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A number of important molecular checkpoints are believed to control the orderly progression of cell cycle events. We have found that the radiation-sensitive Schizosaccharomyces pombe mutant rad3-136 is deficient in two molecular checkpoint functions. Unlike wild-type cells, the mutant cells are unable to arrest in the G2 phase of the cell cycle after DNA damage by -irradiation and are also incapable of maintaining the dependence of mitosis upon the completion of DNA synthesis. An S. pombe genomic clone that complements the UV sensitivity of the rad3-136 mutant completely restores the missing checkpoint functions. The rad3+ gene is also likely to play a role in DNA repair.In recent years major advances have been made in our understanding of the proteins involved in the entry into mitosis in eukaryotic cells (for reviews, see refs. 1-6). However, the regulatory circuits that modulate mitosis are less well-characterized. A variety ofcrucial molecular checkpoint functions ensure that entry into and exit from mitosis occur only after the execution of an orderly progression of events (7). Among such checkpoints are functions that (i) delay mitosis or arrest cells in the G2 phase of the cell cycle in response to DNA damage (8-11), (ii) maintain the dependence of mitosis on the completion of chromosome replication (12,13), and (iii) make the exit from' mitosis dependent upon the completion of spindle assembly and proper chromosome segregation (14, 15). Cells inflicted with DNA damage delay progression into mitosis by arresting the cell cycle temporarily in G2 so that DNA repair can occur (7,16,17). In the budding yeast Saccharomyces cerevisiae (Sa. cerevisiae), the RAD9gene is involved in G2 arrest and the rad9 mutant is sensitive to DNA damage because cells containing damaged DNA fail to arrest (9, 11). Several other mitotic entry checkpoint (mec) mutants have been identified in Sa. cerevisiae (10).The checkpoint controlling the coupling of mitosis to DNA synthesis is abolished in Schizosaccharomyces pombe (Sc. pombe) by overproduction of the phosphatase cdc25, which dephosphorylates the cdc2 kinase and activates mitosis (18)(19)(20)(21). Other genes involved in this checkpoint pathway include pimi (a homolog of the human RCCI gene), mutation of which causes premature initiation of mitosis, and spil (a suppressor of pimi), which encodes a ras-like GTPase homologous to human TC4 (22).The rad3-136 mutant of the fission yeast, Sc. pombe, is sensitive to UV-and y-radiation, refractory to caffeine sensitization after exposure to UV-radiation, and exhibits a reduced frequency of UV-induced forward mutation when compared to wild-type cells (23,24). We show in this paper that the mutant rad3-136 is deficient in two checkpoint functions, G2 arrest after y-irradiation and the coupling of mitosis to DNA synthesis. We also show that a Sc. pombe genomic clone that restores resistance to UV-and -y-radiation completely restores the missing checkpoint functions when reintroduced into the rad3-136 mutant. There also appears to ...

CENP-meta has been identified as an essential, kinesin-like motor protein in Drosophila. The 257-kD CENP-meta protein is most similar to the vertebrate kinetochore-associated kinesin-like protein CENP-E, and like CENP-E, is shown to be a component of centromeric/kinetochore regions of Drosophila chromosomes. However, unlike CENP-E, which leaves the centromere/kinetochore region at the end of anaphase A, the CENP-meta protein remains associated with the centromeric/kinetochore region of the chromosome during all stages of the Drosophila cell cycle. P-element–mediated disruption of the CENP-meta gene leads to late larval/pupal stage lethality with incomplete chromosome alignment at metaphase. Complete removal of CENP-meta from the female germline leads to lethality in early embryos resulting from defects in metaphase chromosome alignment. Real-time imaging of these mutants with GFP-labeled chromosomes demonstrates that CENP-meta is required for the maintenance of chromosomes at the metaphase plate, demonstrating that the functions required to establish and maintain chromosome congression have distinguishable requirements.