulation and therefore allowed MATa/MATa and MAT alpha/MAT alpha diploids to sporulate. RME1 was located on chromosome VII, between LEU1 and ADE6.

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The alpha-mating type locus of Cryptococcus neoformans contains a peptide pheromone gene

Molecular and Cellular Biology ◽

10.1128/mcb.13.3.1962-1970.1993 ◽

1993 ◽

Vol 13 (3) ◽

pp. 1962-1970

Author(s):

T D Moore ◽

J C Edman

Keyword(s):

Cryptococcus Neoformans ◽

Mating Type ◽

Cell Types ◽

Yeast Cells ◽

Mating Types ◽

Type Locus ◽

Reading Frame ◽

Cloning Method ◽

Opportunistic Fungal Pathogen ◽

Specific Fragment

The opportunistic fungal pathogen Cryptococcus neoformans has two mating types, MATa and MAT alpha. The MAT alpha strains are more virulent. Mating of opposite mating type haploid yeast cells results in the production of a filamentous hyphal phase. The MAT alpha locus has been isolated in this study in order to identify the genetic differences between mating types and their contribution to virulence. A 138-bp fragment of MAT alpha-specific DNA which cosegregates with alpha-mating type was isolated by using a difference cloning method. Overlapping phage and cosmid clones spanning the entire MAT alpha locus were isolated by using this MAT alpha-specific fragment as a probe. Mapping of these clones physically defined the MAT alpha locus to a 35- to 45-kb region which is present only in MAT alpha strains. Transformation studies with fragments of the MAT alpha locus identified a 2.1-kb XbaI-HindIII fragment that directs starvation-induced filament formation in MATa cells but not in MAT alpha cells. This 2.1-kb fragment contains a gene, MF alpha, with a small open reading frame encoding a pheromone precursor similar to the lipoprotein mating factors found in Saccharomyces cerevisiae, Ustilago maydis, and Schizosaccharomyces pombe. The ability of the MATa cells to express, process, and secrete the MAT alpha pheromone in response to starvation suggests similar mechanisms for these processes in both cell types. These results also suggest that the production of pheromone is under a type of nutritional control shared by the two cell types.

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

Molecular and Cellular Biology ◽

10.1128/mcb.1.6.522-534.1981 ◽

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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Regulation of STA1 gene expression by MAT during the life cycle of Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.9.9.3992-3998.1989 ◽

1989 ◽

Vol 9 (9) ◽

pp. 3992-3998

Author(s):

A M Dranginis

Keyword(s):

Saccharomyces Cerevisiae ◽

Mating Type ◽

Yeast Cells ◽

Specific Gene ◽

Activity Levels ◽

Sporulation Medium ◽

Mrna Levels ◽

Yeast Saccharomyces Cerevisiae ◽

Type Locus ◽

Diploid Cells

STA1 encodes a secreted glucoamylase of the yeast Saccharomyces cerevisiae var. diastaticus. Glucoamylase secretion is controlled by the mating type locus MAT; a and alpha haploid yeast cells secrete high levels of the enzyme, but a/alpha diploid cells produce undetectable amounts. It has been suggested that STA1 is regulated by MATa2 (I. Yamashita, Y. Takano, and S. Fukui, J. Bacteriol. 164:769-773, 1985), which is a MAT transcript of previously unknown function. In contrast, this work shows that deletion of the entire MATa2 gene had no effect on STA1 regulation but that deletion of MATa1 sequences completely abolished mating-type control. In all cases, glucoamylase activity levels reflected STA1 mRNA levels. It appears that STA1 is a haploid-specific gene that is regulated by MATa1 and a product of the MAT alpha locus and that this regulation occurs at the level of RNA accumulation. STA1 expression was also shown to be glucose repressible. STA1 mRNA was induced in diploids during sporulation along with SGA, a closely linked gene that encodes an intracellular sporulation-specific glucoamylase of S. cerevisiae. A diploid strain with a MATa1 deletion showed normal induction of STA1 in sporulation medium, but SGA expression was abolished. Therefore, these two homologous and closely linked glucoamylase genes are induced by different mechanisms during sporulation. STA1 induction may be a response to the starvation conditions necessary for sporulation, while SGA induction is governed by the pathway by which MAT regulates sporulation. The strain containing a complete deletion of MATa2 grew, mated, and sporulated normally.

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