Centrosomal MPF triggers the mitotic and morphogenetic switches of fission yeast

Abstract

Activation of mitosis promoting factor (MPF) drives mitotic commitment1. In human cells active MPF appears first on centrosomes2. We show that local activation of MPF on the equivalent organelle of fission yeast, the spindle pole body (SPB), promotes Polo kinase activity at the SPBs long before global MPF activation drives mitotic commitment. Artificially promoting MPF or Polo activity at various locations revealed that this local control of Plo1 activity on G2 phase SPBs dictates the timing of mitotic commitment. Cytokinesis of the rod shaped fission yeast cell generates a naïve “new” cell end. Growth is restricted to the experienced old end until a point in G2 phase called “New End Take Off” (NETO) when bipolar growth is triggered3. NETO coincided with MPF activation of Plo1 on G2 phase SPBs4. Both MPF and Polo activities were required for NETO and both induced NETO when ectopically activated at interphase SPBs. NETO promotion by MPF required polo. Thus, local MPF activation on G2 SPBs directs polo kinase to control at least two distinct and temporally separated, cell cycle transitions at remote locations.

The Centrosome and Its Duplication Cycle

“…This defect stemmed from deficient Polo Plo1 -based feedback controls because a simple elevation of Cdc25 levels activated the new SPB to drive cut12.1 cells through a functional mitosis (Tallada et al 2007(Tallada et al , 2009). This evidence for spindle pole -driven mitotic commitment was consolidated by the ability of ectopic activation of either Polo Plo1 or Cdk1 at interphase SPBs to inappropriately promote mitotic commitment (Grallert et al 2013b). These data categorically establish that events on the spindle pole form an integral part of the switch that regulates mitotic commitment.…”

Analysis ofS. pombeSIN protein SPB-association reveals two genetically separable states of the SIN

“…The spindle pole body (SPB) serves as a microtubule-organising centre, and coordination point for cell cycle regulators (Grallert et al, 2012;Grallert et al, 2013;Hagan, 2008). Association of SIN proteins to the SPB plays an important part in the regulation of the SIN (reviewed by Johnson et al, 2012;Simanis, 2003).…”

Live Cell Imaging in Fission Yeast

“…In contrast, because only the first generation of cells will adhere to a lectin-coated surface and daughters float away from the focal plane in this approach, the rapid media exchange around static cells in microfluidic systems makes them ideal for drug exchange during observation. However, if microfluidics are not available, drug exchange can be achieved equally well using a simple Pasteur pipette-mediated exchange of media within a glass-bottomed dish to which cells have been attached using a lectin (Grallert et al 2013b). A further alternative to microfluidics for long-term imaging over several cell cycles is to trap cells under a thick agarose/medium pad on an oxygen-permeable transparent membrane such as the Lumox culture hydrophilic dish ( Fig.…”