Different levels of Bfa1/Bub2 GAP activity are required to prevent mitotic exit of budding yeast depending on the type of perturbations.
In budding yeast, Tem1 is a key regulator of mitotic exit. Bfa1/Bub2 stimulates Tem1 GTPase activity as a GTPase-activating protein (GAP). Lte1 possesses a guanine-nucleotide exchange factor (GEF) domain likely for Tem1. However, recent observations showed that cells may control mitotic exit without either Lte1 or Bfa1/Bub2 GAP activity, obscuring how Tem1 is regulated. Here, we assayed BFA1 mutants with varying GAP activities for Tem1, showing for the first time that Bfa1/Bub2 GAP activity inhibits Tem1 in vivo. A decrease in GAP activity allowed cells to bypass mitotic exit defects. Interestingly, different levels of GAP activity were required to prevent mitotic exit depending on the type of perturbation. Although essential, more Bfa1/Bub2 GAP activity was needed for spindle damage than for DNA damage to fully activate the checkpoint. Conversely, Bfa1/Bub2 GAP activity was insufficient to delay mitotic exit in cells with misoriented spindles. Instead, decreased interaction of Bfa1 with Kin4 was observed in BFA1 mutant cells with a defective spindle position checkpoint. These findings demonstrate that there is a GAP-independent surveillance mechanism of Bfa1/Bub2, which, together with the GTP/GDP switch of Tem1, may be required for the genomic stability of cells with misaligned spindles.
Kim
Junwon
J
Department of Biochemistry and Institute of Life Science and Biotechnology, Yonsei University, Seoul 120-749, Korea.
Jang
Selma Sun
SS
Song
Kiwon
K
D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000502:physiology*; D003598:Cytoskeletal Proteins / Q000502:physiology*; D005434:Flow Cytometry; D020558:GTP Phosphohydrolases / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D008938:Mitosis; D008954:Models, Biological; D008957:Models, Genetic; D016296:Mutagenesis; D009154:Mutation; D010750:Phosphoproteins / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism / Q000502:physiology*; D004718:Saccharomycetales / Q000502:physiology*; D008941:Spindle Apparatus; D020798:Two-Hybrid System Techniques
C119117:BFA1 protein, S cerevisiae; C070351:BUB2 protein, S cerevisiae; D018797:Cell Cycle Proteins; D003598:Cytoskeletal Proteins; D010750:Phosphoproteins; D029701:Saccharomyces cerevisiae Proteins; C490209:KIN1 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; D020558:GTP Phosphohydrolases