Genomic instability induced by furfural-treated in S. cerevisiae [sectored colonies]
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ABSTRACT:
ORGANISM(S): Saccharomyces cerevisiae
PROVIDER: GSE104880 | GEO | 2017/10/13
SECONDARY ACCESSION(S): PRJNA414089
REPOSITORIES: GEO
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Project description:Genomic instability is a common feature found in cancer cells. Accordingly, many tumor suppressor genes identified in familiar cancer syndromes are involved in the maintenance of the stability of the genome during every cell division, and are commonly referred to as caretakers. Inactivating mutations and epigenetic silencing of caretakers are thought to be the most important mechanism that explains cancer-related genome instability. However, little is known of whether transient inactivation of caretaker proteins could trigger genome instability and, if so, what types of instability would occur. In this work, we show that a brief and reversible inactivation, during just one cell cycle, of the key phosphatase Cdc14 in the model organism Saccharomyces cerevisiae is enough to result in diploid cells with multiple gross chromosomal rearrangements and changes in ploidy. Interestingly, we observed that such transient inactivation yields a characteristic fingerprint whereby trisomies are often found in small-sized chromosomes and gross chromosome rearrangements, often associated with concomitant loss of heterozygosity (LOH), are mainly detected on the rDNA-bearing chromosome XII. Taking into account the key role of Cdc14 in preventing anaphase bridges, resetting replication origins and controlling spindle dynamics in a well-defined window within anaphase, we speculate that its transient inactivation causes cells to go through a single mitotic catastrophe with irreversible consequences for the genome stability of the progeny. The details of these experiments are in press in Genetics (Oliver Quevedo, Cristina Ramos-Pérez, Thomas D. Petes, and Félix Machín) but will be described briefly below. The diploid S. cerevisiae strain FM1468 is homozygous for a temperature-sensitive allele of CDC14 which encodes a phosphatase that is important in regulating the cell cycle. The strain was constructed by mating cdc14-1 derivatives of two sequence-diverged haploids, one isogenic with W303-1A (JSC12) and one isogenic with YJM789. The resulting diploid is heterozygous for about 55,000 SNPs (St. Charles, J. and Petes, T. (2013) PLoS Genetics 9: e1003434; PMID 23593029). In addition, the strain is homozygous for the ade2-1 ochre mutation and heterozygous for an insertion of SUP4-o, a gene encoded a tRNA ochre suppressor. The SUP4-o gene is located near the right telomere of chromosome IV and is inserted on the YJM789-derived homolog. The FM1468 strain forms pink colonies because a single copy of SUP4-o partially suppresses the red colony phenotype associated with the ade2 mutation. In the wild-type genetic background, a mitotic crossover between SUP4-o and the centromere of IV produces a red/white sectored colony with the red sector containing no copies of SUP4-o and the white sector containing two copies of SUP4-o (St. Charles and Petes, 2013). To map the position of the crossover and to examine recombination events resulting in loss of heterozygosity (LOH) throughout the genome, we used SNP-specific microarrays (St. Charles et al., 2012; Genetics 190: 1267-1284; PMID 22267500). These arrays contained about 50,000 oligonucleotides, allowing us to monitor LOH at about 13,000 positions in the genome. Each of these single-nucleotide polymorphism (SNP) was represented by four 25-base nucleotides, two for the Watson and Crick strands of the YJM789-derived SNP and two for the Watson and Crick strands of the W303-1A-derived SNP. The sequences and locations of each of these oligonucleotides has been published (St. Charles et al., 2012). As described below, by measuring the level of hybridization to these oligonucleotides, we identified the location of LOH events throughout the genome in FM1468. It should also be noted that the microarrays detect changes in chromosome number in addition to detecting LOH events. A total of 13 sectored colonies were examined. Each colony was given a number and letter indicate the sectors. For example, 1A and 1B are different sectors of one FM1468 colony. The key to the color of these colonies is: 1A (white), 1B (pink); 2A (pink), 2B (red); 3A (white), 3B (pink); 4A (white), 4B (pink); 5A (white), 5B (red); 6A (white), 6B (pink); 7A (white), 7B (pink); 8A (pink), 8B (red); 9A (white), 9B (pink); 10A (white), 10B (pink), 10C (red); 11A (white), 11B (pink), 11C (red); 12A (white), 12B (red); 13A (white), 13B (red). Note that colonies 10 and 11 were tri-colored colonies, and microarrays were done on all three sectors. In summary, microarry analysis was done on 28 sectors derived from 13 colonies.
2015-05-05 | E-GEOD-68530 | biostudies-arrayexpress
Project description:Two types of RNA:DNA associations can lead to genome instability: the formation of R-loops during transcription and the incorporation of ribonucleotide monophosphates (rNMPs) into DNA during replication. Both ribonuclease (RNase) H1 and RNase H2 degrade the RNA component of R-loops, whereas only RNase H2 can remove one or a few rNMPs from DNA. We performed high-resolution mapping of mitotic recombination events throughout the yeast genome in diploid strains of Saccharomyces cerevisiae lacking RNase H1 (rnh1Δ), RNase H2 (rnh201Δ), or both RNase H1 and RNase H2 (rnh1Δ rnh201Δ). We found little effect on recombination in the rnh1Δ strain, but elevated recombination in both the rnh201Δ and the double-mutant strains; levels of recombination in the double mutant were about 50% higher than in the rnh201 single-mutant strain. An rnh201Δ mutant that additionally contained a mutation that reduces rNMP incorporation by DNA polymerase ε (pol2-M644L) had a level of instability similar to that observed in the presence of wild-type Polε. This result suggests that the elevated recombination observed in the absence of only RNase H2 is primarily a consequence of R loops rather than misincorporated rNMPs. The details of these experiments are in press in Genetics This diploid S.cerevisiae strain used here is a hybrid between W303-1A and YJM789. The backgrounds are diverged by about 50,000 polymorphisms that are surveyed by custom CGH microarrays containing SNP-specific probes (St. Charles et al., 2012; Genetics 190: 1267-1284; PMID 22267500). The whole genome array surveyed loss of heterozygosity (LOH) events throughout the genome of in wild type, pol2-M644L, rnh1Δ, rnh201Δ, rnh201Δ pol2-M644L, and rnh1Δ rnh201Δ mutant backgrounds in 10, 12, 10, 19, 17, and 22 samples, respectively. The names of these strains are KO_198,KO_234, KO_73, KO_75, KO_244, and KO_5, respectively. Another assay was performed mapping reciprocal crossover events on the right arm of chromosome IV in the rnh201Δ (strains KO_135 and KO188) and the rnh1Δrnh201Δ (strain KO_132) mutant backgrounds. A reporter system with a wild type ADE2 on the end of chromosome IV allowed us to detect crossovers by seeing a red-white sectored colony. Each reciprocal event was mapped by doing a microarray specific for the right arm of chromosome IV (St. Charles and Petes, 2013; PLoS Genet. 1267-1284 PMCID PMC3616911) on the red and white sides of sectors. 21 sectors were analyzed for the rnh201Δ mutant while 14 were analyzed for the rnh1Δ rnh201Δ mutant. We mapped crossovers on the right arm of chromosome IV in the rnh201Δ mutant in four different diploid isolates: KO135_5 (1R/1W, 2R/2W, 4R/4W, 5R/5W, 12R/12W, and 13R/13W), KO_135_6 (1R/1W, 4R/4W, 5R/5W, 7R/7W, 9R/9W, 11R/11W, 13R/13W, and 17R/17W), KO 188_1 (1R/1W, and 4R/4W), and KO188_2 (6R/6W, 7R/7W, 11R/11W, and 12R/12W). We mapped crossovers on the right arm of chromosome IV in the rnh1Δ rnh201Δ mutant in two different diploid isolates: KO_132_31 (2R/2W, 6R/6W, 8R/8W, 15R/15W, 17R/17W, 21R/21W, 22R/22W, and 24R/24W) and KO_132_29 (3R/3W, 7R/7W, 18R/18W, 24R/24W, and 34R/34W).
2015-09-23 | E-GEOD-73334 | biostudies-arrayexpress
Project description:We have replaced the right arm of chromosome IX in Saccharomyces cerevisiae with a synthetic version to generate synIXR haploids. The synthetic chromosome features multiple sequeunce modifications. We analyzed gene expression by microarray analysis in three synIXR haploids (1D, 6B, and 22D) to detect any changes in synIXR transcripts or global compensatory changes.
2011-09-14 | GSE31326 | GEO
Project description:We have replaced the right arm of chromosome IX in Saccharomyces cerevisiae with a synthetic version to generate synIXR haploids. The synthetic chromosome features multiple sequeunce modifications. We analyzed gene expression by microarray analysis in three synIXR haploids (1D, 6B, and 22D) to detect any changes in synIXR transcripts or global compensatory changes. Biological triplicates of three synIXR haploids (1D, 6B, and 22D) and corresponding controls (either BY4741 or BY4742) were grown to mid-log phase and collected for mRNA isolation.
2011-09-13 | E-GEOD-31326 | biostudies-arrayexpress
Project description:Gene number of the fission yeast Schizosaccharomyces pombe is the lowest in free-living eukaryotes. We further reduced the gene number to examine a minimal gene set required for growth. The genome-reduced strain, named ASP3894, has four chromosomal deletion regions: 168.4 kb deletion in the left arm of chromosome 1 (ALT), 155.4 kb deletion in the right arm of chromosome 1 (ART), 211.7 kb deletion in the left arm of chromosome 2 (BLT) and 121.6 kb deletion in the right arm of chromosome 2 (BRT). As a result of the 657.3 kb genome reduction, 224 genes of approximately 5100 total genes were reduced. ASP3894 transcriptional profiling was compared with a parent strain, ASP3880, that is a non-genome-reduced strain to examine the influence of genome reduction.
2013-05-01 | GSE38620 | GEO
Project description:Saccharomyces cerevisiae synthetic chromosome IV
| PRJNA851090 | ENA
Project description:Saccharomyces cerevisiae switches its mating type (MATa or MATalpha) through gene conversion using one of two possible donors, HMLalpha or HMRa, both of which are maintained by the SIR silencing complex as heterochromatic cassettes on opposing ends of chromosome III. In MATa cells, HMLalpha on the left arm is preferentially chosen as the donor through a mechanism requiring a cis-acting sequence called the recombination enhancer (RE). The left half of the RE is required for this donor preference activity, whereas the right half has been implicated in regulating chromosome III structure. In this study we have identified a MATa-specific Sir2 and condensin binding site within the right half of the RE that maintains chromosome III in a switching-competent conformation by preventing MATa from interacting with the default HMRa donor on the right arm. Within the RE, Sir2 strongly represses transcription of a small MATa-specific gene of unknown function (RDT1). Upon expression of HO endonuclease to induce the switching process, Sir2 redistributes from the RE to the double-stranded DNA break site at MATa, thus derepressing RDT1 transcription to coincide with the timing of mating-type switching. Condensin is also displaced from the RE in response to the HO-induced break, likely contributing to the transient change in chromosome III conformation required for effective switching without disruption of HML and HMR heterochromatin.
2019-05-23 | GSE92715 | GEO
Project description:Saccharomyces cerevisiae switches its mating type (MATa or MATalpha) through gene conversion using one of two possible donors, HMLalpha or HMRa, both of which are maintained by the SIR silencing complex as heterochromatic cassettes on opposing ends of chromosome III. In MATa cells, HMLalpha on the left arm is preferentially chosen as the donor through a mechanism requiring a cis-acting sequence called the recombination enhancer (RE). The left half of the RE is required for this donor preference activity, whereas the right half has been implicated in regulating chromosome III structure. In this study we have identified a MATa-specific Sir2 and condensin binding site within the right half of the RE that maintains chromosome III in a switching-competent conformation by preventing MATa from interacting with the default HMRa donor on the right arm. Within the RE, Sir2 strongly represses transcription of a small MATa-specific gene of unknown function (RDT1). Upon expression of HO endonuclease to induce the switching process, Sir2 redistributes from the RE to the double-stranded DNA break site at MATa, thus derepressing RDT1 transcription to coincide with the timing of mating-type switching. Condensin is also displaced from the RE in response to the HO-induced break, likely contributing to the transient change in chromosome III conformation required for effective switching without disruption of HML and HMR heterochromatin.
2019-05-23 | GSE92714 | GEO
Project description:Gene number of the fission yeast Schizosaccharomyces pombe is the lowest in free-living eukaryotes. We further reduced the gene number to examine a minimal gene set required for growth. The genome-reduced strain, named ASP3894, has four chromosomal deletion regions: 168.4 kb deletion in the left arm of chromosome 1 (ALT), 155.4 kb deletion in the right arm of chromosome 1 (ART), 211.7 kb deletion in the left arm of chromosome 2 (BLT) and 121.6 kb deletion in the right arm of chromosome 2 (BRT). As a result of the 657.3 kb genome reduction, 224 genes of approximately 5100 total genes were reduced. ASP3894 transcriptional profiling was compared with a parent strain, ASP3880, that is a non-genome-reduced strain to examine the influence of genome reduction. Three independent experiments. 3 control ASP3880 samples and 3 genome-reduced strain ASP3894 samples were independently grown in EMM medium and harvested at the logarithmic phase.
2013-05-01 | E-GEOD-38620 | biostudies-arrayexpress
Project description:Chromosome arm length, and a species-specific determinant, define chromosome arm width [S. cerevisiae]
| PRJNA623872 | ENA