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Topoisomerase I plays a critical role in suppressing genome instability at a highly transcribed G-quadruplex-forming sequence.

Yadav P, Harcy V, Argueso JL, Dominska M, Jinks-Robertson S, Kim N - PLoS Genet. (2014)

Bottom Line: Transcription confers a critical strand bias since genome rearrangements at the G4-forming Sμ are elevated only when the guanine-runs are located on the non-transcribed strand.The direction of replication and transcription, when in a head-on orientation, further contribute to the elevated genome instability at a potential G4 DNA-forming sequence.The implications of our identification of Top1 as a critical factor in suppression of instability associated with potential G4 DNA-forming sequences are discussed.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Molecular Genetics, University of Texas Health Science Center at Houston, Houston, Texas, United States of America.

ABSTRACT
G-quadruplex or G4 DNA is a non-B secondary DNA structure that comprises a stacked array of guanine-quartets. Cellular processes such as transcription and replication can be hindered by unresolved DNA secondary structures potentially endangering genome maintenance. As G4-forming sequences are highly frequent throughout eukaryotic genomes, it is important to define what factors contribute to a G4 motif becoming a hotspot of genome instability. Using a genetic assay in Saccharomyces cerevisiae, we previously demonstrated that a potential G4-forming sequence derived from a guanine-run containing immunoglobulin switch Mu (Sμ) region becomes highly unstable when actively transcribed. Here we describe assays designed to survey spontaneous genome rearrangements initiated at the Sμ sequence in the context of large genomic areas. We demonstrate that, in the absence of Top1, a G4 DNA-forming sequence becomes a strong hotspot of gross chromosomal rearrangements and loss of heterozygosity associated with mitotic recombination within the ∼ 20 kb or ∼ 100 kb regions of yeast chromosome V or III, respectively. Transcription confers a critical strand bias since genome rearrangements at the G4-forming Sμ are elevated only when the guanine-runs are located on the non-transcribed strand. The direction of replication and transcription, when in a head-on orientation, further contribute to the elevated genome instability at a potential G4 DNA-forming sequence. The implications of our identification of Top1 as a critical factor in suppression of instability associated with potential G4 DNA-forming sequences are discussed.

No MeSH data available.


Related in: MedlinePlus

The rates of loss of heterozygosity on CHR3 in wild-type backgrounds.A. The total rates of LOH events (5-FOAR). LYS2 or GTOP indicates pTET-LYS2 or pTET-lys2-GTOP allele integrated in each of the strains analyzed. The 95% confidence intervals are indicated by the error bars. B. The rates of LOH initiating at pTET-LYS2 or pTET-lys2-GTOP cassette. LOH events are defined as initiating at pTET-LYS2 or pTET-lys2-GTOP cassette when URA3 and G418R are lost, only YJM789 sequence is present at StyI, BsrBI, and SpeI SNP sites and both YJM789 and YPH45 sequence is present at NarI and HindIII SNP sites (See Table S2, Table 2 and Table 3). There was no statistically significant difference between all rates.
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pgen-1004839-g004: The rates of loss of heterozygosity on CHR3 in wild-type backgrounds.A. The total rates of LOH events (5-FOAR). LYS2 or GTOP indicates pTET-LYS2 or pTET-lys2-GTOP allele integrated in each of the strains analyzed. The 95% confidence intervals are indicated by the error bars. B. The rates of LOH initiating at pTET-LYS2 or pTET-lys2-GTOP cassette. LOH events are defined as initiating at pTET-LYS2 or pTET-lys2-GTOP cassette when URA3 and G418R are lost, only YJM789 sequence is present at StyI, BsrBI, and SpeI SNP sites and both YJM789 and YPH45 sequence is present at NarI and HindIII SNP sites (See Table S2, Table 2 and Table 3). There was no statistically significant difference between all rates.

Mentions: To determine whether highly transcribed G4 motifs elevate LOH on CHR3, we measured the rate of 5-FOAR in WT strains containing pTET-LYS2 (no Sμ sequence) or pTET-lys2-GTOP cassette. In either the SAME or OPPO orientations, the overall rate of LOH (5-FOAR) associated with pTET-LYS2 or pTET-lys2-GTOP was not significantly different (Fig. 4A). Using the RFLP-SNP assay, we identified 3/46 or 9/46 LOH events initiated at the highly transcribed pTET-LYS2 cassette when in the SAME or the OPPO orientation, respectively (Table 3). These proportions were not statistically different from those for LOH initiating at pTET-lys2-GTOP in the SAME or OPPO orientation (by Fisher's exact test; P = 0.31 and 0.39, respectively).


Topoisomerase I plays a critical role in suppressing genome instability at a highly transcribed G-quadruplex-forming sequence.

Yadav P, Harcy V, Argueso JL, Dominska M, Jinks-Robertson S, Kim N - PLoS Genet. (2014)

The rates of loss of heterozygosity on CHR3 in wild-type backgrounds.A. The total rates of LOH events (5-FOAR). LYS2 or GTOP indicates pTET-LYS2 or pTET-lys2-GTOP allele integrated in each of the strains analyzed. The 95% confidence intervals are indicated by the error bars. B. The rates of LOH initiating at pTET-LYS2 or pTET-lys2-GTOP cassette. LOH events are defined as initiating at pTET-LYS2 or pTET-lys2-GTOP cassette when URA3 and G418R are lost, only YJM789 sequence is present at StyI, BsrBI, and SpeI SNP sites and both YJM789 and YPH45 sequence is present at NarI and HindIII SNP sites (See Table S2, Table 2 and Table 3). There was no statistically significant difference between all rates.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4256205&req=5

pgen-1004839-g004: The rates of loss of heterozygosity on CHR3 in wild-type backgrounds.A. The total rates of LOH events (5-FOAR). LYS2 or GTOP indicates pTET-LYS2 or pTET-lys2-GTOP allele integrated in each of the strains analyzed. The 95% confidence intervals are indicated by the error bars. B. The rates of LOH initiating at pTET-LYS2 or pTET-lys2-GTOP cassette. LOH events are defined as initiating at pTET-LYS2 or pTET-lys2-GTOP cassette when URA3 and G418R are lost, only YJM789 sequence is present at StyI, BsrBI, and SpeI SNP sites and both YJM789 and YPH45 sequence is present at NarI and HindIII SNP sites (See Table S2, Table 2 and Table 3). There was no statistically significant difference between all rates.
Mentions: To determine whether highly transcribed G4 motifs elevate LOH on CHR3, we measured the rate of 5-FOAR in WT strains containing pTET-LYS2 (no Sμ sequence) or pTET-lys2-GTOP cassette. In either the SAME or OPPO orientations, the overall rate of LOH (5-FOAR) associated with pTET-LYS2 or pTET-lys2-GTOP was not significantly different (Fig. 4A). Using the RFLP-SNP assay, we identified 3/46 or 9/46 LOH events initiated at the highly transcribed pTET-LYS2 cassette when in the SAME or the OPPO orientation, respectively (Table 3). These proportions were not statistically different from those for LOH initiating at pTET-lys2-GTOP in the SAME or OPPO orientation (by Fisher's exact test; P = 0.31 and 0.39, respectively).

Bottom Line: Transcription confers a critical strand bias since genome rearrangements at the G4-forming Sμ are elevated only when the guanine-runs are located on the non-transcribed strand.The direction of replication and transcription, when in a head-on orientation, further contribute to the elevated genome instability at a potential G4 DNA-forming sequence.The implications of our identification of Top1 as a critical factor in suppression of instability associated with potential G4 DNA-forming sequences are discussed.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Molecular Genetics, University of Texas Health Science Center at Houston, Houston, Texas, United States of America.

ABSTRACT
G-quadruplex or G4 DNA is a non-B secondary DNA structure that comprises a stacked array of guanine-quartets. Cellular processes such as transcription and replication can be hindered by unresolved DNA secondary structures potentially endangering genome maintenance. As G4-forming sequences are highly frequent throughout eukaryotic genomes, it is important to define what factors contribute to a G4 motif becoming a hotspot of genome instability. Using a genetic assay in Saccharomyces cerevisiae, we previously demonstrated that a potential G4-forming sequence derived from a guanine-run containing immunoglobulin switch Mu (Sμ) region becomes highly unstable when actively transcribed. Here we describe assays designed to survey spontaneous genome rearrangements initiated at the Sμ sequence in the context of large genomic areas. We demonstrate that, in the absence of Top1, a G4 DNA-forming sequence becomes a strong hotspot of gross chromosomal rearrangements and loss of heterozygosity associated with mitotic recombination within the ∼ 20 kb or ∼ 100 kb regions of yeast chromosome V or III, respectively. Transcription confers a critical strand bias since genome rearrangements at the G4-forming Sμ are elevated only when the guanine-runs are located on the non-transcribed strand. The direction of replication and transcription, when in a head-on orientation, further contribute to the elevated genome instability at a potential G4 DNA-forming sequence. The implications of our identification of Top1 as a critical factor in suppression of instability associated with potential G4 DNA-forming sequences are discussed.

No MeSH data available.


Related in: MedlinePlus