Castillon, G. A., N. R. Adames, C. H. Rosello, H. S. Seidel, M. S. Longtine, J. A. Cooper & R. A. Heil-Chapdelaine. 2003. Septins have a dual role in controlling mitotic exit in budding yeast. Curr Biol. 13:654-658. PubMed

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Abstract. In Saccharomyces cerevisiae, the spindle position checkpoint ensures that cells do not exit mitosis until the mitotic spindle moves into the mother/bud neck and thus guarantees that each cell receives one nucleus. Mitotic exit is controlled by the small G protein Tem1p. Tem1p and its GTPase activating protein (GAP) Bub2p/Bfa1p are located on the daughter-bound spindle pole body. The GEF Lte1p is located in the bud. This segregation helps keep Tem1p in its inactive GDP state until the spindle enters the neck. However, the checkpoint functions without Lte1p and apparently senses cytoplasmic microtubules in the mother/bud neck. To investigate this mechanism, we examined mutants defective for septins, which compose a ring at the neck. We found that the septin mutants sep7∆ and cdc10∆ are defective in the checkpoint. When movement of the spindle into the neck was delayed, mitotic exit occurred, inappropriately leaving both nuclei in the mother. In sep7∆ and cdc10∆ mutants, Lte1p is mislocalized to the mother. In sep7∆, but not cdc10∆, mutants, inappropriate mitotic exit depends on Lte1p. These results suggest that septins serve as a diffusion barrier for Lte1p, and that Cdc10p is needed for the septin ring to serve as a scaffold for a putative microtubule sensor.

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Movie 1: Time-lapse of Lte1-3GFP in wild type cell. Wide field epifluorescence image collected 1 frame every 2 seconds for 98 seconds. Movie plays two times at 8 fps. Focus on surface of bud.
Movie 2: Time-lapse of wt cell with Cdc3-GFP. Images collected 1 frame every 2 minutes for 66 minutes. Each image is a two dimensional projection of 12 focal planes 0.45µm apart. Movie plays two times.
Movie 3: 3D rotation of wt cell expressing Cdc3-GFP. Shown is nascent septin ring forming adjacent to old septin ring.
Movie 4: 3D rotation of wt cell expressing Cdc3-GFP. Shown is broadening septin ring in neck of small bud.
Movie 5: 3D rotation of wt cell expressing Cdc3-GFP. Shown is hourglass shaped septin ring in neck of large bud.
Movie 6: 3D rotation of 2 wt cells expressing Cdc3-GFP. Shown is the remaining single septin ring in each cell after cell separation.
Movie 7: 3D rotation of sep7∆ cell expressing Cdc3-GFP. Shown is nascent septin ring forming at the nascent bud site of the mother cell. Also shown is remaining ring in daughter cell.
Movie 8: 3D rotation of sep7∆ cell expressing Cdc3-GFP. Shown is hourglass shaped septin ring with longitudinal fissures or breaks in neck of large bud.
Movie 9: 3D rotation of sep7∆ cell expressing Cdc3-GFP. Shown is fissured septin ring during cytokinesis. Fragments of the septin ring are seen in mother and bud.
Movie 10: 3D rotation of connected sep7∆ gin4∆ cells expressing Cdc3-GFP. First shown is nascent septin ring. Then, arrows mark regions of circumferential breaks in the septin stripes within neck – likely regions of attempted cytokinesis.
Movie 11: 3D rotation of sep7∆ gin4∆ cells expressing Cdc3-GFP. Shown are stripes of septins within the neck of a small bud.
Movie 12: 3D rotation of sep7∆ gin4∆ cells expressing Cdc3-GFP. Shown are stripes of septins in large budded cell. Stripes are displaced into the bud. A small amount of Cdc3-GFP is seen in mother.
Movie 13: 3D rotation of cdc10∆ cell expressing Cdc3-GFP. Shown is nascent septin ring of uneven thickness forming at the nascent bud site of the mother cell.
Movie 14: 3D rotation of cdc10∆ cell expressing Cdc3-GFP. Shown is septin ring of uneven thickness in broad neck of small budded cell.
Movie 15: 3D rotation of cdc10∆ cell expressing Cdc3-GFP. Shown is septin ring of uneven thickness displaced from broad neck into the bud of large budded cell.
Movie 16: Time-lapse of cdc10∆ cell with Cdc3-GFP. Images collected 1 frame every 2 minutes for 76 minutes. Each image is a two dimensional projection of 12 focal planes 0.45µm apart. Movie plays two times.