Homothallic Strains Research Articles

MUTANTS SHOWING HETEROTHALLISM FROM A HOMOTHALLIC STRAIN OF SACCHAROMYCES CEREVISIAE

Mutants defective in mating-type conversion were isolated from ascospores of a perfect homothallism strain having the HO HMRaHMLalpha genotype. Eighty mutants, including 11 temperature-dependent mutants showing a or alpha mating potency, were isolated from 10,050 colonies derived from spores mutagenized with ethyl methanesulfonate. Of the 80 mutants, 48 were tested by crossing with an ho HMRaHMLalpha heterothallic and an HO HMRaHMLalpha homothallic strain as standards. The results allowed their division into seven classes. Mutants in the first class were due to mutation of HO to an ho allele. The second class of mutants, csm, lacked the HO function as the result of a mutation unlinked with the HO locus. Mutants in the third and fourth classes were defective in HMRa and HMLalpha gene functions, respectively. Mutants in the fifth and sixth classes were due to a mutation at the mating-type locus showing insensitivity to the function of the homothallic genes. Mutants in the seventh class lacked the expression of mating type as the result of a mutation unlinked to the mating-type locus. All the temperature-dependent mutants lost the mating potency at permissive temperature (25 degrees ). One belonging to the last class of mutants was inferred to be a mutation at a locus necessary for the expression of a mating type at restricted temperature (35 degrees ).

Duplicated genes producing transposable controlling elements for the mating-type differentiation in Saccharomyces cerevisiae.

Mutation of the two homothallic genes, HML alpha/HMLa and HMRa/HMR alpha, in homothallic strains of Saccharomyces cerevisiae was studied. Of 11 mutants of the HML alpha gene, eight were due to a phenotypic mutation from HML alpha to HMLa, i.e., a mutation causing a change in function of the original HML allele to that of the other HML allele (functional mutation), and three were due to a defective mutation at the HML alpha gene, i.e., a mutation causing a nonfunctional allele (nonfunctional mutation). All 14 mutants of the HMRa gene, on the other hand, were due to a phenotypic mutation from HMRa to HMR alpha i.e., a functional mutation. Phenotypic reverse mutations, i.e., HMLa to HML alpha and HMR alpha to HMRa, were also observed in the cultivation of EMS (ethyl methanesulfonate) treated spores having the HO HMR alpha HMLa genotype. Mutation from heterothallic cells to homothallism was observed in a nonfunctional mutant of the HML alpha gene, by mutagenesis with EMS, but not in the functional mutants of the HML alpha and HMRa genes or in the authentic strains having the alpha HO HMR alpha HML alpha (alpha Hp) and a HO HMRa HMLa (a Hq) genotypes. These observations suggest that the functional mutation is not caused by the direct mutation from a homothallic allele to the opposite, but by replacement of a transposable genic element produced from a homothallic locus with a region of a different homothallic locus. These observations also support the controlling-element model and the cassette model, which have been proposed to explain the mating-type differentiation by the homothallic genes.

A MUTATION THAT PERMITS THE EXPRESSION OF NORMALLY SILENT COPIES OF MATING-TYPE INFORMATION IN SACCHAROMYCES CEREVISIAE

Studies of heterothallic and homothallic strains of Saccharomyces cerevisiae have led to the suggestion that mating-type information is located at three distinct sites on chromosome 3, although only information at the mating-type (MAT) locus is expressed (Hicks, Strathern and Herskowitz, 1977). We have found that the recessive mutation cmt permits expression of the normally silent copies of mating-type information at the HMa and HM alpha loci. In haploid strains carrying HMa and HM alpha, the cmt mutation allows the simultaneous expression of both a and alpha information, leading to a nonmating ("MATa/MAT alpha") phenotype. The effects of cmt can be masked by changing the mating-type information at HMa or HM alpha. For example, a cell of genotype MATa hma HM alpha cmt has an a mating type, while a MAT alpha hma HM alpha cmt strain is nonmating. Expression of mating-type information at the HM loci can correct the mating and sporulation defects of the mata* and mat alpha 10 alleles. Meiotic segregants recovered from cmt/cmt diploids carrying the mat mutations demonstrate that these mutants are not "healed" to normal MAT alleles, as is the case in parallel studies using the homothallism gene HO.--All of the results are consistent with the notion that the HMa and hm alpha alleles both code for alpha information, while HM alpha and hma both code for a information. The cmt mutation demonstrates that these normally silent copies of mating-type and sporulation information can be expressed and that the information at these loci is functionally equivalent to that found at MAT. The cmt mutation does not cause interconversions of mating-type alleles at MAT, and it is not genetically linked to MAT, HMa, HM alpha or HO. In cmt heterozygotes, cmt becomes homozygous at a frequency greater than 1% when the genotype at the MAT locus is mata*/MAT alpha or mat alpha 10/MATa.

The sexual process in several heterogamous Chlamydomonas strains in the subgenus Pleiochloris

Summary A soil sample from the Zimbabwe region of Rhodesia yielded a number of clones of Chlamydomonas. All the clonal isolates belong to the subgenus Pleiochloris, they exhibit homothallic and heterogamous sexual reproduction, but they differ in their zygote wall ornamentation. Male gametes are differentiated by successive cleavage of the protoplast to form naked biflagellated sperm. Female gametes are formed by the escape of the protoplast of a vegetative cell from its enclosing cell wall. The clones forming smooth unornamented zygotes are described as a new species Chlamydomonas zimbabwiensis. In this species the investigation of the sexual process is facilitated by three distinct strains, a homothallic strain and two subheterothallic strains. A sexual hormone has been isolated and partically characterized from sperm filtrates which cause vegetative cells to escape from their cell walls and to function as naked female gametes. Four other strains having distinct zygote wall ornamentations appear to be interfertile and respond to the sexual hormone of C. zimbabwiensis as well. Cytological preparations of these strains as well as others in the subgenus Pleiochloris demonstrate the presence of large chromosomes which permit reasonably accurate counts.

The mating system of a homothallic fission yeast

Exposed to iodine vapors, colonies of a homothallic strain of Schizosaccharomyces pombe were of two classes: P, with many black streaks, and d, with scarcely any. Contiguous P and d colonies, but not contiguous P colonies nor contiguous d colonies, gave the iodine junction reaction, a black line along the common boundary of two colonies. Neither class could be purified. On replating, a P colony gave rise to a P plate, which contained mostly P but also d colonies; a d colony gave rise to a d plate, which contained mostly d but also P colonies. The P/d colony ratio of a fresh isolate (if isolated as a P colony) was very high or (if isolated as a d conoly) very low. It fell, if initially high, or rose, if initially low, on subsequent replatings of the same isolate. Maintained for many generations, an isolate attained a fairly constant P/d colony ratio that was less than unity. Tetrad analysis showed 2:2 segregation of the classes. We conclude that a homothallic clone is a mixture of two types of cells: P, which gives rise to a P colony, and d, to a d colony. The two types are sexually complementary and interconvertible. The rate of intercoversion of P to d exceeds that of d to P by a factor of about 2.

CULTURE STUDY OF REPRODUCTIVE CYCLES AND SYSTEMATICA IN MOUGEOTIA TRANSEAUI (CHLOROPHYTA)1,2

ABSTRACT Asexual and sexual reproductive cyries are described and illustrated for a cultured homothallic strain of Mougeotia transeaui Collins. Reproduction is by aplanospores and zygospores. Aplanospore formation precedes zygospore formation and continues longer with regreening of older cultures occurring regularly from precocious germination of aplanospores. Aplanospores typically form when most of the protoplast moves into the central swollen region of a cell and two new cross walls form to delimit an aplanosporangium. Conjugation is scalariform without the movement and fusion of well‐organized gametes. During zygospore maturation, three new cross walls form in the receptive gametangium and conjugation tube to produce a zygosporangium.

Germination and outgrowth of Schizosaccharomyces pombe ascospores isolated by Urografin density gradient centrifugation

A simple method for the isolation of single ascospores of the fission yeast Schizosaccharomyces pombe was examined. Single spores in the 7-day-old sporulating culture of a homothallic strain were separated from remaining vegetative cells by isopycnic centrifugation in the linear gradient from 10 to 60% of Urografin solution at 700 X g for 20 min. Protein content of isolated spores was very low as compared with that of vegetative cells. The isolated spores germinated through the following steps when cultured in a liquid medium at 25--35 degrees C; loss of refractility (darkening) under a phase-contrast microscope, spherical growth (swelling), emergence of germ tubes, elongation of germ tubes, cell plate formation, and cell separation. The absorbance at 650 nm of the spore suspension initially decreased, accompanied by darkening of spores, and then increased with spherical growth. The germination rate of isolated spores reached almost 100%.

Sexual mutants of a heterothallic Mucor species, Mucor pusillus

The Mucorales consist of both homothallic and heterothallic species. Both types can be found in the same species but this is not a common phenomenon. A few homothallic strains are found in the predominantly heterothallic species, Mucor pusillus . It was interesting to see how readily homothallic mutants could be induced in this species. Spores of CBS 253.53(+) were irradiated with gamma rays. Homothallic and (−) mutants were isolated. An albino (+) and a yellowish-brown (+) mutant were also found. The two pigment mutants were further mutated, and again both (−) and homothallic mutants were isolated. Morphological characteristics of M. pusillus such as branching of sporangiophores, size of sporangia, and height of turf are retained by the mutants. The only obvious change is the color of the pigment of some of them. Like the original strain, they grow in the absence of thiamine. The development, structure, and size of the zygospores are typical of M. pusillus . The dark pigment of the zygospores is lacking in the albino homothallic and in crosses between albino (+) and albino (−), but in all other cases the characteristic color is retained. Homothallic mutants which otherwise are identical to their parent are readily induced by mutation of a heterothallic strain. The mere presence or absence of zygospores cannot be used as a classification criterion in this species which contains both hetero- and homothallic members.

Genetic studies of ability to ferment starch in Saccharomyces: Gene polymorphism

Genetic studies were made on the genes determining the ability to ferment starch (STA) in five strains of S. diastaticus different in origins, strain IFO 1046, IFO 1015, Y 8, Y 13 and C1372. With the heterothallic strain IFO 1046, the genes STA1 and STA2 were separated by single spore cultures and crosses to S. cerevisiae. Strain Y 8 did not sporulate, but the progeny having gene STA3 was isolated from the hybrid between the respiratory deficient mutant of strain Y 8 and S. cerevisiae obtained by the minimal plate mating technique. Strain IFO 1015 was the homothallic strain. The progeny having gene STA4 was isolated from the hybrid between IFO 1015 and S. cerevisiae. Strain Y 13 did not sporulate, but the progeny having gene STA5 was isolated according to the same technique as applied to strain Y 8. Haploid strain C1372 retained the gene STA6. The genes STA1 and STA4, and STA3, STA5 and STA6 were shown to be not separable by tetrad analysis.

A NEW GENE AFFECTING THE EFFICIENCY OF MATING-TYPE INTERCONVERSIONS IN HOMOTHALLIC STRAINS OF SACCHAROMYCES CEREVISIAE

Homothallic strains of Saccharomyes cerevisiae are able to switch efficiently from one mating genotype to another. From a single haploid spore arise both a and mating type cells, which then self-mate to produce a colony consisting almost exclusively of nonmating a/ diploid cells. We have isolated a mutant homothallic strain that gives rise to colonies that show bisexual mating behavior. The mating reaction is always asymmetric, that is, in some colonies a mating is much stronger than mating, while others show greater than a mating.-This mating phenotype arises from the presence of three cell types in a colony: some a/ nonmating diploids and an unequal number of a and haploid cells. The predominant haploid type is that of the original cell that gives rise to the colony. This mixture of cell types arises from a very reduced efficiency of homothallic mating-type interconversions in the mutant strain.-The mutation, designated switch (swi1-1), behaves as a single genetic locus. The mutation is centromere linked, but not linked to the mating type locus or to any of the homothallism genes: HO, HMa and HM. The switch mutation does not affect the efficiency of self-mating, but rather directly affects the frequency of interconversion of mating types.

Inhibiton by sulfanilamide of sporulation in Saccharomyces cerevisiae.

The antimetabolite sulfanilamide inhibits sporulation in Saccharomyces cerevisiae strain AP1. Cells exposed to sulfanilamide at various times during the sporulation process become progressively insensitive to the drug, although accumulation of sulfanilamide by the cells increases with time. Vegetative growth of AP1 is practically unaffected by sulfanilamide; pregrowth of the cells in the presence of the drug does not prevent sporulation. Thus, inhibition is confined to the meiotic phase of the cell cycle. Sensitivity to sulfanilamide is independent of pH. Increasing the time cells are exposed to sulfanilamide results in a progressive reduction of ascus formation; however, the inhibition is reversible since sporulation can occur in cells exposed to the drug for greater than 24 h. The drug arrests the cells at a point before commitment to sporulation, since yeast cells exposed to sulfanilamide for 12 h do not complete the sporulation process when returnedto vegetative medium, but resume mitotic growth instead. Meiotic nuclear division is largely prevented by sulfanilamide, and synthesis of RNA and protein is severely retarded. DNA synthesis is inhibited up to 50%; glycogen synthesis is approximately 90% inhibited. Other yeast strains showed varying sensitivity to sulfanilamide; homothallic strains were generally less affected.

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

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.

CORRIGENDA

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 alpha 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 alpha mating type locus can be converted to a functional a locus and subsequently to a functional alpha 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 alpha information, and lead to the cassette model for mating type interconversion. In this model, HMa and hmalpha loci are blocs of unexpressed alpha regulatory information, and HMalpha and hma 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.

Genetic analysis of a cross between two homothallic strains ofPhysarum polycephalum

SUMMARYThe homothallic amoebal clones ofPhysarum polycephalumare of potential use in understanding the developmental genetics of this organism. Such an application requires that complementation and recombination analysis be possible between pairs of homothallic clones. This paper is a report of the formation of mixed plasmodia by pairs of homothallic amoebal clones. In order to detect such mixed plasmodia use was made of two marker genes involved in plasmodial fusion,fusA andfusB. Sporulation of a mixed plasmodium formed from two homothallic (delayed) amoebal clones yielded progeny amoebae which were genetically recombinant. It is deduced from the ratios of various genotypes in these progeny clones that the mixed plasmodium was diploid and that meiosis was associated with sporulation. There is therefore no impediment to the use of the homothallic strains for genetical analysis. The progeny amoebal clones were observed to be showing segregation for the characters homothallic (rapid) and homothallic (delayed). This observation, taken together with other related observations, suggests that the homothallic (delayed) character is produced by mutation of the homothallic (rapid) character. The rare plasmodia formed by a homothallic (delayed) amoebal clone are the result of reversion of this mutation. Amoebal clones of the homothallic (delayed) type are therefore developmental mutants unable to perform the differentiation from amoeba to plasmodium.

The effect of aneuploidy on meiotic crossing over and segregation in yeast.

SUMMARYMeiotic chromosome associations in homothallic strains ofS. cerevisiaewere investigated by analysis of meiotic spore colonies and crossing over in +/+/− trisomics. The segregants of these aneuploids produce phenotypically distinguishable tetrasomic spore colonies. The data indicate that trivalent associations occur with a high frequency in trisomics of chromosome XVII and that the frequency of second division segregation is markedly increased over that found in the normal dipoid.

MATING-TYPE MUTATIONS IN SCHIZOSACCHAROMYCES POMBE: ISOLATION OF MUTANTS AND ANALYSIS OF STRAINS WITH AN h - or h + PHENOTYPE

Mutants defective in various steps of the sexual cycle have been isolated from homothallic strains of Schizosaccharomyces pombe by Bresch, Müller and Egel (1968). These mutants include heterothallic h(+) and h(-) strains. We have isolated additional h(+) and h(- ) mutants from homothallic strains. Those mutants which are due to mutations in the mating-type region were analyzed in detail. Our results show that the mating-type gene mat2 not only has a function in copulation and meiosis, but that it also regulates the formation of the map1 gene product (map1 is a mating-type auxiliary gene). Some of the h( -) mutants have lost only one of the three functions while others are defective in at least two, and perhaps all three, functions. Further, we show that the mat1(-) allele of h(90) strains can mutate to mat1(+) but that mutations in mat2 appear to affect the mutational behavior of mat1. Finally, we describe a new inactive mating-type allele, mat2*, which is different from mat2(0) in that it can mutate to mat2(+).

The genetic instabilities of the mating type locus in fission yeast.

Certain genetic instabilities of the "mating type locus" in the yeast Schizosaccharomyces pombe are interpreted in terms of transposition: Homothallic strains are characterized by two adjacent mating type genes (mat1-mat2+) with sexually complementary functions. One of these genes (mat2+) is able to duplicate itself, and the duplicated copy maps at the position of mat1-. The former function of mat1-is lost (owing to insertion), and only becomes reactivated when the inserted sequence (mat1+) is again excised. Analyses of analogous instabilities expressed by the partially defective mutation mat2+ -B102 have substantiated this transposition scheme. Homothallism is acribed to alternate and mutually exclusive activation of mat1- or mat2+ genes.

Promotion of sporulation by caffeine pretreatment in Saccharomyces cerevisiae. II. Changes in ribonuclease activity during sporulation.

Changes in RNase activity during sporulation of a homothallic diploid strain of Saccharomyces cerevisiae were measured in caffeine-treated and non-treated cells. 1. In caffeine-treated cells soon after the transfer to the sporulation medium a significant increase in RNase activity was observed; in control cells the rise of RNase activity was less and started after a lag period of 5 h. The final activity of RNase activity was about twice as high in caffeine-treated cells as in control cells. 2. Increase in RNase activity during sporulation was sensitive to cycloheximide in control cells, but insensitive in caffeine-treated cells. 3. RNases from vegetative cells and from sporulating ones are different in their Km values. Relation of the changes in RNase activity to premeiotic DNA synthesis is discussed.

A new type of mutation in Schizosaccharomyces pombe: vegetative iodine reaction.

Colonies of Schizosaccharomyces pombe that contain ascospores (e.g., colonies of homothallic strains) turn black after treatment with iodine vapors. Heterothallic strains of S. pombe normally do not show this reaction. In experiments with the latter strains we found mutants which exhibit a positive iodine reaction though they do not contain ascospores. This phenotype is due to mutations in a new gene, vir1 (vegatative iodine reaction). The vir1 locus is not linked with the mating-type genes.--Strains of mating-type h-S are known not to give any spontaneous mating-type mutations. Mating-type mutations were also not found after treatment with nitrous acid.

On Homo- and Heterothallism inSchizosaccharomyces Pombe

The mating-type system of Schizosaccharomyces pombe has previously been studied by Leupold. In his culture, Leupold found heterothallic (h+, h-) and homothallic (h90) strains; the various strains s...