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Saccharomyces cerevisiae cells: control vs positive supercoiling accumulation after 0, 30 and 120 min

ABSTRACT: Positive DNA helical stress accumulates in vivo by the unbalanced relaxation of positive and negative DNA supercoils in Δtop1, top2ts, pGPD:TopA yeast cells. The resulting overwinding of DNA greatly diminishes overall RNA synthesis. Here we show that whereas most genes reduce their transcript levels by several fold, genes situated at less than 100 kb from the chromosomal ends (near 15% of the genome) are gradually unaffected. This positional effect denotes that chromosomal ends are topologically open, thus precluding the accumulation of DNA helical stress in telomere-proximal regions. The progressive escape from the transcription stall observed in all the chromosome extremities indicates also that friction restrictions to DNA twist diffusion, rather than tight topological boundaries, suffice to confine DNA helical tension along eukaryotic chromatin. Keywords: Time course of positive helical tension accumulation.

ORGANISM(S): Saccharomyces cerevisiae

PROVIDER: GSE18240 | GEO | 2009/12/01

SECONDARY ACCESSION(S): PRJNA123503

REPOSITORIES: GEO

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Project description:During chromosome duplication the parental DNA molecule becomes over-wound, or positively supercoiled, in the region ahead of the advancing replication fork. To allow fork progression this superhelical tension has to be removed by topoisomerases, which operate by introducing transient DNA breaks. Positive supercoiling can also be diminished if the advancing fork rotates along the DNA helix, but then sister chromatid intertwinings form in its wake. Despite these insights it remains largely unknown how replicationinduced superhelical stress is dealt with on linear, eukaryotic chromosomes. Here we show that this stress increases with the length of budding yeast chromosomes. This opens for the possibility that superhelical tension is handled on a chromosome scale and not only within topologically closed chromosomal domains as the current view predicts. We found that inhibition of type I topoisomerases leads to a late replication delay of longer, but not shorter chromosomes. This phenotype is also displayed by cells expressing mutated versions of the cohesin- and condensin–related Smc5/6 complex. The chromosomal association of the Smc5/6 complex is shown to increase in response to chromosome lengthening, chromosome circularization, or inactivation of Topoisomerase 2, all having the potential to increase the number of sister chromatid intertwinings 3. Furthermore, non-functional Smc6 reduces the accumulation of intertwined sister plasmids after one round of replication in the absence of Topoisomerase 2 function. Our results demonstrate that the length of a chromosome influences the need of superhelical tension release in Saccharomyces cerevisiae, and allow us to propose a model where the Smc5/6 complex facilitates fork rotation by sequestering nascent chromatid intertwinings which form behind the replication machinery. Smc6、Nse4, and Scc2 profiles with or without topological tension was analyzed (in top2 mutant, cells with circular chromosome, and fragmented chromosome) . 17 ChIP samples and corresponding WCE fractions have been analyzed.

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Saccharomyces cerevisiae cells: control vs positive supercoiling accumulation after 0, 30 and 120 min

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