Many eukaryotic cell types undergo size-dependent cell cycle transitions controlled by the ubiquitous cyclin-dependent kinase Cdk1 (refs 1-4). The proteins that control Cdk1 activity are well described but their links with mechanisms monitoring cell size remain elusive. In the fission yeast Schizosaccharomyces pombe, cells enter mitosis and divide at a defined and reproducible size owing to the regulated activity of Cdk1 (refs 2, 3). Here we show that the cell polarity protein kinase Pom1, which localizes to cell ends, regulates a signalling network that contributes to the control of mitotic entry. This network is located at cortical nodes in the middle of interphase cells, and these nodes contain the Cdk1 inhibitor Wee1, the Wee1-inhibitory kinases Cdr1 (also known as Nim1) and Cdr2, and the anillin-like protein Mid1. Cdr2 establishes the hierarchical localization of other proteins in the nodes, and receives negative regulatory signals from Pom1. Pom1 forms a polar gradient extending from the cell ends towards the cell middle and acts as a dose-dependent inhibitor of mitotic entry, working through the Cdr2 pathway. As cells elongate, Pom1 levels decrease at the cell middle, leading to mitotic entry. We propose that the Pom1 polar gradient and the medial cortical nodes generate information about cell size and coordinate this with mitotic entry by regulating Cdk1 through Pom1, Cdr2, Cdr1 and Wee1.

mid1 is required for the proper placement of the contractile actin ring for cytokinesis at a medial site overlying the nucleus. Here we find that mid1 protein (mid1p) shuttles between the nucleus and a cortical medial broad band during interphase and early mitosis. The position of this broad band, which overlies the nucleus, is linked to nuclear position even in cells with displaced or multiple nuclei. We identified and created mutations in an NLS and in two crm1-dependent NES sequences in mid1p. NES mutations caused mid1p accumulation in the nucleus and loss of function. An NLS mutations greatly reduced nuclear localization but did not perturb cytoplasmic localization or function. mid1p localization to the medial broad band was also not dependent on mid1p PH domain or microtubule and actin cytoskeletons. Overexpression of mid1p produced ectopic cell growth at this band during interphase and abnormal karmellae-like nuclear membrane structures. In plo1-1, mid1p formed a medial broad band but did not incorporate into a tight ring, suggesting that polo kinase plo1p is required for activation of mid1p function. Thus, the mid1p broad band defines a compartment at the medial cell surface, whose localization is linked to the position of the nucleus, and whose function may be to position the plane of cell division.

Proper microtubule organization is essential for cellular processes such as organelle positioning during interphase and spindle formation during mitosis. The fission yeast Schizosaccharomyces pombe presents a good model for understanding microtubule organization. We identify fission yeast ase1p, a member of the conserved ASE1/PRC1/MAP65 family of microtubule bundling proteins, which functions in organizing the spindle midzone during mitosis. Using fluorescence live cell imaging, we show that ase1p localizes to sites of microtubule overlaps associated with microtubule organizing centers at both interphase and mitosis. ase1Delta mutants fail to form overlapping antiparallel microtubule bundles, leading to interphase nuclear positioning defects, and premature mitotic spindle collapse. FRAP analysis revealed that interphase ase1p at overlapping microtubule minus ends is highly dynamic. In contrast, mitotic ase1p at microtubule plus ends at the spindle midzone is more stable. We propose that ase1p functions to organize microtubules into overlapping antiparallel bundles both in interphase and mitosis and that ase1p may be differentially regulated through the cell cycle.