ATP analog-sensitive Pat1 protein kinase for synchronous fission yeast meiosis at physiological temperature |
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Authors: | Lubos Cipak Randy Hyppa Gerald Smith Juraj Gregan |
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Institution: | 1.Max F. Perutz Laboratories; University of Vienna; Vienna, Austria;2.Cancer Research Institute; Slovak Academy of Sciences; Bratislava, Slovakia;3.Fred Hutchinson Cancer Research Center; Division of Basic Sciences; Seattle, WA USA |
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Abstract: | To study meiosis, synchronous cultures are often indispensable, especially for physical analyses of DNA and proteins. A temperature-sensitive allele of the Pat1 protein kinase (pat1-114) has been widely used to induce synchronous meiosis in the fission yeast Schizosaccharomyces pombe, but pat1-114-induced meiosis differs from wild-type meiosis, and some of these abnormalities might be due to higher temperature needed to inactivate the Pat1 kinase. Here, we report an ATP analog-sensitive allele of Pat1 Pat1(L95A), designated pat1-as2] that can be used to generate synchronous meiotic cultures at physiological temperature. In pat1-as2 meiosis, chromosomes segregate with higher fidelity, and spore viability is higher than in pat1-114 meiosis, although recombination is lower by a factor of 2–3 in these mutants than in starvation-induced pat1+ meiosis. Addition of the mat-Pc gene improved chromosome segregation and spore viability to nearly the level of starvation-induced meiosis. We conclude that pat1-as2
mat-Pc cells offer synchronous meiosis with most tested properties similar to those of wild-type meiosis. |
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Keywords: | analog-sensitive fission yeast meiosis Pat1 kinase synchronization |
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