MAPPING AND GENE CONVERSION STUDIES WITH THE STRUCTURAL GENE FOR ISO-1-CYTOCHROME C IN YEAST

ABSTRACT We have investigated the order of the four genes cyc1, rad7, SUP4, and cdc8 which form a tightly linked cluster on the right arm of chromosome X in the yeast Saccharomyces cerevisiae. Crossing over and coconversion data from tetrad analysis established the gene order to be centromere–cyc1–rad7–SUP4. Also cdc8 appeared to be distal to SUP4 on the basis of crossovers that were associated with conversion of SUP4. The frequencies of recombination and the occurrence of coconversions suggest that these four genes are contiguous or at least nearly so. Gene-conversion frequencies for several cyc1 alleles were studied, including cyc1–1, a deletion of the whole gene that extends into the rad7 locus. The cyc1–1 deletion was found to be capable of conversion, though at a frequency some fivefold less than the other alleles studied, and both 3:1 and 1:3 events were detected. In general 1:3 and 3:1 conversion events were equally frequent at all loci studied, and approximately 50% of conversions were accompanied by reciprocal recombination for flanking markers. The orientation of the cyc1 gene could not be clearly deduced from the behavior of the distal marker SUP4 in wild-type recombinants that arose from diploids heteroallelic for cyc1 mutations.

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Homologous recombination involving a large heterology in Escherichia coli.

Genetics ◽

10.1093/genetics/119.4.759 ◽

1988 ◽

Vol 119 (4) ◽

pp. 759-769

Author(s):

K Yamamoto ◽

N Takahashi ◽

H Yoshikura ◽

I Kobayashi

Keyword(s):

Escherichia Coli ◽

Gene Conversion ◽

Kanamycin Resistance ◽

Coli Strain ◽

The Other ◽

Reca Gene ◽

Inverted Repeats ◽

Crossing Over ◽

Wild Type ◽

Parental Allele

Abstract Recombination between two different deletion alleles of a gene (neo) for neomycin and kanamycin resistance was studied in an Escherichia coli sbcA- recB-C- strain. The two homologous regions were in an inverted orientation on the same plasmid molecule. Kanamycin-resistant plasmids were selected and analyzed. The rate of recombination to form kanamycin-resistant plasmids was decreased by mutations in the recE, recF and recJ genes, but was not decreased by a mutation in the recA gene. It was found that these plasmids often possessed one wild-type kanamycin-resistant allele (neo+) while the other neo allele was still in its original (deletion) form. Among kanamycin-resistant plasmids with one wild-type and one parental allele it was often found that the region between the inverted repeats had been flipped (turned around) with respect to sites outside the inverted repeats. These results were interpreted as follows. Gene conversion, analogous to gene conversion in eukaryotic meiosis, is responsible for a unidirectional transfer of information from one neo deletion allele to the other. The flipping of the region between the inverted repeats is interpreted as analogous to the crossing over associated with gene conversion in eukaryotic meiosis. In contrast with a rec+ strain, these products cannot be explained by two rounds of reciprocal crossing over involving a dimeric form as an intermediate. In the accompanying paper we present evidence that gene conversion by double-strand gap repair takes place in the same E. coli strain.

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Oxygen and haem regulate the synthesis of peroxisomal proteins: catalase A, acyl-CoA oxidase and Pex1p in the yeast Saccharomyces cerevisiae; the regulation of these proteins by oxygen is not mediated by haem

Biochemical Journal ◽

10.1042/bj3500313 ◽

2000 ◽

Vol 350 (1) ◽

pp. 313-319 ◽

Cited By ~ 7

Author(s):

Marek SKONECZNY ◽

Joanna RYTKA

Keyword(s):

Saccharomyces Cerevisiae ◽

Mitochondrial Genome ◽

Oxygen Sensor ◽

Transcriptional Factor ◽

The Other ◽

Wild Type ◽

Yeast Saccharomyces Cerevisiae ◽

Regulatory Factors ◽

Genes Encoding ◽

Acyl Coa Oxidase

Saccharomyces cerevisiae genes related to respiration are typically controlled by oxygen and haem. Usually the regulation by these factors is co-ordinated; haem is indicated as the oxygen sensor. However, the responsiveness of peroxisome functions to these regulatory factors is poorly understood. The expression of CTA1, POX1 and PEX1 genes encoding the peroxisomal proteins catalase A, acyl-CoA oxidase and Pex1p peroxin respectively was studied under various conditions: in anaerobiosis, in the absence of haem and in respiratory incompetence caused by the lack of a mitochondrial genome (ρ0). The influence of haem deficiency or ρ0 on peroxisomal morphology was also investigated. Respiratory incompetence has no effect on the expression of CTA1 and POX1, whereas in the absence of haem their expression is markedly decreased. The synthesis of Pex1p is decreased in ρ0 cells and is decreased even more in haem-deficient cells. Nevertheless, peroxisomal morphology in both these types of cell does not differ significantly from the morphology of peroxisomes in wild-type cells. The down-regulating effect of anoxia on the expression of CTA1 and POX1 is even stronger than the effect of haem deficiency and is not reversed by the addition of exogenous haem or the presence of endogenous haem. Moreover, neither of these genes responds to the known haem-controlled transcriptional factor Hap1p. In contrast with the other two genes studied, PEX1 is up-regulated in anaerobiosis. The existence of one or more novel mechanisms of regulation of peroxisomal genes by haem and oxygen, different from those already known in S. cerevisiae, is postulated.

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Depletion of H2A-H2B Dimers in Saccharomyces cerevisiae Triggers Meiotic Arrest by Reducing IME1 Expression and Activating the BUB2-Dependen Branch of the Spindle Checkpoint

Genetics ◽

10.1093/genetics/164.4.1333 ◽

2003 ◽

Vol 164 (4) ◽

pp. 1333-1344

Author(s):

Sean E Hanlon ◽

David N Norris ◽

Andrew K Vershon

Keyword(s):

Saccharomyces Cerevisiae ◽

Spindle Checkpoint ◽

The Other ◽

Histone H2a ◽

Wild Type ◽

Meiotic Arrest ◽

Yeast Saccharomyces Cerevisiae ◽

Chromosomal Alterations ◽

Checkpoint Pathway ◽

Failure To Progress

Abstract In the yeast Saccharomyces cerevisiae, diploid strains carrying homozygous hta1-htb1Δ mutations express histone H2A-H2B dimers at a lower level than do wild-type cells. Although this mutation has only minor effects on mitotic growth, it causes an arrest in sporulation prior to the first meiotic division. In this report, we show that the hta1-htb1Δ mutant exhibits reduced expression of early and middle-sporulation-specific genes and that the meiotic arrest of the hta1-htb1Δ mutant can be partially bypassed by overexpression of IME1. Additionally, deletions of BUB2 or BFA1, components of one branch of the spindle checkpoint pathway, bypass the meiotic arrest. Mutations in the other branch of the pathway or in the pachytene checkpoint are unable to suppress the meiotic block. These observations indicate that depletion of the H2A-H2B dimer blocks sporulation by at least two mechanisms: disruption of the expression of meiotic regulatory genes and activation of the spindle checkpoint. Our results show that the failure to progress through the meiotic pathway is not the result of global chromosomal alterations but that specific aspects of meiosis are sensitive to depletion of the H2A-H2B dimer.

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Cloning, Disruption and Chromosomal Mapping of Yeast LEU3, a Putative Regulatory Gene

Genetics ◽

10.1093/genetics/115.1.91 ◽

1987 ◽

Vol 115 (1) ◽

pp. 91-99

Author(s):

Paula R G Brisco ◽

Thomas S Cunningham ◽

Gunter B Kohlhaw

Keyword(s):

Regulatory Gene ◽

Chromosomal Mapping ◽

Structural Genes ◽

Wild Type ◽

Tetrad Analysis ◽

Yeast Saccharomyces Cerevisiae ◽

The Right ◽

Cloned Gene ◽

Isopropylmalate Dehydrogenase ◽

His3 Gene

ABSTRACT The LEU3 gene of the yeast Saccharomyces cerevisiae, which is involved in the regulation of at least two LEU structural genes (LEU1 and LEU2), has been cloned by complementation of leu3 mutations and shown to reside within a 5.6-kb fragment. Transformation of leu3 mutants with LEU3-carrying multicopy plasmids restored normal, leucine-independent growth behavior in the recipients. It also restored approximately wild-type levels of isopropylmalate isomerase (LEU1) and β-isopropylmalate dehydrogenase (LEU2), which were strongly reduced when exogenous leucine was supplied. Strains containing a disrupted leu3 allele were constructed by deleting 0.7-kb of LEU3 DNA and inserting the yeast HIS3 gene in its place. Like other leu3 mutants, these strains were leaky leucine auxotrophs, owing to a basal level of expression of LEU1 and LEU2. Southern transfer and genetic analyses of strains carrying a disrupted leu3 allele demonstrated that the cloned gene was LEU3, as opposed to a suppressor. Disruption of LEU3 was performed also with a diploid and shown to be nonlethal by tetrad analysis. Northern transfer experiments showed that the LEU3 gene produces mRNA approximately 2.9 kilonucleotides in length. The leu3 marker was mapped to chromosome XII by the spo11 method. Linkage to ura4 by about 44 centiMorgans places leu3 on the right arm of this chromosome.

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REARRANGEMENT OF THE GENETIC MAP OF CHROMOSOME VII OF SACCHAROMYCES CEREVISIAE

Genetics ◽

10.1093/genetics/109.4.661 ◽

1985 ◽

Vol 109 (4) ◽

pp. 661-664

Author(s):

John L Celenza ◽

Marian Carlson

Keyword(s):

Saccharomyces Cerevisiae ◽

Genetic Distance ◽

Genetic Map ◽

Distal Segment ◽

Tetrad Analysis ◽

Distal Marker ◽

The Right

ABSTRACT The genetic map of the right arm of chromosome VII of Saccharomyces cerevisiae includes markers on a distal segment for which meiotic linkage to the centromere-proximal marker cly8 has not previously been demonstrated. According to the currently accepted map, SUF4 is the most distal marker on the right arm. We have shown by tetrad analysis that SUF4 is linked to cly8 and ade6. The genetic distance between SUF4 and cly8 is 29 cM. These data indicate that the genetic map of the right arm of chromosome VII should be revised by inverting the orientation of the distal segment so that SUF4 is located near cly8, and SUC1 and MAL1 are the most distal markers. With this revision, all of the polymeric fermentation markers that have been mapped are located at the ends of chromosomes.

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Meiotic Crossing Over Between Nonhomologous Chromosomes Affects Chromosome Segregation in Yeast

Genetics ◽

10.1093/genetics/146.1.69 ◽

1997 ◽

Vol 146 (1) ◽

pp. 69-78

Author(s):

Sue Jinks-Robertson ◽

Shariq Sayeed ◽

Tamara Murphy

Keyword(s):

Saccharomyces Cerevisiae ◽

Gene Conversion ◽

Chromosome Segregation ◽

Meiotic Recombination ◽

Southern Blot Analysis ◽

Meiotic Chromosome ◽

Crossing Over ◽

Yeast Saccharomyces Cerevisiae ◽

Reciprocal Translocations ◽

Ectopic Recombination

Meiotic recombination between artificial repeats positioned on nonhomologous chromosomes occurs efficiently in the yeast Saccharomyces cerevisiae. Both gene conversion and crossover eventS have been observed, with crossovers yielding reciprocal translocations. In the current study, 5.5-kb ura3 repeats positioned on chromosomes V and XV were used to examine the effect of ectopic recombination on meiotic chromosome segregation. Ura+ random spores were selected and gene conversion vs. crossover events were distinguished by Southern blot analysis. Approximately 15% of the crossover events between chromosomes V and XV were associated with missegregation of one of these chromosomes. The missegregation was manifest as hyperploid spores containing either both translocations plus a normal chromosome, or both normal chromosomes plus one of the translocations. In those cases where it could be analyzed, missegregation occurred at the first meiotic division. These data are discussed in terms of a model in which ectopic crossovers compete efficiently with normal allelic crossovers in directing meiotic chromosome segregation.

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Mechanisms of gene conversion in Saccharomyces cerevisiae.

Genetics ◽

10.1093/genetics/124.1.7 ◽

1990 ◽

Vol 124 (1) ◽

pp. 7-25

Author(s):

H Roman ◽

M M Ruzinski

Keyword(s):

Saccharomyces Cerevisiae ◽

Stationary Phase ◽

Gene Conversion ◽

Close Contact ◽

Light Dose ◽

The Other ◽

Crossing Over ◽

X Rays ◽

Primary Event ◽

Mitotic Cells

Abstract In red-white sectored colonies of Saccharomyces cerevisiae, derived from mitotic cells grown to stationary phase and irradiated with a light dose of x-rays, all of the segregational products of gene conversion and crossing over can be ascertained. Approximately 80% of convertants are induced in G1, the remaining 20% in G2. Crossing over, in the amount of 20%, is found among G1 convertants but most of the crossovers are delayed until G2. About 20% of all sectored colonies had more than one genotype in one or the other sector, thus confirming the hypothesis that conversion also occurs in G2. The principal primary event in G2 conversion is a single DNA heteroduplex. It is suggested that the close contact that this implies carries over to G2 when crossing over and a second round of conversion occurs.

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Interaction between mismatch repair and genetic recombination in Saccharomyces cerevisiae.

Genetics ◽

10.1093/genetics/137.1.19 ◽

1994 ◽

Vol 137 (1) ◽

pp. 19-39 ◽

Cited By ~ 14

Author(s):

E Alani ◽

R A Reenan ◽

R D Kolodner

Keyword(s):

Saccharomyces Cerevisiae ◽

Gene Conversion ◽

Mismatch Repair ◽

Base Pair ◽

Genetic Recombination ◽

Sequence Similarity ◽

Amino Acid Sequence Similarity ◽

Wild Type ◽

Yeast Saccharomyces Cerevisiae ◽

Heteroduplex Dna

Abstract The yeast Saccharomyces cerevisiae encodes a set of genes that show strong amino acid sequence similarity to MutS and MutL, proteins required for mismatch repair in Escherichia coli. We examined the role of MSH2 and PMS1, yeast homologs of mutS and mutL, respectively, in the repair of base pair mismatches formed during meiotic recombination. By using specifically marked HIS4 and ARG4 alleles, we showed that msh2 mutants displayed a severe defect in the repair of all base pair mismatches as well as 1-, 2- and 4-bp insertion/deletion mispairs. The msh2 and pms1 phenotypes were indistinguishable, suggesting that the wild-type gene products act in the same repair pathway. A comparison of gene conversion events in wild-type and msh2 mutants indicated that mismatch repair plays an important role in genetic recombination. (1) Tetrad analysis at five different loci revealed that, in msh2 mutants, the majority of aberrant segregants displayed a sectored phenotype, consistent with a failure to repair mismatches created during heteroduplex formation. In wild type, base pair mismatches were almost exclusively repaired toward conversion rather than restoration. (2) In msh2 strains 10-19% of the aberrant tetrads were Ab4:4. (3) Polarity gradients at HIS4 and ARG4 were nearly abolished in msh2 mutants. The frequency of gene conversion at the 3' end of these genes was increased and was nearly the frequency observed at the 5' end. (4) Co-conversion studies were consistent with mismatch repair acting to regulate heteroduplex DNA tract length. We favor a model proposing that recombination events occur through the formation and resolution of heteroduplex intermediates and that mismatch repair proteins specifically interact with recombination enzymes to regulate the length of symmetric heteroduplex DNA.

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Spontaneous mitotic recombination in yeast: the hyper-recombinational rem1 mutations are alleles of the RAD3 gene.

Genetics ◽

10.1093/genetics/119.2.289 ◽

1988 ◽

Vol 119 (2) ◽

pp. 289-301

Author(s):

B A Montelone ◽

M F Hoekstra ◽

R E Malone

Keyword(s):

Saccharomyces Cerevisiae ◽

Gene Conversion ◽

Excision Repair ◽

Mitotic Recombination ◽

Crossing Over ◽

Wild Type ◽

Recombination Repair ◽

Mitotic Gene Conversion ◽

Type Gene ◽

Repair Genes

Abstract The RAD3 gene of Saccharomyces cerevisiae is required for UV excision-repair and is essential for cell viability. We have identified the rem1 mutations (enhanced spontaneous mitotic recombination and mutation) of Saccharomyces cerevisiae as alleles of RAD3 by genetic mapping, complementation with the cloned wild-type gene, and DNA hybridization. The high levels of spontaneous mitotic gene conversion, crossing over, and mutation conferred upon cells by the rem1 mutations are distinct from the effects of all other alleles of RAD3. We present preliminary data on the localization of the rem1 mutations within the RAD3 gene. The interaction of the rem1 mutant alleles with a number of radiation-sensitive mutations is also different than the interactions reported for previously described (UV-sensitive) alleles of RAD3. Double mutants of rem1 and a defect in the recombination-repair pathway are inviable, while double mutants containing UV-sensitive alleles of RAD3 are viable. The data presented here demonstrate that: (1) rem1 strains containing additional mutations in other excision-repair genes do not exhibit elevated gene conversion; (2) triple mutants containing rem1 and mutations in both excision-repair and recombination-repair are viable; (3) such triple mutants containing rad52 have reduced levels of gene conversion but wild-type frequencies of crossing over. We have interpreted these observations in a model to explain the effects of rem1. Consistent with the predictions of the model, we find that the size of DNA from rem1 strains, as measured by neutral sucrose gradients, is smaller than wild type.

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Mitotic and meiotic gene conversion of Ty elements and other insertions in Saccharomyces cerevisiae.

Genetics ◽

10.1093/genetics/122.4.759 ◽

1989 ◽

Vol 122 (4) ◽

pp. 759-772

Author(s):

A Vincent ◽

T D Petes

Keyword(s):

Saccharomyces Cerevisiae ◽

Gene Conversion ◽

Point Mutations ◽

Crossing Over ◽

Yeast Saccharomyces Cerevisiae ◽

Ty Elements ◽

Meiotic Gene ◽

Mitotic Gene Conversion

Abstract We examined meiotic and mitotic gene conversion events involved in deletion of Ty elements and other insertions from the genome of the yeast Saccharomyces cerevisiae. We found that Ty elements and one other insertion were deleted by mitotic gene conversion less frequently than point mutations at the same loci. One non-Ty insertion similar in size to Ty, however, did not show this bias. Mitotic conversion events deleting insertions were more frequently associated with crossing over than those deleting point mutations. In meiosis, conversion events duplicating the element were more common than those that deleted the element for one of the loci (HIS4) examined.

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