Dataset Information

Immediate visualization of recombination events and chromosome segregation defects in fission yeast meiosis.

ABSTRACT: Schizosaccharomyces pombe, also known as fission yeast, is an established model for studying chromosome biological processes. Over the years, research employing fission yeast has made important contributions to our knowledge about chromosome segregation during meiosis, as well as meiotic recombination and its regulation. Quantification of meiotic recombination frequency is not a straightforward undertaking, either requiring viable progeny for a genetic plating assay, or relying on laborious Southern blot analysis of recombination intermediates. Neither of these methods lends itself to high-throughput screens to identify novel meiotic factors. Here, we establish visual assays novel to Sz. pombe for characterizing chromosome segregation and meiotic recombination phenotypes. Genes expressing red, yellow, and/or cyan fluorophores from spore-autonomous promoters have been integrated into the fission yeast genomes, either close to the centromere of chromosome 1 to monitor chromosome segregation, or on the arm of chromosome 3 to form a genetic interval at which recombination frequency can be determined. The visual recombination assay allows straightforward and immediate assessment of the genetic outcome of a single meiosis by epi-fluorescence microscopy without requiring tetrad dissection. We also demonstrate that the recombination frequency analysis can be automatized by utilizing imaging flow cytometry to enable high-throughput screens. These assays have several advantages over traditional methods for analyzing meiotic phenotypes.

SUBMITTER: Li D

PROVIDER: S-EPMC6823302 | biostudies-literature | 2019 Sep

REPOSITORIES: biostudies-literature

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Immediate visualization of recombination events and chromosome segregation defects in fission yeast meiosis.

Li Dmitriy D Roca Marianne M Yuecel Raif R Lorenz Alexander A

Chromosoma 20190209 3

Schizosaccharomyces pombe, also known as fission yeast, is an established model for studying chromosome biological processes. Over the years, research employing fission yeast has made important contributions to our knowledge about chromosome segregation during meiosis, as well as meiotic recombination and its regulation. Quantification of meiotic recombination frequency is not a straightforward undertaking, either requiring viable progeny for a genetic plating assay, or relying on laborious Sout ...[more]

PMID: 30739171

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Project description:CDKs (cyclin-dependent kinases) associate with different cyclins to form different CDK-complexes that are fundamental for an ordered cell cycle progression, and the coordination of this progression with different aspects of the cellular physiology. During meiosis programmed DNA double-strand breaks (DSBs) initiate recombination that in addition to generating genetic variability are essential for the reductional chromosome segregation during the first meiotic division, and therefore for genome stability and viability of the gametes. However, how meiotic progression and DSB formation are coordinated, and the role CDKs have in the process, is not well understood. We have used single and double cyclin deletion mutants, and chemical inhibition of global CDK activity using the cdc2-asM17 allele, to address the requirement of CDK activity for DSB formation and recombination in fission yeast. We report that several cyclins (Cig1, Cig2, and the meiosis-specific Crs1) control DSB formation and recombination, with a major contribution of Crs1. Moreover, complementation analysis indicates specificity at least for this cyclin, suggesting that different CDK complexes might act in different pathways to promote recombination. Down-regulation of CDK activity impinges on the formation of linear elements (LinEs, protein complexes required for break formation at most DSB hotspot sites). This defect correlates with a reduction in the capability of one structural component (Rec25) to bind chromatin, suggesting a molecular mechanism by which CDK controls break formation. However, reduction in DSB formation in cyclin deletion mutants does not always correspondingly correlate with a proportional reduction in meiotic recombination (crossovers), suggesting that specific CDK complexes might also control downstream events balancing repair pathways. Therefore, our work points to CDK regulation of DSB formation as a key conserved feature in the initiation of meiotic recombination, in addition to provide a view of possible roles CDK might have in other steps of the recombination process.

Dataset Information

Immediate visualization of recombination events and chromosome segregation defects in fission yeast meiosis.

Publications

Immediate visualization of recombination events and chromosome segregation defects in fission yeast meiosis.

Similar Datasets

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