Phosphorylation cues exit from mitosis The entry and exit from the cell cycle are controlled by waves of protein phosphorylation and degradation events. Fujimitsu et al. describe the precise mechanism by which the cell cycle machinery controls exit from mitosis. The critical event is activation of a ubiquitin ligase, the anaphase-promoting complex or cyclosome (APC/C). The authors used purified components and the Xenopus egg extract system to show that two subunits of APC/C were directly phosphorylated by cyclin-dependent kinase 1 (CDK1). Phosphorylation of one subunit helped recruit CDK1 for further phosphorylation of another subunit. The second subunit interacted with the APC/C activator and target of anticancer therapy known as Cdc20. Science , this issue p. 1121
Fission yeast Atf1 is a member of the ATF/CREB basic leucine zipper (bZIP) family of transcription factors with strong homology to mammalian ATF2. Atf1 regulates transcription in response to stress stimuli and also plays a role in controlling heterochromatin formation and recombination. However, its DNA binding independent role is poorly studied. Here, we report that Atf1 has a distinct role in regulating the anaphase-promoting complex/cyclosome (APC/C) ubiquitin ligase. We have identified atf1(+) as a dose-dependent suppressor of apc5-1, a mutation causing mitotic arrest. Remarkably, the suppression is not dependent upon the bZIP domain and is therefore independent of the ability of Atf1 to bind DNA. Interestingly, Atf1 physically binds the APC/C in vivo. Furthermore, we show that addition of purified Atf1 proteins into a cell-free system stimulates ubiquitylation of cyclin B and securin by the APC/C. These results reveal a novel role for Atf1 in cell cycle control through protein-protein interaction.
Expression of a human tumour-derived p53 His 273 cDNA induced growth arrest in fission yeast Schizosaccharomyces pombe. Based on the p53-induced growth arrest, we cloned an extragenic suppressor, termed tms1, by complementation. The open reading frame of the tms1 gene corresponded to a protein of 347 amino acids with a calculated mass of 37380 Da. The transcriptional start site of the tms1 gene was mapped and, in addition, the corresponding cDNA was isolated and expressed in Escherichia coli. Recombinant tms1 protein served as an antigen to produce specific polyclonal antibodies to aid identification of the tms1-gene-product in total yeast lysates. Comparison of the deduced amino acid sequence of tms1 with available databases revealed significant similarity to dehydrogenases, suggesting that the tms1 protein itself might possess dehydrogenase activity.
Homologous recombination (HR) is important for maintaining genome integrity and for the process of meiotic chromosome segregation and the generation of variation.HR is regulated throughout the cell cycle, being prevalent in the S and G 2 phases and suppressed in the G 1 phase.Here we show that the anaphasepromoting complex/cyclosome (APC/C) regulates homologous recombination in the fission yeast Schizosaccharomyces pombe by ubiquitylating Rhp54 (an ortholog of Rad54).We show that Rhp54 is a novel APC/C substrate that is destroyed in G 1 phase in a KEN-box-and Ste9/Fizzy-related manner.The biological consequences of failing to temporally regulate HR via Rhp54 degradation are seen in haploid cells only in the absence of antirecombinase Srs2 function and are more extensive in diploid cells, which become sensitive to a range of DNA-damaging agents, including hydroxyurea, methyl methanesulfonate, bleomycin, and UV.During meiosis, expression of nondegradable Rhp54 inhibits interhomolog recombination and stimulates sister chromatid recombination.We thus propose that it is critical to control levels of Rhp54 in G 1 to suppress HR repair of double-strand breaks and during meiosis to coordinate interhomolog recombination.
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