scholarly journals MUTANTS OF THE KILLER PLASMID OF SACCHAROMYCES CEREVISIAE DEPENDENT ON CHROMOSOMAL DIPLOIDY FOR EXPRESSION AND MAINTENANCE

Genetics ◽  
1976 ◽  
Vol 82 (2) ◽  
pp. 273-285
Author(s):  
Reed B Wickner
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Elevated Temperature ◽  
Mating Type ◽  
Chromosomal Gene ◽  
Wild Type ◽  
Opposite Mating Type ◽  
Diploid Clone ◽  
Wild Type Phenotype ◽  
Chromosomal Genes ◽  
Killer Plasmid

ABSTRACT Mutants of the killer plasmid of Saccharomyecs cerevisiaehave been isolated that depend upon chromosomal diploidy for the expression of plasmid functions and for replication or maintenance of the plasmid itself. These mutants are not defective in any chromosomal gene needed for expression or replication of the killer plasmid.—Haploids carrying these mutant plasmids (called d for diploid-dependent) are either unable to kill or unable to resist being killed or both and show frequent loss of the plasmid. The wild-type phenotype (K+R+) is restored by mating the d plasmid-carrying strain with either (a) a wild-type sensitive strain which apparently has no killer plasmid; (b) a strain which has been cured of the killer plasmid by growth at elevated temperature; (c) a strain which has been cured of the plasmid by growth in the presence of cycloheximide; (d) a strain which has lost the plasmid because it carries a mutation in a chromosomal mak gene; or (e) a strain of the opposite mating type which carries the same d plasmid and has the same defective phenotype, indicating that the restoration of the normal phenotype is not due to recombination between plasmid genomes or complementation of plasmid or chromosomal genes.—Sporulation of the phenotypically K+R+ diploids formed in matings between d and wild-type nonkiller strains yields tetrads, all four of whose haploid spores are defective for killing or resistance or maintenance of the plasmid or a combination of these. Every defective phenotype may be found among the segregants of a single diploid clone carrying a d plasmid. These defective segregants resume the normal killer phenotype in the diploids formed when a second round of mating is performed, and the segregants from a second round of meiosis and sporulation are again defective.

scholarly journals DELETION OF MITOCHONDRIAL DNA BYPASSING A CHROMOSOMAL GENE NEEDED FOR MAINTENANCE OF THE KILLER PLASMID OF YEAST

Genetics ◽  
1977 ◽  
Vol 87 (3) ◽  
pp. 441-452
Author(s):  
Reed B Wickner
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitochondrial Dna ◽  
Mitochondrial Genome ◽  
Antimycin A ◽  
Chromosomal Gene ◽  
Wild Type ◽  
Suppressor Mutations ◽  
Induced Mutants ◽  
Chromosomal Genes ◽  
Killer Plasmid

ABSTRACT Strains of Saccharomyces cerevisiae carrying a 1.4 × 106 dalton double-stranded (ds) RNA in virus-like particles (the killer plasmid or virus) secrete a toxin that is lethal to strains not carrying this plasmid (virus). The mak10 gene is one of 24 chromosomal genes (called pets, mak1, mak2,…) that are needed to maintain and replicate the killer plasmid. We report here isolation of spontaneous and induced mutants in which the killer plasmid is maintained and replicated in spite of a defect in the mak10 gene. The bypass (or suppressor) mutations in these strains are in the mitochondrial genome. Respiratory deficiency produced by various chromosomal pet mutations, by chloramphenicol, or by antimycin A, does not bypass the mak10-1 mutation. Several spontaneous mak10-1 killer strains have about 12-fold more of the killer plasmid ds RNA than do wild-type killers. Although the absence of mitochondrial DNA bypasses mak10-1, it does not bypass pets-1, mak1-1, mak3-1, mak4-1, mak5-1, mak6-1, mak7-1, or mak8-1.

scholarly journals TWO CHROMOSOMAL GENES REQUIRED FOR KILLING EXPRESSION IN KILLER STRAINS OF SACCHAROMYCES CEREVISIAE

Genetics ◽  
1976 ◽  
Vol 82 (3) ◽  
pp. 429-442
Author(s):  
Reed B Wickner ◽  
Michael J Leibowitz
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Molecular Weight ◽  
Mutant Phenotype ◽  
Wild Type ◽  
Double Stranded Rna ◽  
Killer Strain ◽  
Chromosomal Genes ◽  
Complementation Groups ◽  
Killer Plasmid ◽  
Type Strains

ABSTRACT The killer character of yeast is determined by a 1.4 × 106 molecular weight double-stranded RNA plasmid and at least 12 chromosomal genes. Wild-type strains of yeast that carry this plasmid (killers) secrete a toxin which is lethal only to strains not carrying this plasmid (sensitives). —— We have isolated 28 independent recessive chromosomal mutants of a killer strain that have lost the ability to secrete an active toxin but remain resistant to the effects of the toxin and continue to carry the complete cytoplasmic killer genome. These mutants define two complementation groups, kex1 and kex2. Kex1 is located on chromosome VII between ade5 and lys5. Kex2 is located on chromosome XIV, but it does not show meiotic linkage to any gene previously located on this chromosome. —— When the killer plasmid of kex1 or kex2 strains is eliminated by curing with heat or cycloheximide, the strains become sensitive to killing. The mutant phenotype reappears among the meiotic segregants in a cross with a normal killer. Thus, the kex phenotype does not require an alteration of the killer plasmid. —— Kex1 and kex2 strains each contain near-normal levels of the 1.4 × 106 molecular weight double-stranded RNA, whose presence is correlated with the presence of the killer genome.

ESCAPE FROM MATING-TYPE INCOMPATIBILITY IN BISEXUAL (A + a) NEUROSPORA HETEROKARYONS

1975 ◽  
Vol 17 (3) ◽  
pp. 441-449 ◽  
Author(s):  
A. M. DeLange ◽  
A. J. F. Griffiths
Keyword(s):  
Mating Type ◽  
Loss Of Function ◽  
Wild Type ◽  
Opposite Mating Type ◽  
Type Locus ◽  
Heterokaryon Incompatibility ◽  
Recessive Mutant ◽  
Type Rate ◽  
Wild Type Rate ◽  
Incompatibility Locus

In Neurospora crassa, strains of opposite mating type generally do not form stable heterokaryons because the mating type locus acts as a heterokaryon incompatibility locus. However, when one A and one a strain, having complementing auxotrophic mutants, are placed together on minimal medium, growth may occur, although the growth is generally slow. In this study, escape from such slow growth to that at a wild type or near-wild type rate was observed. The escaped cultures are stable heterokaryons, mostly having lost the mating type allele function from one component nucleus, so that the nuclear types are heterokaryon compatible. Either A or a mating type can be lost. This loss of function has been attributed to deletion since only one nuclear type could be recovered in all heterokaryons except one, but deletion spanning adjacent loci has been directly demonstrated in a minority of cases. Alternatively when one component strain is tol and the other tol+ (tol being a recessive mutant suppressing the heterokaryon incompatibility associated with mating type), escape may occur by the deletion or mutation of tol+, also resulting in heterokaryon compatibility. An induction mechanism for escape is speculated upon.

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.

scholarly journals Mistranslating tRNA identifies a deleterious S213P mutation in the Saccharomyces cerevisiae eco1-1 allele

2020 ◽  
Author(s):  
Yanrui Zhu ◽  
Matthew D. Berg ◽  
Phoebe Yang ◽  
Raphaël Loll-Krippleber ◽  
Grant W. Brown ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Elevated Temperature ◽  
Amino Acid ◽  
Genetic Disease ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Standard Genetic Code ◽  
Gene Encoding ◽  
Chromatid Cohesion ◽  
Sensitive Allele

ABSTRACTMistranslation occurs when an amino acid not specified by the standard genetic code is incorporated during translation. Since the ribosome does not read the amino acid, tRNA variants aminoacylated with a non-cognate amino acid or containing a non-cognate anticodon dramatically increase the frequency of mistranslation. In a systematic genetic analysis, we identified a suppression interaction between tRNASerUGG, G26A, which mistranslates proline codons by inserting serine, and eco1-1, a temperature sensitive allele of the gene encoding an acetyltransferase required for sister chromatid cohesion. The suppression was partial with a tRNA that inserts alanine at proline codons and not apparent for a tRNA that inserts serine at arginine codons. Sequencing of the eco1-1 allele revealed a mutation that would convert the highly conserved serine 213 within β7 of the GCN5-related N-acetyltransferase core to proline. Mutation of P213 in eco1-1 back to the wild-type serine restored function of the enzyme at elevated temperature. Our results indicate the utility of mistranslating tRNA variants to identify functionally relevant mutations and identify eco1 as a reporter for mistranslation. We propose that mistranslation could be used as a tool to treat genetic disease.

scholarly journals tRNA thiolation links translation to stress responses in Saccharomyces cerevisiae

2015 ◽  
Vol 26 (2) ◽  
pp. 270-282 ◽  
Author(s):  
Jadyn R. Damon ◽  
David Pincus ◽  
Hidde L. Ploegh
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Endoplasmic Reticulum ◽  
Elevated Temperature ◽  
Heat Shock ◽  
Stress Responses ◽  
Wild Type ◽  
Trna Modifications ◽  
Shock Response ◽  
Tightly Coupled

Although tRNA modifications have been well catalogued, the precise functions of many modifications and their roles in mediating gene expression are still being elucidated. Whereas tRNA modifications were long assumed to be constitutive, it is now apparent that the modification status of tRNAs changes in response to different environmental conditions. The URM1 pathway is required for thiolation of the cytoplasmic tRNAs tGluUUC, tGlnUUG, and tLysUUU in Saccharomyces cerevisiae. We demonstrate that URM1 pathway mutants have impaired translation, which results in increased basal activation of the Hsf1-mediated heat shock response; we also find that tRNA thiolation levels in wild-type cells decrease when cells are grown at elevated temperature. We show that defects in tRNA thiolation can be conditionally advantageous, conferring resistance to endoplasmic reticulum stress. URM1 pathway proteins are unstable and hence are more sensitive to changes in the translational capacity of cells, which is decreased in cells experiencing stresses. We propose a model in which a stress-induced decrease in translation results in decreased levels of URM1 pathway components, which results in decreased tRNA thiolation levels, which further serves to decrease translation. This mechanism ensures that tRNA thiolation and translation are tightly coupled and coregulated according to need.

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.

scholarly journals [HOK], A NEW YEAST NON-MENDELIAN TRAIT, ENABLES A REPLICATION-DEFECTIVE KILLER PLASMID TO BE MAINTAINED

Genetics ◽  
1982 ◽  
Vol 100 (2) ◽  
pp. 159-174
Author(s):  
Reed B Wickner ◽  
Akio Toh-E
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Copy Number ◽  
Double Stranded Rna ◽  
Protein Toxin ◽  
Defective Mutant ◽  
Chromosomal Genes ◽  
Killer Plasmid ◽  
New Yeast ◽  
Lower Copy ◽  
Host Strains

ABSTRACT The K1 killer plasmid, [KIL-k1], of Saccharomyces cerevisiae is a 1.25 × 106 dalton linear double-stranded RNA plasmid coding for a protein toxin and immunity to that toxin. The [KIL-sd1] plasmid is a replication-defective mutant of [KIL-k1] that depends on one of the recessive chromosomal superkiller (ski  -) mutations for its maintenance (Toh-e and Wickner 1979). This report concerns a means by which [KIL-sd1] can be stably maintained in a SKI  + host. Strains carrying a plasmid we call [HOK] (helper of killer) stably maintain [KIL-sd1]. [HOK] segregates 4 [HOK]:0 in meiotic crosses and is efficiently transferred by cytoplasmic mixing (heterokaryon formation). [HOK] depends for its maintenance on the products of PET18, MAK3, and MAK10, three chromosomal genes needed to maintain [KIL-k1], but is independent of 10 other MAK genes and of MKT1. [HOK] is not mitochondrial DNA and is unaffected by agents which convert ψ+ strains to ψ-. [HOK] is also distinct from the previously described plasmids [URE3], 20S RNA, 2 µ DNA, and [EXL]. Strains lacking [HOK] consistently have a four-fold lower copy number of L double-stranded RNA than strains carrying [HOK].

scholarly journals Mutations affecting donor preference during mating type interconversion in Saccharomyces cerevisiae.

Genetics ◽  
1995 ◽  
Vol 139 (4) ◽  
pp. 1495-1510 ◽  
Author(s):  
K S Weiler ◽  
L Szeto ◽  
J R Broach
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Chromosome Segregation ◽  
Mating Type ◽  
Null Allele ◽  
Alpha Cells ◽  
Opposite Mating Type ◽  
Donor Preference ◽  
Donor Locus ◽  
Type Information ◽  
Selection Of

Abstract Homothallic strains of Saccharomyces cerevisiae can convert mating type from a to alpha or alpha to a as often as every generation, by replacing genetic information specifying one mating type at the expressor locus, MAT, with information specifying the opposite mating type. The cryptic mating type information that is copied and inserted at MAT is contained in either of two loci, HML or HMR. The particular locus selected as donor during mating type interconversion is regulated by the allele expressed at MAT. MATa cells usually select HML, and MAT alpha cells usually select HMR, a process referred to as donor preference. To identify factors required for donor preference, we isolated and characterized a number of mutants that frequently selected the nonpreferred donor locus during mating type interconversion. Many of these mutants were found to harbor chromosome rearrangements or mutations at MAT or HML that interfered with the switching process. However, one mutant carried a recessive allele of CHL1, a gene previously shown to be required for efficient chromosome segregation during mitosis. Homothallic strains of yeast containing a null allele of CHL1 exhibited almost random selection of the donor locus in a MATa background but were normal in their ability to select HMR in a MAT alpha background. Our results indicate that Chl1p participates in the process of donor selection and are consistent with a model in which Chl1p helps establish an intrinsic bias in donor preference.

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