Mitotic chromosome loss induced by methyl benzimidazole-2-yl-carbamate as a rapid mapping method in Saccharomyces cerevisiae

1982 ◽  
Vol 2 (9) ◽  
pp. 1080-1087
Author(s):  
J S Wood
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Chromosome ◽  
Chromosomal Location ◽  
Mapping Method ◽  
Chromosome Loss ◽  
Diploid Strain ◽  
Simple Method ◽  
Rapid Mapping

Mitotic chromosome loss induced by methyl benzimidazole-2-yl-carbamate has been utilized as a rapid and simple method for assigning genes to individual chromosomes in Saccharomyces cerevisiae. This technique relied on the segregation of heterozygous markers in a diploid strain after methyl benzimidazole-2-yl-carbamate treatment due to loss of whole chromosomes. Correlations between the expression of an unmapped gene and that of a previously mapped recessive marker indicated chromosomal linkage. Depending on whether the unmapped gene and the marker were located in coupling or in repulsion, either positive or negative correlations were seen. The chromosomal location of several previously mapped genes were confirmed as a test of the method, and one previously unmapped gene, nib1, was mapped.

scholarly journals Mitotic chromosome loss induced by methyl benzimidazole-2-yl-carbamate as a rapid mapping method in Saccharomyces cerevisiae.

1982 ◽  
Vol 2 (9) ◽  
pp. 1080-1087 ◽  
Author(s):  
J S Wood
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Chromosome ◽  
Chromosomal Location ◽  
Mapping Method ◽  
Chromosome Loss ◽  
Diploid Strain ◽  
Simple Method ◽  
Rapid Mapping

Mitotic chromosome loss induced by methyl benzimidazole-2-yl-carbamate has been utilized as a rapid and simple method for assigning genes to individual chromosomes in Saccharomyces cerevisiae. This technique relied on the segregation of heterozygous markers in a diploid strain after methyl benzimidazole-2-yl-carbamate treatment due to loss of whole chromosomes. Correlations between the expression of an unmapped gene and that of a previously mapped recessive marker indicated chromosomal linkage. Depending on whether the unmapped gene and the marker were located in coupling or in repulsion, either positive or negative correlations were seen. The chromosomal location of several previously mapped genes were confirmed as a test of the method, and one previously unmapped gene, nib1, was mapped.

Detection of induced mitotic chromosome loss in Saccharomyces cerevisiae — an interlaboratory study

1989 ◽  
Vol 224 (1) ◽  
pp. 31-78 ◽  
Author(s):  
S.G. Whittaker ◽  
F.K. Zimmermann ◽  
B. Dicus ◽  
W.W. Piegorsch ◽  
S. Fogel ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Chromosome ◽  
Interlaboratory Study ◽  
Chromosome Loss

Mutational analysis of centromere DNA from chromosome VI of Saccharomyces cerevisiae

1988 ◽  
Vol 8 (6) ◽  
pp. 2523-2535
Author(s):  
J H Hegemann ◽  
J H Shero ◽  
G Cottarel ◽  
P Philippsen ◽  
P Hieter
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Chromosome ◽  
Mutational Analysis ◽  
Point Mutations ◽  
Vector System ◽  
Chromosome Loss ◽  
Wild Type ◽  
Sequence Elements ◽  
Chromosome Fragments

Saccharomyces cerevisiae centromeres have a characteristic 120-base-pair region consisting of three distinct centromere DNA sequence elements (CDEI, CDEII, and CDEIII). We have generated a series of 26 CEN mutations in vitro (including 22 point mutations, 3 insertions, and 1 deletion) and tested their effects on mitotic chromosome segregation by using a new vector system. The yeast transformation vector pYCF5 was constructed to introduce wild-type and mutant CEN DNAs onto large, linear chromosome fragments which are mitotically stable and nonessential. Six point mutations in CDEI show increased rates of chromosome loss events per cell division of 2- to 10-fold. Twenty mutations in CDEIII exhibit chromosome loss rates that vary from wild type (10(-4)) to nonfunctional (greater than 10(-1)). These results directly identify nucleotides within CDEI and CDEIII that are required for the specification of a functional centromere and show that the degree of conservation of an individual base does not necessarily reflect its importance in mitotic CEN function.

scholarly journals Isolation of the gene encoding the Saccharomyces cerevisiae centromere-binding protein CP1.

1990 ◽  
Vol 10 (6) ◽  
pp. 2458-2467 ◽  
Author(s):  
R E Baker ◽  
D C Masison
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Disruption ◽  
Mitotic Chromosome ◽  
Binding Protein ◽  
Chromosome Loss ◽  
Chromosome X ◽  
Gene Encoding ◽  
Yeast Saccharomyces Cerevisiae ◽  
Unexpected Consequence ◽  
Cell Doubling Time

CP1 is a sequence-specific DNA-binding protein of the yeast Saccharomyces cerevisiae which recognizes the highly conserved DNA element I (CDEI) of yeast centromeres. We cloned and sequenced the gene encoding CP1. The gene codes for a protein of molecular weight 39,400. When expressed in Escherichia coli, the CP1 gene directed the synthesis of a CDEI-binding protein having the same gel mobility as purified yeast CP1. We have given the CP1 gene the genetic designation CEP1 (centromere protein 1). CEP1 was mapped and found to reside on chromosome X, 2.0 centimorgans from SUP4. Strains were constructed in which most of CEP1 was deleted. Such strains lacked detectable CP1 activity and were viable; however, CEP1 gene disruption resulted in a 35% increase in cell doubling time and a ninefold increase in the rate of mitotic chromosome loss. An unexpected consequence of CP1 gene disruption was methionine auxotrophy genetically linked to cep1. This result and the recent finding that CDEI sites in the MET25 promoter are required to activate transcription (D. Thomas, H. Cherest, and Y. Surdin-Kerjan, J. Mol. Biol. 9:3292-3298, 1989) suggest that CP1 is both a kinetochore protein and a transcription factor.

scholarly journals MITOTIC CHROMOSOME LOSS IN A DISOMIC HAPLOID OF SACCHAROMYCES CEREVISIAE

Genetics ◽  
1975 ◽  
Vol 79 (3) ◽  
pp. 383-396
Author(s):  
D A Campbell ◽  
S Fogel ◽  
K Lusnak
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mating Type ◽  
Mitotic Chromosome ◽  
Marker Genes ◽  
Chromosome Loss ◽  
Type Locus ◽  
Group Iii ◽  
Mating Type Locus ◽  
Independent Events ◽  
Spontaneous Chromosome

ABSTRACT Experiments designed to characterize the incidence of mitotic chromosome loss in a yeast disomic haploid were performed. The selective methods employed utilize the non-mating property of strains disomic for linkage group III and heterozygous at the mating type locus. The principal findings are: (1) The frequency of spontaneous chromosome loss in the disome is of the order 10-4 per cell; this value approximates the frequency in the same population of spontaneous mitotic exchange resulting in homozygosity at the mating type locus. (2) The recovered diploids are pure clones, and thus represent unique events in the disomic haploid. (3) Of the euploid chromosomes recovered after events leading to chromosome loss, approximately 90% retain the parental marker configuration expected from segregation alone; however, the remainder are recombinant for marker genes, and are the result of mitotic exchanges in the disome, especially in regions near the centromere. The recombinant proportion significantly exceeds that expected if chromosome loss and mitotic exchange in the disome were independent events. The data are consistent with a model proposing mitotic nondisjunction as the event responsible for chromosome loss in the disomic haploid.

scholarly journals Meiosis in Saccharomyces cerevisiae mutants lacking the centromere-binding protein CP1.

Genetics ◽  
1992 ◽  
Vol 131 (1) ◽  
pp. 43-53 ◽  
Author(s):  
D C Masison ◽  
R E Baker
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Chromosome ◽  
Binding Protein ◽  
Pleiotropic Effect ◽  
Chromosome Loss ◽  
Spore Viability ◽  
Chromosome Fragments ◽  
Unpaired Chromosome ◽  
Yeast Meiosis ◽  
Chromosome I

Abstract CP1 (encoded by the CEP1 gene) is a centromere binding protein of Saccharomyces cerevisiae that binds to the conserved DNA element I (CDEI) of yeast centromeres. To investigate the function of CP1 in yeast meiosis, we analyzed the meiotic segregation of CEN plasmids, nonessential chromosome fragments (CFs) and chromosomes in cep1 null mutants. Plasmids and CFs missegregated in 10-20% of meioses with the most frequent type of aberrant event being precocious sister segregation at the first meiotic division; paired and unpaired CFs behaved similarly. An unpaired chromosome I homolog (2N + 1) also missegregated at high frequency in the cep1 mutant (7.6%); however, missegregation of other chromosomes was not detected by tetrad analysis. Spore viability of cep1 tetrads was significantly reduced, and the pattern of spore death was nonrandom. The inviability could not be explained solely by chromosome missegregation and is probably a pleiotropic effect of cep1. Mitotic chromosome loss in cep1 strains was also analyzed. Both simple loss (1:0 segregation) and nondisjunction (2:0 segregation) were increased, but the majority of loss events resulted from nondisjunction. We interpret the results to suggest that CP1 generally promotes chromatid-kinetochore adhesion.

Genetic effects of methyl benzimidazole-2-yl-carbamate on Saccharomyces cerevisiae

1982 ◽  
Vol 2 (9) ◽  
pp. 1064-1079
Author(s):  
J S Wood
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Spindle ◽  
High Frequency ◽  
Mitotic Chromosome ◽  
Genetic Effect ◽  
Genetic Effects ◽  
Chromosome Loss ◽  
Polyploid Cells ◽  
Mitotic Inhibitor ◽  
Chromosome Nondisjunction

The genetic effects of the mitotic inhibitor methyl benzimidazole-2-yl-carbamate (MBC) have been studied in Saccharomyces cerevisiae. MBC had little or no effect on the frequency of mutation. In some experiments MBC caused an increase in the frequency of mitotic recombination; however, this effect was small and not reproducible. The primary genetic effect of MBC was to induce mitotic chromosome loss at a high frequency. Chromosome loss occurred at equal frequencies for all chromosomes tested (13 of 16). Cells which had lost multiple chromosomes were found more frequently than predicted if individual chromosome loss events were independent. The probability of loss for a particular chromosome increased with length of time cells were incubated with MBC. MBC treatment also increased the frequency at which polyploid cells were found. These results suggested that MBC acted to disrupt the structure or function of the mitotic spindle and cause chromosome nondisjunction.

scholarly journals Mutational analysis of centromere DNA from chromosome VI of Saccharomyces cerevisiae.

1988 ◽  
Vol 8 (6) ◽  
pp. 2523-2535 ◽  
Author(s):  
J H Hegemann ◽  
J H Shero ◽  
G Cottarel ◽  
P Philippsen ◽  
P Hieter
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Chromosome ◽  
Mutational Analysis ◽  
Point Mutations ◽  
Vector System ◽  
Chromosome Loss ◽  
Wild Type ◽  
Sequence Elements ◽  
Chromosome Fragments

Saccharomyces cerevisiae centromeres have a characteristic 120-base-pair region consisting of three distinct centromere DNA sequence elements (CDEI, CDEII, and CDEIII). We have generated a series of 26 CEN mutations in vitro (including 22 point mutations, 3 insertions, and 1 deletion) and tested their effects on mitotic chromosome segregation by using a new vector system. The yeast transformation vector pYCF5 was constructed to introduce wild-type and mutant CEN DNAs onto large, linear chromosome fragments which are mitotically stable and nonessential. Six point mutations in CDEI show increased rates of chromosome loss events per cell division of 2- to 10-fold. Twenty mutations in CDEIII exhibit chromosome loss rates that vary from wild type (10(-4)) to nonfunctional (greater than 10(-1)). These results directly identify nucleotides within CDEI and CDEIII that are required for the specification of a functional centromere and show that the degree of conservation of an individual base does not necessarily reflect its importance in mitotic CEN function.

scholarly journals ASSOCIATION OF DISOMIC CHROMOSOME LOSS WITH EMS-INDUCED CONVERSION IN YEAST

Genetics ◽  
1980 ◽  
Vol 96 (3) ◽  
pp. 613-625
Author(s):  
Douglas Campbell
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Causal Relationship ◽  
Mitotic Chromosome ◽  
Experimental Tests ◽  
Chromosome Loss ◽  
Constant Independent ◽  
Yeast Saccharomyces Cerevisiae ◽  
Proposed Model ◽  
Mitotic Gene Conversion ◽  
Chromosome Iii

ABSTRACT Experimental tests with the yeast Saccharomyces cerevisiae of a previously proposed model suggesting a causal relationship between disomic chromosome loss (n + 1 → n) and centromere-adjacent mitotic gene conversion were performed. Disomic haploid cells heteroallelic at two loci on the left arm of chromosome III were exposed to ethyl methanesulfonate (EMS) under nonlethal conditions; EMS-induced prototrophic gene convertants were selected and tested for coincident chromosome loss. The principal results are: (1) The frequency of chromosome loss among EMS-induced gene convertants selected to arise near the centromere is markedly enhanced over basal levels and remains constant, independent of EMS exposure. There is little such enhancement among EMS-induced convertants selected to arise far from the centromere. (2) Chromosome loss is almost completely associated with induced conversion of the centromere-proximal allele at the centromere-adjacent heteroallelic locus. This result is identical to (and confirms) results found previously for spontaneous loss-associated conversion. (3) The conversion polarity at the centromere-adjacent locus among unselected (nonloss-associated) induced or spontaneous mitotic convertants is identical to that among meiotic convertants and markedly favors the contromere-distal allele. These findings are wholly consistent with, and strengthen, the hypothesis that structural involvement of centromeric regions in nearby recombinational events may interfere with proper segregational function and lead to mitotic chromosome loss.

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