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First published online September 26, 2003
doi: 10.1242/10.1242/jcs.00807

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Events at the end of mitosis in the budding and fission yeasts

Cell Cycle Control Laboratory, Swiss Institute for Experimental Cancer Research (ISREC), 1066 Epalinges, Switzerland

View larger version (41K): [in a new window]   Fig. 1. The effects of altering SIN and MEN activity. Panel A shows wild-type S. pombe cells stained with DAPI (to reveal the DNA) and Calcofluor (to reveal the position of the division septum). Panel B shows cells in which SIN signaling is defective. Note that cells become elongated and multinucleated, as nuclear division continues in the absence of cytokinesis. Panel C shows the effects of increased SIN activity. Note that the cells undergo multiple rounds of septum formation without cell cleavage. The red cross indicates that loss-of-function mutations produce the indicated phenotype. The bold black type `OP' indicates that increased expression of a gene produces the indicated phenotype. Scale bar, 10 µm. Panel D shows an arrested S. cerevisiae cdc15 mutant. The cells are stained with DAPI (blue) and antibody to tubulin (red). Note that the chromosomes are separated, and that the spindle is highly elongated. This image is provided by Rosella Visintin and Angelika Amon (MIT, Cambridge, MA). Panel E shows the mutant dbf2-2. The phosphoprotein phosphatase has been tagged with GFP. Note that in the arrested cells, Cdc14p remains in the nucleolus. This image is provided by Elmar Schiebel (Patterson Institute for Cancer Research, Christie Hospital, Manchester, UK).

View larger version (32K): [in a new window]   Fig. 2. Localization of SIN proteins. The figure shows schematically the localization of SIN proteins and some of the regulators through the cell cycle. The data are taken from references cited in the text. The colored cutout within the nucleus in the localization sequence for S. pombe flp1 represents the nucleolus. In the S. pombe cells, the grey horizontal bar within them represents the mitotic spindle, while the short bar at the cell center represents the position of the contractile ring. The localization of Par1p is similar to that of Mob1p, whereas Scw1p is cytoplasmic throughout the cell cycle. There is also a significant cytoplasmic pool of Mob1p and Par1p that is not indicated, for reasons of clarity. It is noteworthy that signal intensity of some of the proteins at the spindle pole body varies significantly through the cell cycle. To convey this, the intensity has been assigned on a arbitrary scale of 1 (present, but very faint) to 5 (present, strongest signals observed). The absolute signal strengths vary from protein to protein, and so the numbers give no indication of the amounts of protein relative to each other.

View larger version (27K): [in a new window]   Fig. 3. How the SIN is wired. This figure is a representation of how the principal components of the SIN might interact. This is based upon localization studies, and the activity of proteins in different mutant backgrounds. For the sake of simplicity, the signal transduction event is portrayed as being linear, although there is no biochemical evidence to rule out a more complex arrangement, such as branching at the level of Spg1p. With the exception of the action of Byr4p-Cdc16p on Spg1p, direct regulation of one protein by another in vitro has not been demonstrated. Green arrows indicate septum-promoting events. Orange ball and stick symbols indicate the presumed point of action of negative regulators of SIN signaling. Inhibitors of SIN signaling are indicated in red, protein kinases in yellow, and phosphoprotein phosphatases in mauve. The roles and targets of Flp1p, Etd1p and Pld6p are unclear. Plo1p acts at or near the top of the network but its target has not been identified. Spg1p is shown twice, since it can form at least two complexes on the spindle pole body, one with Cdc7p, and the other with Byr4p-Cdc16p. Note that it is likely that Plo1p interacts with other spindle pole body components in addition to Cut12p. The large gray box represents the spindle pole body. The Mob1p-Sid2p complex is shown twice, once on the spindle pole body and once at the cell cortex, associated with the contractile actin ring. Zfs1 and Scw1 are shown downstream of the main SIN signaling module; their point of action is unclear.

View larger version (31K): [in a new window]   Fig. 4. How the MEN is wired. The basic interactions among components of the MEN and some of its regulators are shown. Orange ball and stick symbols indicate inhibitory events. Green arrows indicate activating events. Purple arrows indicate substrates of Cdc14p. In all the cases indicated (except Lte1p, where this has not been assessed), the protein is activated to perform its function as a result of dephosphorylation by Cdc14p. The large purple arrows indicate that events are brought about by the indicated proteins, while the black arrow represents release of Cdc14p from the nucleolus. The grey ovals indicate functional groups of proteins. The gray box is meant to represent the Nud1p-based scaffold, upon which the MEN signaling proteins assemble at the spindle pole body. As in Fig. 3, red boxes indicate inhibitors of MEN components, and protein kinases are in yellow boxes. Panel A shows the state of the regulators of mitotic exit in early mitosis, before all the chromosomes have attached to the spindle. The spindle checkpoint blocks both the degradation of Cdc28p-CLB and activation of the FEAR proteins. The phosphoprotein phosphatase Cdc14p is sequestered in the nucleolus, and activated Cdc28p-CLB inhibits formation of APC/CCdh1p and activation of Cdc15p. Panel B shows the state of the regulators of mitotic exit during anaphase. The FEAR proteins release Cdc14p from the nucleolus. Cdc5p also inhibits Bfa1p, allowing Tem1p to accumulate in the GTP-bound form. One of the substrates of Cdc14p is Cdc15p, activating MEN signaling. This keeps Cdc14p out of the nucleolus and activates Dbf2p-Mob1p, which bring about cytokinesis and help maintain Cdc14p outside the nucleolus. Cdc14p increases the level of the CKI Sic1p, both by increasing its expression via Swi5p and by preventing degradation of Sic1p. It also targets Cdh1p, allowing formation of APC/CCdh1p to promote mitotic cyclin destruction during the M-G1 transition. Further details are given in the text. Panel C shows how this self perpetuating mitotic exit circuit is broken to allow B-type cyclins to accumulate in preparation for S-phase. Cdc14 promotes expression of Amn1p (see text for details), which directly interferes with the association of Cdc15p and Tem1p. Another target of Cdc14p is Bfa1p, which reactivates it, thereby promoting GTP hydrolysis by Tem1p and inactivation of signaling. Finally, the FEAR proteins Spo12p and Cdc5p are targets of APC/CCdh1p, which decreases FEAR activity. The reduction of MEN and FEAR activity is presumed to result in the return of Cdc14p to the nucleolus, although how this is achieved is not clear. Finally, Amn1p degradation is promoted through phosphorylation by activated Cdc28p-CLN, which allows Tem1p-Cdc15p interaction to occur later in the cycle.

View larger version (30K): [in a new window]   Fig. 5. Localization of MEN proteins. The localization of the principal components of the MEN is shown, following the style of Pereira and Schiebel (Pereira and Schiebel, 2001). Since different studies do not always agree regarding the intensity of different protein signals, this is not indicated.