<i>Saccharomyces cerevisiae</i>
            : A Diffusible Sex Factor

Duntze, W.; MacKay, Vivian L.; Manney, Thomas R.

doi:10.1126/science.168.3938.1472

Cited by 203 publications

(90 citation statements)

References 4 publications

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“…The morphological alteration noted is indistinguishable from the effect previously described by Duntze and coworkers (6) and attributed by them to the action of an extracellular factor produced by a strains on cells of the opposite mating type. The coincident inhibition of mating and change of cell shape were also observed in mixtures of fractionated cells taken from late-log-phase cultures.…”

Section: Mating Optimizationsupporting

confidence: 54%

Synchronous Mating in Yeast

Sena

Radin

Fogel

1973

Proc. Natl. Acad. Sci. U.S.A.

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Mating by cellular fusion is essential for segregational and recombinational analysis of nuclear or cytoplasmic inheritance in Saccharomyces cerevisiae. However, a detailed description of the mating reaction at the molecular level has not been established. In part, this reflects the inadequacy of current experimental methods for studying mating in yeast. In heterothallic yeasts mating occurs when cells with complementary mating types are mixed under appropriate conditions. The conjugation reaction (1, 2) then progresses as a sequence of ordered events that can be subdivided on a gross visual and cytological basis into three principal phases: (a) mating initiation, which involves conjugant pairing and sexual agglutination, (b) cell fusion (plasmogamy), and (c) nuclear fusion (karyogamy). The completed reaction sequence yields a morphologically distinct zygote that can, depending on nutritional conditions, either proliferate mitotically or enter into meiotic development and ascospore formation.A prerequisite for studies aimed at understanding mating and sexuality is the ability to obtain sizable, homogeneous cell populations that undergo synchronous mating under defined conditions. We report here our initial studies on mating and a protocol for producing highly efficient synchronously mating populations of yeast in liquid minimal medium. From these populations, purified zygote suspensions are readily isolated in quantities suitable for extensive genetic and biochemical analyses. MATERIALS AND METHODSStrains. Two heterothallic haploid strains of S. cerevisiae containing complementary leucine and tryptophan markers were isolated as single spore clones: 5032A a leu2-27 tryl-1 and 5032B a leul-12 trp4. Growth Mating Procedure. The single cell fraction obtained from the zonal rotor was centrifuged to remove the sorbitol, resuspended in 100 ml of YNB to a density of 6 to 10 X 106 cells per ml in a 500-ml Erlenmyer flask, and shaken at 300. The beginning of incubation is defined as zero time for the mating reaction. A cell sample was immediately withdrawn from the mating mixture to determine the ratio of a and a unbudded cells obtained from the rotor. Samples taken throughout the first 180 min of the mating reaction were fixed immediately with an equal volume of 3.7% formaldehyde in 0.05 M phosphate buffer (pH 7.0).Zygote Isolation. After 120 and 150 min of incubation, 30-40 ml of the cell suspension was harvested by centrifugation, resuspended in l ml of water, sonicated for 5 sec, and then layered onto a 25-ml linear 10-30% sorbitol gradient. The gradient was centrifuged at 1000 rpm in a Sorvall HB4

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Section: Mating Optimizationsupporting

confidence: 54%

Synchronous Mating in Yeast

Sena

Radin

Fogel

1973

Proc. Natl. Acad. Sci. U.S.A.

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“…Our findings indicate that U. maydis dikaryons respond to two signals: the mating pheromones and a putative plant signal (see below). In S. cerevisiae, the signal for the pseudohyphal pathway is clearly not the mating pheromones (Liu et al 1993), which are not produced in a/~ cells (Duntze et al 1970).…”

Section: Fuz7 Is Required For Several A-dependent Processesmentioning

confidence: 99%

Identification of fuz7, a Ustilago maydis MEK/MAPKK homolog required for a-locus-dependent and -independent steps in the fungal life cycle.

Banuett¹,

Herskowitz²

1994

Genes Dev.

175

143

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Ustilago maydis is a plant pathogenic Basidiomycete fungus that exhibits dimorphismuit has a haploid, yeast-like phase and a dikaryotic, filamentous phase that is pathogenic. Establishment and maintenance of these two forms are controlled by two mating type loci, a and b. The a locus is thought to govern fusion of haploid cells to form a dikaryon and is also required for filamentous growth of the dikaryon. It encodes two components of a pheromone response pathway: pheromones and receptors. We report the identification of the U. maydis fuz7 gene, which codes for a putative dual specificity serine/threonine tyrosine kinase of the MAP kinase kinase (MAPKK/MEK) family, by homology with other members of the family. Analysis of mutants deleted for fuz7 shows that it participates in different facets of the life cycle: It is necessary for a-locus-dependent processes, such as conjugation tube formation, filament formation, and maintenance of filamentous growth, and for a-locus-independent processes, such as tumor induction and teliospore germination, fuz7 is the first U. maydis gene distinct from the b locus required for fungal pathogenicity. We propose that fuz7 is involved in at least two pathways, one of which responds to the pheromones coded by the a locus and the other to putative signals from the plant.

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“…The Escherichia coli and Neurospora crassa enzymes appear to convert indoleglycerol phosphate to tryptophan (the sum of reactions A and B) without releasing free indole from the enzymc [2,3]. This mechanism predominates also with the Xaccharomyces cerevisiae enzyme, but significant amounts of free indole are also formed [4]. In bacteria tryptophan synthase can bc dissociated Abbreviations.…”

mentioning

confidence: 99%

“…Genetic studies have been carried out [4] with the fungus S. cerevisiae pertainent to the structure of tryptophan synthase and its reaction mechanism. It was shown also by Manney 17 J and by Katsunuma et al [S] that the yeast enzyme is inactivated in an enzymatic process.…”

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confidence: 99%

Tryptophan Synthase from Yeast

Wolf

Hoffmann

1974

European Journal of Biochemistry

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Tryptophan synthase was purified from Xaccharomyces cerevisiae with the aid of affinity chromatography up to a specific activity of 525 units/g protein. Electrophoresis on polyacrylamide gel showed one main protein component. Gel filtration on Sephadex G-150 indicates an average molecluar weight of 143 000. Dissociation with dodecylsulfate followed by dodecylsulfate-acrylamide gcl electrophoresis leads to the conclusion that the enzyme is probably cornposed of four subunits of equal size. The Km-values with respect to its substrates indoleglycerol phosphate, indole and L-serine were estimated. L-Serine was shown to increase the K,-value for indoleglycerol phosphate about 7-fold. Several indole analogues inhibit the tryptophan synthase reaction (B) (i.e. indole + serine --f tryptophan); indoleacrylic acid is the most effective. The pH-optimum of tryptophan synthase is around 6.9 to 7.0. Sulfhydryl modification by 5,5'-dithiobis(2-nitrobenzoic acid) and p-chloromercuriphenylsulfonic acid results in a large loss of tryptophan synthase activity, which cannot be prevented by the substrates L-serine and indole. The tryptophan synthase inactivating system from yeast affects the catalysis of both reactions (A) (i.e. indoleglycerol phosphate + indole + glyceraldehyde 3-phosphate) and (B).Tryptophan synthase (L-serine hydrolyase, adding indole) catalyzes the final reaction in tryptophan hiosynthesis :In all microorganisms in which i t has been studied, the enzyme is also capable of catalyzing the following two half reactions [ 11 :Indole-glycerol-P -+ Indole + n-glyceraldehyde-3-P.Pyridoxal phosphate is required as a coenzyme in reactions (AB) and (B). The Escherichia coli and Neurospora crassa enzymes appear to convert indoleglycerol phosphate to tryptophan (the sum of reactions A and B) without releasing free indole from the enzymc [2,3]. This mechanism predominates also with the Xaccharomyces cerevisiae enzyme, but significant amounts of free indole are also formed [4]. In bacteria tryptophan synthase can bc dissociated Abbreviations. 5,5'-Dithiobis(2-nitrobenzoic acid), Nbs,; p-chloromercuriphenylsulfonic acid, pCI-HgBzOH.Enzymes. Tryptophan synthase (or r,-serine hydrolyase (adding indole) (EC 4.2.1.20). __.-readily and reversibly into two distinct components designated A and B [l], the A protein catalyzing reaction (A), the B protein reaction (B). The rates of reactions catalyzed by the individual components, however, are, in general, much lower than that catalyzed by the AB complex. In contrast, the enzyme from the fungus Neurospora is not dissociable and acts only as the whole unit, though i t is composed of two pairs of polypeptide chains [5,6].Genetic studies have been carried out [4] with the fungus S. cerevisiae pertainent to the structure of tryptophan synthase and its reaction mechanism. It was shown also by Manney 17 J and by Katsunuma et al. [S] that the yeast enzyme is inactivated in an enzymatic process. The inactivating system consists of two proteolytic enzymes and associated inhibitors [9,9a,Bb]. Th...

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Saccharomyces cerevisiae : A Diffusible Sex Factor

Cited by 203 publications

References 4 publications

Synchronous Mating in Yeast

Synchronous Mating in Yeast

Identification of fuz7, a Ustilago maydis MEK/MAPKK homolog required for a-locus-dependent and -independent steps in the fungal life cycle.

Tryptophan Synthase from Yeast

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